Bio

Clinical Focus


  • Pediatric Pulmonary
  • Pulmonary Medicine/Cystic Fibrosis, Pediatric

Academic Appointments


Administrative Appointments


  • Chairman of Pediatrics, Stanford University (2008 - Present)
  • Physician in Chief, Lucile Packard Children's Hospital (2008 - Present)
  • Director, Child Health Research Institute at Stanford, Stanford University (2011 - Present)
  • Paediatrician in Chief, Hospital for Sick Children (SickKids) (1996 - 2006)
  • Chairman of Pediatrics, University of Toronto (1996 - 2006)

Honors & Awards


  • President, Fleischner Society (2007)
  • Fellow, Canadian Academy of Health Sciences (2006)
  • Edwin L Kendig Award, American Academy of Pediatrics/American College of Chest Physicians (2005)
  • Inaugural President, Pediatric Chairs of Canada (2002-2004)
  • Scientific Accomplishment Award, American Thoracic Society (2001)

Professional Education


  • Board Certification: Pediatric Pulmonary, American Board of Pediatrics (1986)
  • Fellowship:Columbia University College of Physicians and Surgeons (1981) NY
  • Board Certification: Pediatrics, American Board of Pediatrics (1980)
  • Residency:University of Manitoba (1978) Canada
  • Residency:University of Manitoba (1977) Canada
  • Internship:University of Manitoba (1976) Canada
  • Medical Education:University of Manitoba (1975) Canada
  • FRCPC, Royal Coll Phys & Surg, Paediatric Respiratory Medicine (1989)
  • FRCPC, Royal College Phys & Surg, Paediatrics (1980)
  • MD, University of Manitoba, Medicine (1975)
  • BSc, University of Manitoba, Science (1971)

Research & Scholarship

Current Research and Scholarly Interests


We wish to develop new therapies to prevent and treat the respiratory distress syndrome (RDS) and its sequelae, bronchopulmonary dysplasia (BPD). In the pursuit of this goal my laboratory has conducted research on how the lung’s airspaces become fluid filled (mechanisms of pulmonary edema) and how airspace fluid is cleared under both physiologic (fetal lung liquid at birth) and pathophysiologic (pulmonary edema) conditions. Since the rate limiting step in airspace fluid clearance is the active transport of Na+ by distal lung epithelium, our studies had investigated the effect of lung maturity, mediators and hormones on the pre and post-translational regulation of the epithelial Na+ channel (ENaC). My current research involves a population based study to discover the genetic influences on the development of BPD.

Teaching

2013-14 Courses


Publications

Journal Articles


  • A Genome-Wide Association Study (GWAS) for Bronchopulmonary Dysplasia. Pediatrics Wang, H., St Julien, K. R., Stevenson, D. K., Hoffmann, T. J., Witte, J. S., Lazzeroni, L. C., Krasnow, M. A., Quaintance, C. C., Oehlert, J. W., Jelliffe-Pawlowski, L. L., Gould, J. B., Shaw, G. M., O'Brodovich, H. M. 2013; 132 (2): 290-297

    Abstract

    Twin studies suggest that heritability of moderate-severe bronchopulmonary dysplasia (BPD) is 53% to 79%, we conducted a genome-wide association study (GWAS) to identify genetic variants associated with the risk for BPD.The discovery GWAS was completed on 1726 very low birth weight infants (gestational age = 25(0)-29(6/7) weeks) who had a minimum of 3 days of intermittent positive pressure ventilation and were in the hospital at 36 weeks' postmenstrual age. At 36 weeks' postmenstrual age, moderate-severe BPD cases (n = 899) were defined as requiring continuous supplemental oxygen, whereas controls (n = 827) inhaled room air. An additional 795 comparable infants (371 cases, 424 controls) were a replication population. Genomic DNA from case and control newborn screening bloodspots was used for the GWAS. The replication study interrogated single-nucleotide polymorphisms (SNPs) identified in the discovery GWAS and those within the HumanExome beadchip.Genotyping using genomic DNA was successful. We did not identify SNPs associated with BPD at the genome-wide significance level (5 × 10(-8)) and no SNP identified in previous studies reached statistical significance (Bonferroni-corrected P value threshold .0018). Pathway analyses were not informative.We did not identify genomic loci or pathways that account for the previously described heritability for BPD. Potential explanations include causal mutations that are genetic variants and were not assayed or are mapped to many distributed loci, inadequate sample size, race ethnicity of our study population, or case-control differences investigated are not attributable to underlying common genetic variation.

    View details for DOI 10.1542/peds.2013-0533

    View details for PubMedID 23897914

  • Progress in understanding the genetics of bronchopulmonary dysplasia SEMINARS IN PERINATOLOGY Shaw, G. M., O'Brodovich, H. M. 2013; 37 (2): 85-93

    Abstract

    Bronchopulmonary dysplasia (BPD) is the most common chronic lung disease in infants. Its treatment imposes considerable healthcare burden and costs in the perinatal and early childhood period and patients are usually left with lifelong deficits in lung function. Evidence exists for different pathophysiologic pathways that can promote the structural changes that characterize BPD, including the impairment in alveolarization; however, there is increasing interest regarding heritable factors that may predispose very low birth weight infants to BPD. Our review focuses on recent publications that have investigated genetic factors that may potentially contribute to such reported heritability. These publications point us toward some possible genomic candidates for further study, but certainly do not identify any particular gene or gene pathway that would be inferred to be contributing substantially to the underlying etiology of BPD.

    View details for DOI 10.1053/j.semperi.2013.01.004

    View details for Web of Science ID 000317870800005

    View details for PubMedID 23582962

  • High Quality Genome-Wide Genotyping from Archived Dried Blood Spots without DNA Amplification. PLoS One St Julien KR, Jelliffe-Pawlowski LL, Shaw GM, Stevenson DK, O'Brodovich HM, Krasnow MA, Stanford BPD Study Group (Gould JB, Oehlert JW, Quaintance C, Wang H, Lazzeroni L, Hoffman T, Witte J, Lorey F) 2013; 8 (5): e4710

    View details for PubMedID 23737996

  • Risk of bronchopulmonary dysplasia by second trimester maternal serum levels of alpha-fetoprotein, human chorionic gonadotrophin and unconjugaged Estriol Pediatric Research Jelliffe-Pawlowski LL, Shaw G, Stevenson D, Oehlert JW, Quaintance C, Santos AJ, Baer RJ, Currier RJ, O'Brodovich H, Gould JB 2012; 71 (4): 399-406
  • Lung-derived soluble mediators are pathogenic in ventilator-induced lung injury AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Jaecklin, T., Engelberts, D., Otulakowski, G., O'Brodovich, H., Post, M., Kavanagh, B. P. 2011; 300 (4): L648-L658

    Abstract

    Ventilator-induced lung injury (VILI) due to high tidal volume (V(T)) is associated with increased levels of circulating factors that may contribute to, or be markers of, injury. This study investigated if exclusively lung-derived circulating factors produced during high V(T) ventilation can cause or worsen VILI. In isolated perfused mouse lungs, recirculation of perfusate worsened injury (compliance impairment, microvascular permeability, edema) induced by high V(T). Perfusate collected from lungs ventilated with high V(T) and used to perfuse lungs ventilated with low V(T) caused similar compliance impairment and permeability and caused a dose-dependent decrease in transepithelial electrical resistance (TER) across rat distal lung epithelial monolayers. Circulating soluble factors derived from the isolated lung thus contributed to VILI and had deleterious effects on the lung epithelial barrier. These data demonstrate transferability of an injury initially caused exclusively by mechanical ventilation and provides novel evidence for the biotrauma hypothesis in VILI. Mediators of the TER decrease were heat-sensitive, transferable via Folch extraction, and (following ultrafiltration, 3 kDa) comprised both smaller and larger molecules. Although several classes of candidate mediators, including protein cytokines (e.g., tumor necrosis factor-?, interleukin-6, macrophage inflammation protein-1?) and lipids (e.g., eicosanoids, ceramides, sphingolipids), have been implicated in VILI, only prostanoids accumulated in the perfusate in a pattern consistent with a pathogenic role, yet cyclooxygenase inhibition did not protect against injury. Although no single class of factor appears solely responsible for the decrease in barrier function, the current data implicate lipid-soluble protein-bound molecules as not just markers but pathogenic mediators in VILI.

    View details for Web of Science ID 000288943900017

    View details for PubMedID 21239530

  • Lung-derived soluble mediators are pathogenic in ventilator-induced lung injury Am J Physiol Lung Cell Mol Physiol Jaecklin T, Engelberts D, Otulakowski G, O'Brodovich H, Post M, Kavanagh BP 2011; 300: L648-L658
  • Long-term terbutaline exposure stimulates alpha(1)-Na+K+-ATPase expression at posttranscriptional level in rat fetal distal lung epithelial cells AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Rahman, M. S., Gandhi, S., Otulakowski, G., Duan, W., Sarangapani, A., O'Brodovich, H. 2010; 298 (1): L96-L104

    Abstract

    Transepithelial Na(+) transport through epithelial Na(+) channels (ENaC) on the apical membrane and Na(+)-K(+)-ATPase activity on the basolateral membrane of distal lung epithelial cells are critical for alveolar fluid clearance. Acute exposure to beta-adrenergic agonists stimulates lung fluid clearance by increasing Na(+) transport. We investigated the effects of chronic exposure to the beta(2)-adrenergic agonist terbutaline on the transepithelial Na(+) transport in rat fetal distal lung epithelia (FDLE). FDLE monolayers exposed to 10(-4) M terbutaline for 48 h had significantly increased propanolol-blockable transepithelial total and amiloride-sensitive short-circuit current (I(sc)); however, when these chronically exposed monolayers were acutely exposed to additional beta-agonists and intracellular cAMP upregulators, there was no further increase in I(sc). Monolayers exposed to terbutaline for >48 h had I(sc) similar to control cells. Ouabain-sensitive Na(+)-K(+)-ATPase activity was increased in 48-h terbutaline-exposed FDLE whose apical membranes were permeabilized with nystatin. In contrast, terbutaline did not increase amiloride-sensitive apical membrane I(sc) in FDLE whose basolateral membranes were permeabilized with nystatin. Terbutaline treatment did not affect alpha-, beta-, or gamma-ENaC mRNA or alpha-ENaC protein steady-state levels, but increased total cellular levels and rate of synthesis of alpha(1)-Na(+)-K(+)-ATPase protein in FDLE in the absence of any change in alpha(1)-Na(+)-K(+)-ATPase mRNA. Total cellular beta(1)-Na(+)-K(+)-ATPase mRNA and protein levels were not affected by terbutaline. These data suggest that FDLE have different responses from adult type II epithelial cells when chronically exposed to terbutaline, and their increased transepithelial Na(+) transport occurs via a posttranscriptional increase in alpha(1)-Na(+)-K(+)-ATPase expression.

    View details for DOI 10.1152/ajplung.00158.2009

    View details for Web of Science ID 000272827900012

    View details for PubMedID 19880505

  • NHLBI Training Workshop Report: The Vanishing Pediatric Pulmonary Investigator and Recommendations for Recovery PEDIATRIC PULMONOLOGY Ferkol, T., Zeitlin, P., Abman, S., Blaisdell, C. J., O'Brodovich, H. 2010; 45 (1): 25-33

    Abstract

    The adequacy of the pipeline of advanced pulmonary fellows to supply appropriately trained and committed researchers to enter academic careers was the major topic of a recently held National Heart Lung and Blood Institute NHLBI Workshop: Respiratory Medicine-Related Research Training for Adult and Pediatric Fellows. The special challenges and opportunities for the academic pediatric pulmonary trainee were discussed as part of this workshop and are discussed as a companion paper to the report by the full workshop. Surveys were conducted of pediatric chairs of academic departments and pediatric pulmonary training directors in the United States to examine the current status and opportunities for the pediatric pulmonary trainee. Strategies for recruitment and retention of talented young trainees and junior faculty are proposed.

    View details for DOI 10.1002/ppul.21155

    View details for Web of Science ID 000273625000003

    View details for PubMedID 20025052

  • A Scientist Track Investigator Program to Support Early Career Outcomes for Clinician Scientists JOURNAL OF PEDIATRICS Rosenblum, N. D., Bazett-Jones, D. P., O'Brodovich, H. 2009; 155 (5): 603-U25

    View details for DOI 10.1016/j.jpeds.2009.07.047

    View details for Web of Science ID 000271570900001

    View details for PubMedID 19840610

  • Global and Gene-Specific Translational Regulation in Rat Lung Development AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Otulakowski, G., Duan, W., O'Brodovich, H. 2009; 40 (5): 555-567

    Abstract

    During the peripartum period, the lung must respond to dramatic changes in circulating hormones, nutritional factors, and physiologic signals during its transition to becoming the organ of gas exchange. Protein synthesis consumes a significant proportion of metabolic resources and is inhibited by many environmental stresses. We hypothesized that translational control mechanisms play a role in the perinatal lung. Immunoblots of late-gestation (Fetal Day [FD] 17-22) rat lung extracts revealed gradual decreases in phosphorylated forms of the mammalian target of rapamycin effectors, eukaryotic initiation factor (eIF) 4E-binding protein, p70 S6 kinase, and ribosomal protein S6, followed by sharp increases on Postnatal Day 1 (P1). Immunohistochemistry showed phospho-S6 staining was most prominent in epithelial cells of the large and small airways. m(7)GTP-sepharose pulldown experiments showed a decrease in association of translation initiation factor, eIF4E, with its inhibitor, eIF4E-binding protein, and a concomitant increase in eIF4E association with eIF4G immediately after birth, and polysome profiles confirmed a decrease in abundance of large polysomes between FD19 and FD22, which was reversed on P1. Microarray analysis of polysomal versus total RNA from FD19, FD22, and P1 lungs was used to identify specific genes, the association of which with large polysomes changed either pre- or postnatally. RT-PCR and Northern blotting were used to confirm translational changes in selected candidate genes, including a prenatal increase in IL-18 and a postnatal decrease in regulatory subunit 2 of protein phosphatase 1. Translational regulation of IL-18 and protein phosphatase 1 regulatory (inhibitor) subunit 2 is gene-specific, as these changes contrast with the corresponding global changes in polysome abundance.

    View details for DOI 10.1165/rcmb.2008-0284OC

    View details for Web of Science ID 000265620300006

    View details for PubMedID 18952566

  • Glucocorticoid-Mediated Repression of REDD1 mRNA Expression in Rat Fetal Distal Lung Epithelial Cells PEDIATRIC RESEARCH Otulakowski, G., Duan, W., Sarangapani, A., Gandhi, S., O'Brodovich, H. 2009; 65 (5): 514-519

    Abstract

    REDD1 (Regulated in Development and DNA Damage-1) is a stress-response gene that represses mammalian target of rapamycin (mTOR) thus decreasing protein synthesis. In contrast to studies using cell lines and adult alveolar type II (ATII) cells, we find that REDD1 mRNA levels did not increase in rat fetal distal lung epithelia (FDLE) or fetal lung fibroblasts grown in primary cultures and then exposed to 3% O2. REDD1 mRNA expression was repressed by dexamethasone (DEX) in FDLE and ATII, but induced by DEX in fibroblasts. Lung epithelial cell lines, A549 and MLE-15, showed increases in REDD1 mRNA in response to hypoxia and DEX. The effect of DEX on REDD1 mRNA and protein in FDLE and fibroblasts was dose- and time-dependent. Inhibitor studies support repression of REDD1 mRNA by DEX in FDLE was mediated via glucocorticoid receptor and not by nongenomic effects of glucocorticoids via MAPK pathways. The half-life of REDD1 mRNA was shorter in DEX-exposed FDLE compared with hormone-free media suggesting that DEX reduced REDD1 mRNA stability in FDLE. These studies indicate that REDD1 expression in response to hypoxia and DEX is cell-type specific and that physiologically appropriate levels of PO2 should be used when investigating fetal lung development.

    View details for Web of Science ID 000265448700006

    View details for PubMedID 19127203

  • Pulmonary Neuroendocrine Cell-Secreted Factors May Alter Fetal Lung Liquid Clearance PEDIATRIC RESEARCH Gandhi, S. G., Law, C., Duan, W., Otulakowski, G., O'Brodovich, H. 2009; 65 (3): 274-278

    Abstract

    The neuroendocrine system is most active at birth and may play a role in the transition from fetal to postnatal life, in particular in the lungs' transition from fluid secretion to fluid absorption. Pulmonary neuroendocrine cells do release dopamine (DA), serotonin, and gastrin-releasing peptide but their effects on lung ion and fluid transport are poorly understood. Therefore, we studied their effects on fetal distal lung explants and primary cultures of fetal distal lung epithelium (FDLE). We show that DA, but neither serotonin nor gastrin-releasing peptide, alters ion and fluid transport, in a dose-dependent manner. DAs effects were abrogated by D1/D2 receptor blockers in FDLE but not in explants. Propranolol abrogated DAs effects in both models. DA increased intracellular cAMP levels in FDLE. Terbutaline, forskolin, and isobutylmethylxanthine did not increase short circuit current (Isc) in DA-treated cells, despite a further increase in cAMP. We conclude that at least one, but not all mediators released by pulmonary neuroendocrine cells alter distal lung epithelial ion transport.

    View details for Web of Science ID 000263543000004

    View details for PubMedID 19092725

  • PROTEASE-ACTIVATED RECEPTOR (PAR)1 ALTERS BIOELECTRIC PROPERTIES OF DISTAL LUNG EPITHELIA WITHOUT COMPROMISING BARRIER FUNCTION EXPERIMENTAL LUNG RESEARCH Moraes, T. J., Rafii, B., Niessen, F., Suzuki, T., Martin, R., Vachon, E., Vogel, W., Ruf, W., O'Brodovich, H., Downey, G. P. 2009; 35 (2): 136-154

    Abstract

    Proteinases contribute to the pathogenesis of various lung diseases, partly through activating cell surface receptors by limited proteolytic cleavage. The authors provide evidence that in primary cultures of distal lung epithelia, basolateral protease-activated receptor 1 activation rapidly reduces transepithelial resistance but does not alter paracellular permeability to small uncharged solutes. Changes in transepithelial resistance were partially blocked by ion transport inhibitors and were completely blocked by placing cells in low chloride buffer. In vivo studies did not reveal enhanced lung permeability in response to pulmonary or intravenous administration of protease-activated receptor 1 activators. This information is relevant as strategies to inhibit protease-activated receptor 1 signaling are considered in order to preserve lung epithelial barrier function.

    View details for DOI 10.1080/01902140802490723

    View details for Web of Science ID 000263870100004

    View details for PubMedID 19263282

  • Visualizing water clearance in the lung with MRI MAGNETIC RESONANCE IN MEDICINE Vidarsson, L., Helm, E., O'Brodovich, H., Macgowan, C. K. 2008; 60 (1): 230-235

    Abstract

    Current indirect measurements of alveolar fluid clearance (AFC) suggest that the rate of fluid clearance correlates with morbidity and mortality in patients with pulmonary edema. In a traditional AFC-measurement, fluid laced with a tracer macromolecule is instilled into the lung and thereafter repeated samples of the instilled fluid are extracted from the lung's fluid-filled airspaces. The change in concentration of the tracer molecule indicates the AFC-rate. In this work, a new MRI technique was developed to image lung water clearance by adding Gadolinium-DTPA to the instilled fluid. As fluid is absorbed by the animal, the concentration of gadolinium will increase, reducing the T(1) relaxation time. By repeatedly measuring the T(1) relaxation time, the AFC can be tracked over time with high spatial resolution. The new technique was tested both in phantoms and 10 Yorkshire piglets.

    View details for DOI 10.1002/mrm.21644

    View details for Web of Science ID 000257267700028

    View details for PubMedID 18581395

  • Amiloride-insensitive Na+ and fluid absorption in the mammalian distal lung AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY O'Brodovich, H., Yang, P., Gandhi, S., Otulakowski, G. 2008; 294 (3): L401-L408

    Abstract

    The ability of the distal lung epithelia to actively transport Na+, with Cl- and water following, from the alveolar spaces inversely correlates with morbidity and mortality of infants, children, and adults with alveolar pulmonary edema. It is now recognized, in contrast to many other Na+ transporting epithelia, that at least half of this active transport is not sensitive to amiloride, which inhibits the epithelial Na+ channel. This paper reviews amiloride-insensitive Na+ and fluid transport in the mammalian distal lung unit under basal conditions and speculates on potential explanations for this amiloride-insensitive transport. It also provides new information, using primary cultures of rat fetal distal lung epithelia and alveolar type II cells grown under submersion and air-liquid interface culture conditions, regarding putative blockers of this transport.

    View details for DOI 10.1152/ajplung.00431.2007

    View details for Web of Science ID 000253822800003

    View details for PubMedID 18162600

  • Steroid and oxygen effects on elF4F complex, mTOR, and ENaC translation in fetal lung epithelia AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Otulakowski, G., Duan, W., Gandhi, S., O'Brodovich, H. 2007; 37 (4): 457-466

    Abstract

    Fetal distal lung epithelium (FDLE) must increase amiloride-sensitive epithelial Na(+) channel (ENaC) activity during the perinatal period to increase Na(+) transport and fluid clearance. Glucocorticosteroid (GC) levels increase, there is a 7-fold increase in Po(2) at birth, and we have previously shown that dexamethasone (DEX)-induced alpha-ENaC mRNA is efficiently translated only under postnatal (21%) O(2) (Otulakowski et al., AJRCMB 2006;34:204-212). Translation of mRNAs with long GC-rich 5'UTRs, such as alpha-ENaC mRNA, are sensitive to the amount of eIF4F, the mRNA 5'-cap binding complex composed of eIF4E and eIF4G. We now show, by Western blotting and m(7)GTP-Sepharose pull-down experiments, that in FDLE cultured under 3% O(2), DEX decreases formation of eIF4F and increases association of eIF4E with its inhibitor 4E-BP by changing 4E-BP phosphorylation. Conversely, FDLE cultured at 21% O(2) expressed lower levels of 4E-BP and maintained eIF4E-eIF4G association independent of DEX. Phosphorylation of 4E-BP is regulated by the kinase mTOR. Under 3% O(2), DEX decreased abundance of phosphorylated forms of the mTOR effectors, S6 kinase and ribosomal protein S6. Neither effect was associated with changes in REDD1, an upstream regulator of mTOR. When mTOR was inhibited (3 nM rapamycin) there was reduced 4E-BP phosphorylation, fewer ribosomes on alpha-ENaC mRNA, and decreased amiloride-sensitive short-circuit current, but no change in ribosomal loading onto any of beta- or gamma-ENaC or cytokeratin 18 mRNAs. We speculate that at birth increased Po(2) acts with GC through an mTOR-related pathway to increase alpha-ENaC protein synthesis, thereby promoting lung fluid absorption.

    View details for DOI 10.1165/rcmb.2007-0055OC

    View details for Web of Science ID 000249898400012

    View details for PubMedID 17556672

  • Purification and characterization of a recombinant rat prohaptoglobin expressed in baculovirus-infected Sf9 insect cells PROTEIN EXPRESSION AND PURIFICATION Hillar, A., Otulakowski, G., O'Brodovich, H. 2007; 55 (2): 246-256

    Abstract

    To generate hemoglobin-free full-length haptoglobin the cDNA encoding rat haptoglobin alphabeta subunits was cloned into shuttle vector pVT-Bac-His and used to produce a recombinant baculovirus Autographa californica Nuclear Polyhedrosis Virus (AcNPV) as an expression vector, named HpAcNPV. Recombinant virus was used to infect Spodoptera frugiperda (Sf9) insect cells. The 50 kDa protein expressed was mostly secreted into the culture medium at relatively high titer (15 microg/mL) and was found to be rat prohaptoglobin having a vector-derived N-terminal extension of 37 amino acids, containing both a hexahistidine tag and an enterokinase recognition sequence. The protein was successfully purified by a three step procedure including nickel-linked agarose and DEAE-Sepharose chromatography steps. Hemoglobin was not detected in the purified preparations. Purified recombinant rat prohaptoglobin protein was also found to be glycosylated, and to be capable of forming a complex with rat hemoglobin in vitro.

    View details for DOI 10.1016/j.pep.2007.06.004

    View details for Web of Science ID 000250070400004

    View details for PubMedID 17681809

  • Effects of cardiogenic edema fluid on ion and fluid transport in the adult lung AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Gandhi, S. G., Rafii, B., Harris, M. S., Garces, A., Mahuran, D., Chen, X., Bao, H., Jain, L., Eaton, D. C., Otulakowski, G., O'Brodovich, H. 2007; 293 (3): L651-L659

    Abstract

    We have previously shown that cardiogenic pulmonary edema fluid (EF) increases Na(+) and fluid transport by fetal distal lung epithelia (FDLE) (Rafii B, Gillie DJ, Sulowski C, Hannam V, Cheung T, Otulakowski G, Barker PM and O'Brodovich H. J Physiol 544: 537-548, 2002). We now report the effect of EF on Na(+) and fluid transport by the adult lung. We first studied primary cultures of adult type II (ATII) epithelium and found that overnight exposure to EF increased Na(+) transport, and this effect was mainly due to factors other than catecholamines. Plasma did not stimulate Na(+) transport in ATII. Purification of EF demonstrated that at least some agent(s) responsible for the amiloride-insensitive component resided within the globulin fraction. ATII exposed to globulins demonstrated a conversion of amiloride-sensitive short-circuit current (I(sc)) to amiloride-insensitive I(sc) with no increase in total I(sc). Patch-clamp studies showed that ATII exposed to EF for 18 h had increased the number of highly selective Na(+) channels in their apical membrane. In situ acute exposure to EF increased the open probability of Na(+)-permeant ion channels in ATII within rat lung slices. EF did increase, by amiloride-sensitive pathways, the alveolar fluid clearance from the lungs of adult rats. We conclude that cardiogenic EF increases Na(+) transport by adult lung epithelia in primary cell culture, in situ and in vivo.

    View details for DOI 10.1152/ajplung.00464.2006

    View details for Web of Science ID 000249125100020

    View details for PubMedID 17557800

  • The role of alpha-, beta-, and gamma-ENaC subunits in distal lung epithelial fluid absorption induced by pulmonary edema fluid AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Elias, N., Rafii, B., Rahman, M., Otulakowski, G., Cutz, E., O'Brodovich, H. 2007; 293 (3): L537-L545

    Abstract

    Edema fluid (EF) increases epithelial Na(+) transport by rat fetal distal lung epithelia (FDLE) and induces net lung fluid absorption in fetal mouse lung explants [Rafii B, Gillie DJ, Sulowski C, Hannam V, Cheung T, Otulakowski G, Barker PM, O'Brodovich H. J Physiol (Lond) 544: 537-548, 2002]. We now show that EF increases fluid absorption across monolayers of rat FDLE in a dose-dependent manner. To study the role of subunits of the epithelial Na(+) channel (ENaC) in the phenomena, we cultured explants from the distal lungs of 16-day gestational age wild-type (WT) or alpha-, beta-, or gamma-ENaC knockout or heterozygote (HT) mice. WT explants cultured in media continuously expanded over time as a result of net fluid secretion. In contrast, when explants were exposed to EF for 24 h, net fluid absorption occurred. EF-exposed explants had normal histology, but marked changes were seen after Triton X-100 or staurosporine exposure. Transmission electron microscopy showed EF promoted lamellar body formation and abundant surfactant in the explants' lumens. EF-induced changes in explant size were similar in alpha-ENaC knockout, WT, and HT littermate fetal lung explants (P > 0.05). In contrast, EF's effect was attenuated in beta- and gamma-ENaC knockouts (P < 0.05) vs. WT and HT littermate fetal lung explants. EF exposure slightly decreased or had no effect on mRNA levels for alpha-ENaC in various mouse genotypes but decreased expression of beta- and gamma-ENaC subunit mRNAs (P < 0.01) across all genotype groups. We conclude that beta- and gamma-, but not alpha-, ENaC subunits are essential for EF to exert its maximal effect on net fluid absorption by distal lung epithelia.

    View details for DOI 10.1152/ajplung.00373.2006

    View details for Web of Science ID 000249125100005

    View details for PubMedID 17513453

  • Amiloride-insensitive nasal potential difference varies with the menstrual cycle in cystic fibrosis PEDIATRIC PULMONOLOGY Sweezey, N. B., Smith, D., Corey, M., Ellis, L., Carpenter, S., Tullis, D. E., Durie, P., O'Brodovich, H. M. 2007; 42 (6): 519-524

    Abstract

    There is no adequate explanation for gender-based differences in rates of mortality and of deterioration in pulmonary function in cystic fibrosis (CF) patients. One potential explanation is that gender hormones (sex steroids) may modulate the severity of CF lung disease, the principal cause of mortality in CF, by altering respiratory transepithelial ion transport.To determine whether respiratory epithelial ion transport varied during the menstrual cycle of CF females.The nasal transepithelial electrical potential difference (NPD) was determined as a measure of ion transport across human respiratory epithelium, coincident with measurements of endogenous serum hormone levels in the luteal and follicular phases of the menstrual cycle in CF females aged 16-22 years.The component of the NPD that is insensitive to the Na(+) transport blocker amiloride, but not the amiloride-sensitive component, changed in association with endogenous, menstrual cycle-induced changes in serum levels of progesterone and estrogen (P=0.02, n=7, paired t-test). Measurements using Cl(-) free perfusates suggested that the changes are not a result of Cl(-) conductance.Our results suggest that in CF respiratory epithelium amiloride-insensitive, but not amiloride-sensitive, ion transport is altered by female gender hormones in vivo. We speculate that amiloride-insensitive ion transport may contribute to the regulation of human airway surface fluid.

    View details for DOI 10.1002/ppul.20624

    View details for Web of Science ID 000246872000006

    View details for PubMedID 17469152

  • Performance of a career development and compensation program at an academic health science center PEDIATRICS O'Brodovich, H., Beyene, J., Tallett, S., MacGregor, D., Rosenblum, N. D. 2007; 119 (4): E791-E797

    Abstract

    The academic physicians of our department developed a novel Career Development and Compensation Program to outline job expectations, enhance career development, and provide a peer-review process to assess performance. The Career Development and Compensation Program was founded on the principle that sustained achievement in education, clinical care, or research should be valued, supported, and rewarded in an equivalent manner and that reward for clinical work should not be limited by the focus of the university on research and education. The objective of this study was to determine whether the principles of the Career Development and Compensation Program were sustained during the initial 7 years of its implementation.The outcome of the 7 triennial reviews that occurred from 1999 to 2005 was evaluated. For the purposes of some analyses, physicians were classified as predominately clinical (clinician-specialists and clinician-teachers), predominately education (clinician-educators), or predominately research (clinician-investigators and clinician-scientists).Each of the job profiles had a similar probability to increase a level within the Career Development and Compensation Program at the time of triennial review. Similarly, all 5 job profiles had a similar rate of increase in their level in relation to the total number of years of experience at an academic health science center. Neither the university academic rank nor gender of the physician affected the probability of increasing a level at the time of the triennial review.The peer-reviewed Career Development and Compensation Program recognizes sustained achievement in each area of education, clinical care, and research in an equivalent manner with no detectable effect of academic rank or gender.

    View details for DOI 10.1542/peds.2006-2207

    View details for Web of Science ID 000245406200001

    View details for PubMedID 17339386

  • Thyroid hormone and Na+-K+-ATPase: more than simple transcription AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Otulakowski, G., O'Brodovich, H. 2007; 292 (1): L4-L5

    View details for DOI 10.1152/ajplung.00332.2006

    View details for Web of Science ID 000243399900002

    View details for PubMedID 16951130

  • Clinical and education workload measurements using personal digital assistant-based software PEDIATRICS MacGregor, D. L., Tallett, S., MacMillan, S., Gerber, R., O'Brodovich, H. 2006; 118 (4): E985-E991

    Abstract

    There are no accepted and practical measures of the relative clinical and educational activities of pediatricians who work in an academic health science center. Such measures are necessary for justification of existing and future human resource plans and evaluation of the activities and performance of physicians. The limited literature on the measurement of physician workload usually focuses on a specific subspecialty group and does not account for such issues as indirect patient care, such as telephone calls or e-mail consultations; variables that affect the delivery of clinical care, including patient acuity and complexity; and the presence of students during the patient care activities. After completing a pilot study that assessed the educational workload of faculty members, we adapted existing personal digital assistant technology and software to document clinical and educational activities.Twenty full-time physicians from 4 subspecialty pediatric divisions participated in a 2-week evaluation project in May through June 2005. Clinical activities, with and without trainees, and educational activities were collected with the use of personal digital assistants. Software allowed an individualized division-specific drop-down menu. Information that was collected included clinical (location of activity, diagnosis, and time requirement) and educational activities. After completion of a 2-week data collection period, each physician was asked to complete a 5-question evaluation form.The project was completed successfully with capture of additional clinical and educational activities. A 5-question evaluation form was completed by 70% of the participants at the end of the 2-week data collection. Data on clinical and educational activities were analyzed qualitatively and graphed.This method of workload data collection added significant information in capturing activities that are not measured in traditional workload evaluations for either clinical activities, such as e-mail, telephone, and patient information review, or educational endeavors, including mentoring and educational lectures and presentations.

    View details for DOI 10.1542/peds.2006-0515

    View details for Web of Science ID 000240959300079

    View details for PubMedID 17015518

  • Oxygen and glucocorticoids modulate alpha ENaC mRNA translation in fetal distal lung epithelium AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Otulakowski, G., Rafii, B., Harris, M., O'Brodovich, H. 2006; 34 (2): 204-212

    Abstract

    Glucocorticoid hormones play an important role in fetal lung maturation. It is unknown how they interact with changes in O2 tension, which play an important role in converting the lung from a fluid-secreting to a fluid-absorbing organ at birth. Airspace fluid absorption arises from active transepithelial Na+ transport with the amiloride-sensitive epithelial Na channel (ENaC), consisting of alpha, beta, and gamma subunits, representing the rate-limiting step under nonpathologic conditions. We investigated the individual and combined effects of dexamethasone (DEX) and PO2 on alphaENaC mRNA levels, rate of alphaENaC protein synthesis, and amiloride-sensitive short-circuit current in primary cultures of rat fetal distal lung epithelial cells. DEX significantly induced alphaENaC mRNA in fetal (3%) and postnatal (21%) O2, but increases in alphaENaC protein synthesis and function occurred only when epithelia were grown under a postnatal PO2. Sucrose density gradient analyses showed that DEX treatment of cells cultured at 3% O2 decreased the association of alphaENaC mRNA with large polysomes and enhanced the association with small polysomes. Conversely, incubation of DEX-treated cells in 21% O2 restored alphaENaC mRNA association with large polysomes. No significant changes were seen in the overall polyribosome profiles or in the distribution of mRNAs encoding beta and gamma subunits of ENaC or cytokeratin 18, indicating specific modulation of alphaENaC mRNA translation. These data suggest that postnatal O2 exposure may be important for efficient translation of the alphaENaC mRNA.

    View details for DOI 10.1165/rcmb.2005-0273OC

    View details for Web of Science ID 000235115700010

    View details for PubMedID 16210692

  • Fluid clearance from the lungs of newborns, infants and children PAEDIATRIC RESPIRATORY REVIEWS O'Brodovich, H. 2006; 7: S62-S63

    View details for DOI 10.1016/j.prrv.2006.04.173

    View details for Web of Science ID 000243007400020

    View details for PubMedID 16798598

  • Fitness to fly in an infant with congenital lobar emphysema AVIATION SPACE AND ENVIRONMENTAL MEDICINE MacLean, J. E., Manson, D., Gray, G., O'Brodovich, H. 2005; 76 (10): 989-991

    Abstract

    Determining fitness to fly is a difficult task for physicians, especially for those caring for children with respiratory disorders, as the available information for guidance is scarce. This case describes the use of a flight simulation in a decompression chamber in order to assess fitness to fly in an 18-mo-old infant with congenital lobar emphysema (CLE). The case discussion focuses on the need for an understanding of flight physiology in order for physicians to determine the most appropriate method to assess fitness to fly in children with medical concerns.

    View details for Web of Science ID 000232347500014

    View details for PubMedID 16235886

  • Pulmonary edema in infants and children CURRENT OPINION IN PEDIATRICS O'Brodovich, H. 2005; 17 (3): 381-384

    Abstract

    To provide an overview of the pathogenesis of pulmonary edema and describe recent discoveries related to the clearance of airspace fluid and potential new therapies for this life-threatening disorder.It is clinically important to determine the mechanisms responsible for the clearance of fluid from the airspaces. At birth inadequate clearance of fetal lung liquid is one of the two mechanisms leading to respiratory distress syndrome in the premature infant. Adults with heart failure or adult respiratory distress syndrome survive when they had active absorption of airspace fluid, yet have greater morbidity or die when they show no evidence of active fluid clearance. Humans who are susceptible to high-altitude pulmonary edema have less ability to actively transport fluid across their respiratory epithelium.New approaches to increase the active clearance of fluid from the airspaces, combined with further improvements in the intensive care and monitoring of patients with serious illnesses, will lead to improved care for patients with lung diseases characterized by increased lung water content.

    View details for Web of Science ID 000229268400007

    View details for PubMedID 15891430

  • Pediatric chairs of Canada: Academic pediatric workforce JOURNAL OF PEDIATRICS Scott, B., Frewen, T., O'Brodovich, H. 2004; 145 (4): 425-426

    View details for DOI 10.1016/j.jpeds.2004.08.017

    View details for Web of Science ID 000224417500001

    View details for PubMedID 15480356

  • Expression of epithelial sodium channel alpha-subunit mRNAs with alternative 5 '-untranslated regions in the developing human lung AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Banasikowska, K., Post, M., Cutz, E., O'Brodovich, H., Otulakowski, G. 2004; 287 (3): L608-L615

    Abstract

    In preparation for birth, lung epithelia must switch from net fluid secretion, required for lung development, to net absorption, which prepares the lungs for postnatal gas exchange. The apical membrane amiloride-sensitive epithelial Na channel (ENaC) is the rate-limiting step for Na+ and fluid absorption. Expression of alpha-ENaC mRNA has been detected in human lung as early as the embryonic stage of development. However, humans express multiple transcripts for alpha-ENaC, containing differing 5'-untranslated regions (UTR) with unknown effects on protein translation, and different ontogenies for individual transcripts could provide a novel mechanism for developmental regulation of ENaC function. To assess the relative expression of the two most abundant alpha-ENaC transcripts (alpha-ENaC1 and alpha-ENaC2) during lung development, we performed nonradioactive in situ hybridization using probes specific to the alternative 5'-UTRs. Both transcripts were expressed throughout intrauterine lung development (8 to 40 wk gestation), and expression was localized to the surface epithelial cells of the conductive and respiratory airways in both ciliated cells and nonciliated Clara cells. alpha-ENaC mRNA expression was also identified in the serous cells of the submucosal glands surrounding the proximal airways. In the mature prenatal lung, subsets of alveolar type II (ATII) cells expressed one or both of the alpha-ENaC transcripts. Our observations demonstrate that a developmentally regulated switch between alpha-ENaC 5'-UTR variants is not the trigger by which the developing human lung becomes a fluid-absorbing organ at birth, that individual ATII cells express neither, one, or both of the alpha-ENaC transcripts, and that the overall expression is linked to epithelial cell differentiation and lung maturation.

    View details for DOI 10.1152/ajplung.00031.2004

    View details for Web of Science ID 000223251000023

    View details for PubMedID 15169674

  • Differential translational efficiency of ENaC subunits during lung development AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Otulakowski, G., Rafii, B., O'Brodovich, H. 2004; 30 (6): 862-870

    Abstract

    The amiloride-sensitive epithelial Na(+) channel (ENaC), the rate-limiting step in epithelial Na(+) transport, consists of three subunits: alpha, beta, and gamma. The abundance of mRNA encoding the alpha-subunit far surpasses the amount for other subunits, and considerably exceeds the predicted subunit protein stoichiometry. We evaluated 5'-untranslated region (UTR) expression and found that fetal rat lung uses alternative 5'UTRs for alpha-ENaC during development. Sucrose density gradient analysis of postnuclear supernatants from fetal rat lung homogenates demonstrated that all three ENaC subunits were associated with high molecular weight polysomes, indicating active translation of the mRNAs, but translational efficiency was much lower for the alpha-subunit. Sucrose density gradient distributions were comparable for the endogenously expressed alpha-ENaC 5'UTRs in rat lung at Fetal Day 20 or Postnatal Day 1 using Northern analysis. Although birth resulted in a global decrease in lung mRNA translation, the loading of ribosomes on ENaC subunit mRNAs was largely unaffected. Evaluation of cytokeratin 18 and vimentin mRNAs in these gradients suggested a cell-specific effect. We conclude that there are different translational efficiencies for ENaC subunits and that perinatal processes globally modulate lung mRNA translation.

    View details for DOI 10.1165/rcmb.2003-0381OC

    View details for Web of Science ID 000221835100014

    View details for PubMedID 14672917

  • Low expression of human epithelial sodium channel in airway epithelium of preterm infants with respiratory distress PEDIATRICS Helve, O., Pitkanen, O. M., Andersson, S., O'Brodovich, H., Kirjavainen, T., Otulakowski, G. 2004; 113 (5): 1267-1272

    Abstract

    Active ion transport is critical to postnatal clearance of lung fluid. The importance of epithelial sodium channel (ENaC) in this clearance has been demonstrated in animal studies in which alpha-ENaC knockout mice died postnatally as a result of respiratory insufficiency. In animals, the expression of alpha-ENaC in respiratory epithelium is dependent on gestational age, but when assessed by in situ hybridization in the human (h), the mRNA is present from the earliest stages of pulmonary development. Therefore, the purpose of the present investigation was to quantify mRNA of the alpha-, beta-, and gamma-hENaC subunits of newborn preterm infants with respiratory distress and compare the gene expression data against those detected in healthy term infants. In addition, the effect of systemic dexamethasone therapy on the 3 hENaC subunits was studied in 4 preterm infants who received prolonged assisted ventilation.The expression of subunits of hENaC was determined in samples taken from nasal respiratory epithelium of 7 healthy term infants (gestation age: 39.3 +/- 0.9 weeks [mean +/- standard deviation) and 5 preterm infants (gestational age: 27.2 +/- 0.9 weeks) with respiratory distress syndrome within 5 hours of birth. Betamethasone had been given to all mothers of preterm infants. In 4 additional preterm infants who still required assisted ventilation at 43 +/- 6 days postnatal age, the expression of alpha-hENaC was determined in samples taken before and during treatment with dexamethasone.Preterm infants with respiratory distress syndrome had low expression of all hENaC subunits relative to healthy term infants (alpha-hENaC: 5.38 +/- 2.01 [amol/fmol cytokeratin 18] vs 9.13 +/- 2.26; beta-hENaC: 2.44 +/- 1.43 vs 4.25 +/- 1.10; gamma-hENaC: 2.43 +/- 0.11 vs 6.81 +/- 3.24). Each of the 4 preterm infants who were treated with dexamethasone at approximately 1 month of age showed an increase in expression of alpha-hENaC and beta-hENaC subunit normalized to cytokeratin 18.All 3 subunits of the hENaC are low in preterm relative to full-term infants. alpha-hENaC mRNA in respiratory epithelium is increased by therapeutic doses of glucocorticosteroid. Low expression of alpha-hENaC in human respiratory epithelium may play a role in the pathogenesis of respiratory distress in preterm infants.

    View details for Web of Science ID 000221169200015

    View details for PubMedID 15121940

  • Acquisition of competencies during the pediatric residency: A Canadian perspective JOURNAL OF PEDIATRICS Tallett, S., O'Brodovich, H. 2004; 144 (3): 289-290

    View details for DOI 10.1016/j.jpeds.2003.11.021

    View details for Web of Science ID 000220155800001

    View details for PubMedID 15001928

  • Airspace fluid clearance in the normal and edematous newborn lung: Cardiogenic pulmonary edema fluid alters the clearance of airspace fluid PEDIATRIC PULMONOLOGY O'Brodovich, H. 2004: 121-122

    View details for DOI 10.1002/ppul.70077

    View details for Web of Science ID 000189078500048

    View details for PubMedID 15029623

  • Deficient Na+ channel expression in newborn respiratory distress syndrome PEDIATRIC PULMONOLOGY O'Brodovich, H. 2004: 141-142

    View details for DOI 10.1002/ppul.70085

    View details for Web of Science ID 000189078500056

    View details for PubMedID 15029631

  • Evaluation of a peer-reviewed career development and compensation program for physicians at an academic health science center PEDIATRICS O'Brodovich, H., Pleinys, R., Laxer, R., Tallett, S., Rosenblum, N., Sass-Kortsak, C. 2003; 111 (1)

    Abstract

    The Department of Pediatrics at the Hospital for Sick Children, which is funded by an alternative payment plan, has implemented a novel career development and compensation program (CDCP). Job activity profiles were used to more clearly define job expectations, benchmarks guided career development, and peer review was used to assess performance. The objective of this study was to evaluate the departmental pediatricians' satisfaction with the CDCP.Pediatricians, all of whom had undergone CDCP annual reviews, could participate if they had undergone the in-depth triennial CDCP review. Each received a 5-point Likert scale-based questionnaire that asked how well the CDCP had conformed to the principles identified by the department during the development of the CDCP. Anonymous, confidential responses were collated and used to guide focus groups that discussed areas of greatest concern and attempted to identify solutions. Focus groups were led by external facilitators who were experienced in qualitative research. They audiotaped the sessions, transcribed the comments, and analyzed the data with the assistance of a qualitative analysis application.Sixty of the eligible 88 pediatricians participated, and 74% of their responses were that the CDCP had addressed the original principles "somewhat," "to a great extent," or "extremely well." The remainder indicated that some of the principles were either "not addressed" or "only to a small extent" by the CDCP. Results from the 11 focus groups (46 participants) indicated that the CDCP was an improvement over the previous method of career development and determination of the rate of remuneration. Most were also still in agreement with the purpose and design principles. Although they did not want the CDCP to undergo a major redesign, they identified areas that need improvement. Short-, medium-, and long-term action plans to address these areas are under way.Pediatricians at the health science center of the Hospital for Sick Children remain supportive of the CDCP.

    View details for Web of Science ID 000180135200004

    View details for PubMedID 12509591

  • Pulmonary oedema fluid induces non-alpha-ENaC-dependent Na+ transport and fluid absorption in the distal lung JOURNAL OF PHYSIOLOGY-LONDON Rafii, B., Gillie, D. J., Sulowski, C., Hannam, V., Cheung, T., Otulakowski, G., Barker, P. M., O'Brodovich, H. 2002; 544 (2): 537-548

    Abstract

    To determine if pulmonary oedema fluid (EF) alters ion and fluid transport of distal lung epithelium (DLE), EF was collected from rats in acute heart failure. EF, but not plasma, increased amiloride-insensitive short circuit current (I(sc)) and Na(+)-K(+) ATPase protein content and pump activity of DLE grown in primary culture. Inhibitors of Cl(-) transport or cGMP-gated cation channels had a significant (P < 0.05), but limited ability to block the increased I(sc). EF increased amiloride-insensitive, but not amiloride-sensitive, DLE apical membrane Na(+) conductance. The level of mRNA encoding epithelial sodium channel (ENaC) subunits was unchanged (alpha, beta), or decreased (gamma, P < 0.05) in EF-exposed DLE. EF also induced an amiloride-insensitive increase in the potential difference across murine tracheal cysts. Distal lung explants from late gestation wild-type and alpha-ENaC-deficient fetal mice, which normally expand due to liquid secretion, decreased in size due to liquid absorption when exposed to EF. Trypsin digestion or heat treatment of EF abrogated the ability of EF to increase amiloride-insensitive I(sc) in DLE and liquid absorption by distal lung explants. Thus proteins or protein-dependent factors within cardiogenic EF induce an alpha-ENaC-independent and amiloride-insensitive apical membrane Na(+) conductance and liquid absorption in the distal lung.

    View details for DOI 10.1113/jphysiol.2002.024612

    View details for Web of Science ID 000178875000021

    View details for PubMedID 12381825

  • Pulmonary interstitial glycogenosis - A new variant of neonatal interstitial lung disease AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE Canakis, A. M., Cutz, E., Manson, D., O'Brodovich, H. 2002; 165 (11): 1557-1565

    Abstract

    We present the clinical, radiologic, and pathologic findings in lung biopsies from seven infants with atypical neonatal lung disease. All seven infants presented with tachypnea, hypoxemia, and diffuse interstitial infiltrates with overinflated lungs on chest radiographs in the first month of life. Lung biopsies from all cases showed similar pathology, with expansion of the interstitium by spindle-shaped cells containing periodic acid-Schiff positive diastase labile material consistent with glycogen. Immunohistochemical staining showed these cells to be vimentin positive but negative for leucocyte common antigen, lysozyme, and other macrophage markers. Electron microscopy revealed primitive interstitial mesenchymal cells with few cytoplasmic organelles and abundant monoparticulate glycogen. Minimal or no glycogen was seen in the alveolar lining cells. Five cases were treated with pulse corticosteroids; hydroxychloroquine was added in one case. Six of seven infants have shown a favorable clinical outcome. One infant died from complications of extreme prematurity and bronchopulmonary dysplasia. Three cases that have been followed for at least 6 years have shown clinical resolution and radiographic improvement. We propose the term "pulmonary interstitial glycogenosis" of the neonate for this new entity to be differentiated from other forms of interstitial lung disease. Because abundant glycogen is not normally found in pulmonary interstitial cells, we postulate an abnormality in lung cytodifferentiation involving interstitial mesenchymal cells.

    View details for DOI 10.1164/rccm.2105139

    View details for Web of Science ID 000176069400019

    View details for PubMedID 12045133

  • Promoter analysis of the gene encoding the beta-subunit of the rat amiloride-sensitive epithelial sodium channel AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Bremner, H. R., Freywald, T., O'Brodovich, H. M., Otulakowski, G. 2002; 282 (1): L124-L134

    Abstract

    The amiloride-sensitive epithelial Na(+) channel (ENaC), found in the apical membrane of Na(+)-absorptive epithelia, is made up of three differentially regulated subunits: alpha, beta, and gamma. We undertook a study of the 5'-end of the gene encoding the beta-ENaC subunit in the rat. 5'-Rapid amplification of cDNA ends and RNase protection assays indicated multiple transcription start sites over a 50-bp region. Sequencing 1.3 kb of the 5'-flanking DNA revealed putative binding sites for PEA3, Sp1, activator protein (AP)-1 and Oct-1 but neither a TATA box nor consensus sites for steroid hormone receptor binding. Transient transfections of reporter constructs driven by beta-ENaC 5'-flanking DNA in the representative epithelial cell lines Madin-Darby canine kidney, MLE-15, and Caco-2 revealed a negative element present between positions -424 and -311 that affected basal transcription rates. Gel shift assays showed protein-DNA binding activity of an AP-1 consensus site in this region; however, mutation of the AP-1 site did not abrogate the repressive activity of the region in transient transfections. Deletion of two clusters of Sp1 consensus binding sites between -1 and -51 bp and between -169 and -211 bp indicated that the proximal cluster was essential to basal promoter activity in transfected cell lines. In a comparison of these data with those in published studies on alpha- and gamma-ENaC promoters, the beta- and gamma-subunit promoters appear to be more similar to each other than to the alpha-promoter.

    View details for Web of Science ID 000172686200016

    View details for PubMedID 11741824

  • Pulmonary edema fluid movement within the lung AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY O'Brodovich, H. 2001; 281 (6): L1324-L1326

    View details for Web of Science ID 000172094400002

    View details for PubMedID 11704525

  • Translational activation and repression by distinct elements within the 5 '-UTR of ENaC alpha-subunit mRNA AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Otulakowski, G., Freywald, T., Wen, Y. X., O'Brodovich, H. 2001; 281 (5): L1219-L1231

    Abstract

    The rat amiloride-sensitive epithelial Na(+) channel (rENaC), the rate-limiting step in epithelial Na(+) transport, consists of three subunits, alpha, beta, and gamma. We hypothesized that alpha-rENaC translation is regulated via its 5'-untranslated region (UTR). Transient transfections of alpha-rENaC promoter-reporter constructs in representative epithelial cell lines demonstrated up to fivefold differences in activity among constructs containing different amounts of the alpha-rENaC 5'-UTR sequence. Differences in reporter protein activity did not parallel differences in reporter mRNA, demonstrating that 5'-UTR regulation must be at the level of translation. Specifically, translation was enhanced by a region extending from +53 to +211 bp downstream from the transcription start site and repressed by the region between +367 and +499 bp. Examination of the 5'-UTR sequence revealed an out-of-frame initiation codon within the repressive region, 43 bp upstream from the start of the alpha-rENaC open reading frame. Mutational analysis of this upstream start codon indicated that it plays, at most, a minor role in impeding translation both in vitro and in vivo, suggesting that additional mechanisms of translational regulation are operative.

    View details for Web of Science ID 000171502700022

    View details for PubMedID 11597914

  • Career development and compensation: Strategies for physicians in academic health science centers - A perspective from a Canadian academic health science center JOURNAL OF PEDIATRICS O'Brodovich, H. 2001; 139 (2): 171-172

    View details for Web of Science ID 000170360100001

    View details for PubMedID 11487737

  • Fetal lung liquid secretion - Insights using the tools of inhibitors and genetic knock-out experiments AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY O'Brodovich, H. 2001; 25 (1): 8-10

    View details for Web of Science ID 000170221800003

    View details for PubMedID 11472968

  • Expression of alpha-, beta-, and gamma-hENaC mRNA in the human nasal, bronchial, and distal lung epithelium AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE Pitkanen, O. M., Smith, D., O'Brodovich, H., Otulakowski, G. 2001; 163 (1): 273-276

    Abstract

    The amount of fluid covering the epithelium of the airways and alveolar space is modulated by active transport of Na+ from the lumen through the apical membrane Na+ permeant ion channels towards the interstitial space. We have measured the subunit expression of the amiloride-sensitive human Na+ channel (hENaC) by concomitant assessment of alpha-, beta-, and gamma-hENaC mRNA in the nasal, bronchial, and peripheral lung epithelia of adult patients undergoing lobectomy secondary to lung cancer. The study employed quantitative competitive reverse-transcriptase-polymerase chain reaction and qualitative in situ hybridization techniques. The hENaC mRNA content of each sample was normalized to the amount of epithelial cell-specific cytokeratin 18 (CK18) mRNA. Nasal epithelium contained significantly more (p < 0.05) alpha-hENaC mRNA (18 +/- 5 SD amol/fmol CK18), than bronchus (8 +/- 2 SD amol/fmol) and peripheral lung (9 +/- 2 SD amol/fmol). The ratio of gamma-hENaC/alpha-hENaC mRNA concentration was lowest in the nasal area, and it increased significantly towards the distal lung regions. The change in beta-hENaC mRNA was less profound. In situ hybridization studies of bronchial and peripheral lung sections selectively revealed expression of alpha-hENaC mRNA in superficial epithelium and submucosal glands of large airways, in bronchiolar epithelium, and in alveolar cells. We conclude that the relative expression of the hENaC subunit genes changes from the proximal to distal regions of the human respiratory tract.

    View details for Web of Science ID 000166540100045

    View details for PubMedID 11208656

  • Transplant immunosuppression increases and prolongs transgene expression following adenoviral-mediated transfection of rat lungs JOURNAL OF HEART AND LUNG TRANSPLANTATION Cassivi, S. D., Liu, M. Y., Boehler, A., Pierre, A., Tanswell, A. K., O'Brodovich, E., Mullen, J. B., Slutsky, A. S., Keshavjee, S. H. 2000; 19 (10): 984-994

    Abstract

    Gene therapy provides the potential to modify donor organs to better withstand transplantation, but this has yet to be realized. In vivo gene transfer using adenoviral vectors has had limited success because of host immune response that induces inflammation and limits the amount and duration of transgene expression. We hypothesize that transplantation immunosuppression can attenuate the post-transfection host-immune response to allow for improved gene transfer following adenoviral-mediated transfection.We intratracheally transfected with adenovirus containing the beta-galactosidase gene and randomized the rats to either the immunosuppression group, receiving daily cyclosporine, azathioprine, and methylprednisolone, or the control group, receiving no immunosuppression. We evaluated transgene expression and post-transfection inflammation at time points ranging from 1 day to 5 weeks.Following transfection, control rats showed relatively low levels of transgene expression, which rapidly decreased to non-detectable levels. In contrast, immunosuppressed rats demonstrated significantly higher levels of transgene expression overall (p < 0.00005), peaking at almost 3 times that of the control group (p < 0.02), and showing prolonged and elevated transgene expression at 5 weeks (p < 0.02). On histologic sections of the lungs, immunosuppressed rats exhibited overall lesser grades of post-transfection inflammation.Transplant immunosuppression provides the means to attenuate the severe immune response to adenoviral-mediated gene transfection and thereby increase and prolong transgene expression.

    View details for Web of Science ID 000090034100010

    View details for PubMedID 11044694

  • Preventing endotoxin-stimulated alveolar macrophages from decreasing epithelium Na+ channel (ENaC) mRNA levels and activity PEDIATRIC RESEARCH Dickie, A. J., Rafii, B., Piovesan, J., Davreux, C., Ding, J. W., Tanswell, A. K., Rotstein, O., O'Brodovich, H. 2000; 48 (3): 304-310

    Abstract

    The acute respiratory distress syndrome is characterized by impairment of the alveolar-capillary barrier. Our laboratory has shown that distal lung epithelial cell (DLEC) amiloride-sensitive Na+ transport is impaired by in vitro coculture with endotoxin (lipopolysaccharide)-stimulated alveolar macrophages (AM) through an L-arginine-dependent mechanism. To investigate the effect of this model on mRNA levels of the rat epithelial Na+ channel, mature fetal rat DLEC monolayers were incubated for 16 h with rat AM (1 x 10(7)) and lipopolysaccharide (10 microg/mL), or the cell-free supernatant of lipopolysaccharide-stimulated rat AM. Such exposure resulted in a profound decrease in mRNA expression for all subunits (alpha, beta, and gamma) of the rat epithelial Na+ channel, without affecting 18S RNA levels. This effect was prevented by the antioxidant N-acetylcysteine. In separate experiments, confluent DLEC monolayers were exposed to lipopolysaccharide-stimulated AM supernatant for 16 h with or without N-acetylcysteine and DTT and studied in Ussing chambers. As previously demonstrated in our laboratory, AM supernatant resulted in a significant (p < 0.05) impairment of DLEC Na+ transport, as reflected by a decrease in the amiloride-sensitive component of short-circuit current (control, 3.96 +/- 0.18 microA/cm2 versus supernatant, 2.34 +/- 0.56 microA/cm2; p < 0.05). This effect was significantly reversed by N-acetylcysteine (3.55 +/- 0.48 microA/cm2), but not by DTT (1.87 +/- 0.21 microA/cm2). N-acetylcysteine, but not DTT, increased DLEC thiol levels. These studies elucidate mechanisms by which activated AM impair alveolar epithelial barrier function in an in vitro model of acute lung injury.

    View details for Web of Science ID 000088845900008

    View details for PubMedID 10960494

  • Epithelial Na+ channel (ENaC) expression in the developing normal and abnormal human perinatal lung AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE Smith, D. E., Otulakowski, G., Yeger, M., Post, M., Cutz, E., O'Brodovich, H. M. 2000; 161 (4): 1322-1331

    Abstract

    Impaired lung epithelial Na(+) channel (ENaC) activity at the time of birth results in respiratory distress. To investigate potential mechanisms, the ontogeny and cellular distribution of the alphaENaC subunit mRNA expression was studied in normal, immature, and abnormal (hypoplastic) human fetal lungs using nonradioisotopic in situ hybridization. Surprisingly, alphaENaC expression was detected at the embryonic stage of normal lung development (4 to 5 wk gestation) when expression was localized to the fetal lung bud epithelium. By late gestation, ENaC was expressed in the conductive and respiratory airway epithelium, serous cells, and the distal lung unit in an alveolar type II (ATII) epitheliumlike distribution. Significant alphaENaC expression was found in newborn lung diseases associated with respiratory distress. One explanation is that alphaENaC mRNA is constitutively expressed, and that activity is regulated, at least in part, at the post-transcriptional level. Alternative explanations are that the expression of the beta or gammaENaC subunits may be impaired in certain newborn lung diseases or that alternate Na(+) permeant channels or transporters are important to lung liquid absorption in humans at birth.

    View details for Web of Science ID 000086573400041

    View details for PubMedID 10764330

  • Oxygen induction of epithelial Na+ transport requires heme proteins AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Rafii, B., Coutinho, C., Otulakowski, G., O'Brodovich, H. 2000; 278 (2): L399-L406

    Abstract

    Fetal distal lung epithelial (FDLE) cells exposed to a postnatal O(2) concentration of 21% have higher epithelial Na(+) channel (ENaC) mRNA levels and Na(+) transport relative to FDLE cells grown in a fetal O(2) concentration of 3%. To investigate the mechanism of this process, FDLE monolayers were initially cultured in 3% O(2), and then some were switched to a 21% O(2) environment. Incubation of FDLE cells with the iron chelator deferoxamine, CoCl(2), NiCl(2), or an inhibitor of heme synthesis prevented or diminished the O(2) induction of amiloride-sensitive short-circuit current in FDLE cells. Similarly, defer- oxamine and cobalt prevented O(2)-induced ENaC mRNA expression. Exposure of FDLE cells grown under hypoxic conditions to carbon monoxide increased both ENaC mRNA expression and amiloride-sensitive short-circuit current. We therefore concluded that induction of ENaC mRNA expression and amiloride-sensitive Na(+) transport in FDLE cells by a physiological increase in O(2) concentration seen at birth requires iron and heme proteins.

    View details for Web of Science ID 000085278500024

    View details for PubMedID 10666125

  • Hormonal regulation and genomic organization of the human amiloride-sensitive epithelial sodium channel alpha subunit gene PEDIATRIC RESEARCH Chow, Y. H., Wang, Y., Plumb, J., O'Brodovich, H., Hu, J. 1999; 46 (2): 208-214

    Abstract

    To investigate the regulation of the amiloride-sensitive epithelial sodium channel (ENaC) expression, we have characterized the genomic structure and performed promoter analyses of the alpha subunit of the human (h) ENaC gene. Genomic clones containing the alphahENaC gene were isolated and subjected to restriction-mapping analysis. The alphahENaC gene was shown to be composed of 13 exons and 12 introns. Primer extension analysis confirmed that transcription initiation occurred at the beginning of the reported alphahENaC cDNA, but also indicated potential heterogenous initiation sites. Examination of a 3.1 kb 5' flanking sequence revealed a notable absence of CCAAT or TATA-like elements but suggested three GC boxes and several putative transcription factor binding sites, including a glucocorticoid response element (GRE) consensus. A 250 bp minimal promoter was capable of directing expression of a secreted alkaline phosphatase reporter. This promoter activity was enhanced 2.5- and 4-fold by upstream flanking sequences. Dexamethasone treatment induced levels of expression from the longer, GRE-containing promoter fragments from 8- to 20-fold, but not from the minimal promoter. Precise deletion of the 15-bp, dyad GRE sequence completely abolishes the response of reporter expression to dexamethasone induction. These experiments indicate that glucocorticoid augmentation of lung epithelial Na+ transport occurs, at least in part, by direct stimulation of transcription of the ENaC genes.

    View details for Web of Science ID 000081690500014

    View details for PubMedID 10447117

  • Alternate funding arrangements to compensate physicians at Academic Health Science Centers JOURNAL OF PEDIATRIC GASTROENTEROLOGY AND NUTRITION O'Brodovich, H. 1999; 29 (1): 8-9

    View details for Web of Science ID 000081057800004

    View details for PubMedID 10400094

  • Structure and hormone responsiveness of the gene encoding the alpha-subunit of the rat amiloride-sensitive epithelial sodium channel AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Otulakowski, G., Rafii, B., Bremner, H. R., O'Brodovich, H. 1999; 20 (5): 1028-1040

    Abstract

    The rat amiloride-sensitive epithelial sodium channel (rENaC) is the rate-limiting step for vectorial transport of Na+ across tight epithelia. The complex is composed of three subunits, alpha, beta, and gamma. Expression of the subunits has been shown to be tissue-specific and developmentally and hormonally regulated. To study mechanisms involved in transcriptional regulation of alpharENaC, we determined the genomic organization of the alpharENaC gene. By 5' rapid amplification of cDNA ends and primer extension, two transcriptional start sites were detected 453 base pairs (bp) apart, resulting in alternative 5' untranslated region (UTR) lengths of 515 or 62 bp. The longer 5' UTR is more prevalent in fetal lung than in adult lung or kidney. The 5' untranslated and coding regions are contained within 12 exons, with the translation start site located within the first exon. Sequence analysis of approximately 1,500 bp of 5' flanking DNA identified putative binding sites for transcription factors PEA3, SP1, AP-1, nuclear factor-kappaB, and thyroid and glucocorticoid receptors. alpharENaC promoter-reporter gene constructs produced low levels of reporter gene activity in transiently transfected cells, which could be increased by dexamethasone (DEX) treatment. Tri-iodothyronine treatment alone had no effect but potentiated stimulation by DEX.

    View details for Web of Science ID 000080388900022

    View details for PubMedID 10226074

  • Metabolic instability of plasmid DNA in the cytosol: a potential barrier to gene transfer GENE THERAPY Lechardeur, D., Sohn, K. J., Haardt, M., Joshi, P. B., Monck, M., Graham, R. W., Beatty, B., Squire, J., O'Brodovich, H., LUKACS, G. L. 1999; 6 (4): 482-497

    Abstract

    Inefficient nuclear delivery of plasmid DNA is thought to be one of the daunting hurdles to gene transfer, utilizing a nonviral delivery system such as polycation-DNA complex. Following its internalization by endocytosis, plasmid DNA has to be released into the cytosol before its nuclear entry can occur. However, the stability of plasmid DNA in the cytoplasm, that may play a determinant role in the transfection efficiency, is not known. The turnover of plasmid DNA, delivered by microinjection into the cytosol, was determined by fluorescence in situ hybridization (FISH) and quantitative single-cell fluorescence video-image analysis. Both single- and double-stranded circular plasmid DNA disappeared with an apparent half-life of 50-90 min from the cytoplasm of HeLa and COS cells, while the amount of co-injected dextran (MW 70,000) remained unaltered. We propose that cytosolic nuclease(s) are responsible for the rapid-degradation of plasmid DNA, since (1) elimination of plasmid DNA cannot be attributed to cell division or to the activity of apoptotic and lysosomal nucleases; (2) disposal of microinjected plasmid DNA was inhibited in cytosol-depleted cells or following the encapsulation of DNA in phospholipid vesicles; (3) generation and subsequent elimination of free 3'-OH ends could be detected by the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling assay (TUNEL), reflecting the fragmentation of the injected DNA; and finally (4) isolated cytosol, obtained by selective permeabilization of the plasma membrane, exhibits divalent cation-dependent, thermolabile nuclease activity, determined by Southern blotting and 32P-release from end-labeled DNA. Collectively, these findings suggest that the metabolic instability of plasmid DNA, caused by cytosolic nuclease, may constitute a previously unrecognized impediment for DNA translocation into the nucleus and a possible target to enhance the efficiency of gene delivery.

    View details for Web of Science ID 000079560200005

    View details for PubMedID 10476208

  • Regulation of an amiloride-sensitive Na+-permeable channel by a beta(2)-adrenergic agonist, cytosolic Ca2+ and Cl- in fetal rat alveolar epithelium JOURNAL OF PHYSIOLOGY-LONDON Marunaka, Y., Niisato, H., O'Brodovich, H., Eaton, D. C. 1999; 515 (3): 669-683

    Abstract

    1. In cell-attached patches formed on the apical membrane of fetal alveolar epithelium, terbutaline (a specific beta2-adrenergic agonist) increased the open probability (Po) of an amiloride-sensitive Na+-permeable non-selective cation (NSC) channel (control, 0.03 +/- 0.04; terbutaline, 0.62 +/- 0.18; n = 8, P < 0. 00001) by increasing the mean open time 100-fold without any significant change in the mean closed time and without any change in the single channel conductance (control, 27.8 +/- 2.3 pS; terbutaline, 28.2 +/- 2.1 pS; n = 8). 2. The Po of the unstimulated channel increased when the apical membrane was depolarized due to a decrease in the closing rate and an increase in the opening rate, while the Po of the terbutaline-stimulated channel did not depend on the membrane potential. 3. Increased cytosolic [Ca2+] also increased the Po of the channel in a manner consistent with one Ca2+-binding site on the cytosolic surface of the channel. Terbutaline increased the sensitivity of the channel to cytosolic Ca2+ by shifting the concentration of cytosolic Ca2+ ([Ca2+]c) required for half-maximal activation to a lower [Ca2+]c value, leading to an increase in Po. 4. An increase in the cytosolic Cl- concentration ([Cl-]c) decreased the Po of the channel consistent with two Cl--binding sites by increasing the closing rate without any significant change in the opening rate. Terbutaline increased Po by reducing the effect of cytosolic Cl- to promote channel closing. 5. Taken together, these observations indicate that terbutaline activates a Ca2+-activated, Cl--inhibitable, amiloride-sensitive, Na+-permeable NSC channel in fetal rat alveolar epithelium in two ways: first, through an increase in Ca2+ sensitivity, and second, through a reduction in the effect of cytosolic Cl- to promote channel closing.

    View details for Web of Science ID 000079792600005

    View details for PubMedID 10066896

  • Roles of Ca2+ and protein tyrosine kinase in insulin action on cell volume via Na+ and K+ channels and Na+/K+/2Cl(-) cotransporter in fetal rat alveolar type II pneumocyte JOURNAL OF MEMBRANE BIOLOGY Marunaka, Y., Niisato, N., O'Brodovich, H., Post, M., Tanswell, A. K. 1999; 168 (1): 91-101

    Abstract

    The aim of the present study was to investigate the roles of Ca2+ and protein tyrosine kinase (PTK) in the insulin action on cell volume in fetal rat (20-day gestational age) type II pneumocytes. Insulin (100 nm) increased cell volume in the presence of extracellular Ca2+ (1 mm), while cell shrinkage was induced by insulin in the absence of extracellular Ca2+ (<1 nm). This insulin action in a Ca2+-containing solution was completely blocked by co-application of bumetanide (50 microm, an inhibitor of Na+/K+/2Cl- cotransporter) and amiloride (10 microm, an inhibitor of epithelial Na+ channel), but not by the individual application of either bumetanide or amiloride. On the other hand, the insulin action on cell volume in a Ca2+-free solution was completely blocked by quinine (1 mm, a blocker of Ca2+-activated K+ channel), but not by bumetanide and/or amiloride. These observations suggest that insulin activates an amiloride-sensitive Na+ channel and a bumetanide-sensitive Na+/K+/2Cl- cotransporter in the presence of 1 mm extracellular Ca2+, that the stimulatory action of insulin on an amiloride-sensitive Na+ channel and a bumetanide-sensitive Na+/K+/2Cl- cotransporter requires Ca2+, and that in a Ca2+-free solution insulin activates a quinine-sensitive K+ channel but not in the presence of 1 mm Ca2+. The insulin action on cell volume in a Ca2+-free solution was almost completely blocked by treatment with BAPTA (10 microm) or thapsigargin (1 microM, an inhibitor of Ca2+-ATPase which depletes the intracellular Ca2+ pool). Further, lavendustin A (10 microm, an inhibitor of receptor type PTK) blocked the insulin action in a Ca2+-free solution. These observations suggest that the stimulatory action of insulin on a quinine-sensitive K+ channel is mediated through PTK activity in a cytosolic Ca2+-dependent manner. Lavendustin A, further, completely blocked the activity of the Na+/K+/2Cl- cotransporter in a Ca2+-free solution, but only partially blocked the activity of the Na+/K+/2Cl- cotransporter in the presence of 1 mm Ca2+. This observation suggests that the activity of the Na+/K+/2Cl- cotransporter is maintained through two different pathways; one is a PTK-dependent, Ca2+-independent pathway and the other is a PTK-independent, Ca2+-dependent pathway. Further, we observed that removal of extracellular Ca2+ caused cell shrinkage by diminishing the activity of the amiloride-sensitive Na+ channel and the bumetanide-sensitive Na+/K+/2Cl- cotransporter, and that removal of extracellular Ca2+ abolished the activity of the quinine-sensitive K+ channel. We conclude that the cell shrinkage induced by removal of extracellular Ca2+ results from diverse effects on the cotransporter and Na+ and K+ channels.

    View details for Web of Science ID 000078969000009

    View details for PubMedID 10051692

  • Induction of epithelial sodium channel (ENaC) expression and sodium transport in distal lung epithelia by oxygen AMILORIDE-SENSITIVE SODIUM CHANNELS Rafii, B., Tanswell, A. K., Pitkanen, O., O'Brodovich, H. 1999; 47: 239-254
  • HSC clinical trials controversy continues NATURE MEDICINE Buchwald, M., O'Brodovich, H. 1999; 5 (1): 2-3

    View details for Web of Science ID 000077885000002

    View details for PubMedID 9883819

  • Sodium channels in alveolar epithelial cells: Molecular characterization, biophysical properties, and physiological significance ANNUAL REVIEW OF PHYSIOLOGY Matalon, S., O'Brodovich, H. 1999; 61: 627-661

    Abstract

    At birth, fetal distal lung epithelial (FDLE) cells switch from active chloride secretion to active sodium (Na+) reabsorption. Sodium ions enter the FDLE and alveolar type II (ATII) cells mainly through apical nonselective cation and Na(+)-selective channels, with conductances of 4-26 pS (picoSiemens) in FDLE and 20-25 pS in ATII cells. All these channels are inhibited by amiloride with a 50% inhibitory concentration of < 1 microM, and some are also inhibited by [N-ethyl-N-isopropyl]-2'-4'-amiloride (50% inhibitory concentration of < 1 microM). Both FDLE and ATII cells contain the alpha-, beta-, and gamma-rENaC (rat epithelial Na+ channels) mRNAs; reconstitution of an ATII cell Na(+)-channel protein into lipid bilayers revealed the presence of 25-pS Na+ single channels, inhibited by amiloride and [N-ethyl-N-isopropyl]-2'-4'-amiloride. A variety of agents, including cAMP, oxygen, glucocorticoids, and in some cases Ca2+, increased the activity and/or rENaC mRNA levels. The phenotypic properties of these channels differ from those observed in other Na(+)-absorbing epithelia. Pharmacological blockade of alveolar Na+ transport in vivo, as well as experiments with newborn alpha-rENaC knock-out mice, demonstrate the importance of active Na+ transport in the reabsorption of fluid from the fetal lung and in reabsorbing alveolar fluid in the injured adult lung. Indeed, in a number of inflammatory diseases, increased production of reactive oxygen-nitrogen intermediates, such as peroxynitrite (ONOO-), may damage ATII and FDLE Na+ channels, decrease Na+ reabsorption in vivo, and thus contribute to the formation of alveolar edema.

    View details for Web of Science ID 000079229000026

    View details for PubMedID 10099704

  • Investigation of the anticoagulant mechanisms of a covalent antithrombin-heparin complex JOURNAL OF BIOLOGICAL CHEMISTRY Berry, L., Stafford, A., Fredenburgh, J., O'Brodovich, H., Mitchell, L., Weitz, J., Andrew, M., Chan, A. K. 1998; 273 (52): 34730-34736

    Abstract

    Recently, we developed a covalent antithrombin-heparin complex (ATH) as a possible treatment for respiratory distress syndrome. ATH reacted rapidly with thrombin and efficiently catalyzed the inhibition of either thrombin or factor Xa by exogenous antithrombin. In order to investigate mechanisms for the conjugate's unusual anticoagulant properties, changes in fluorescence due to covalent linkage or addition of exogenous antithrombin were studied in relation to reaction with thrombin derivatives or factor Xa. The emission spectrum of ATH was similar to that of antithrombin plus heparin mixtures. ATH quickly inhibited thrombin or factor Xa activities, as measured by a fluorogenic substrate. Fluorescein-labeled heparin was displaced from either thrombin or active site blocked thrombin by ATH, indicating that thrombin must bind to the conjugate's heparin moiety. Interaction of thrombin with ATH's heparin component was confirmed by a slow reaction rate of conjugate with a thrombin mutant that has weak heparin binding. Total intrinsic fluorescence increased when exogenous antithrombin was added to ATH, indicating that the catalytic mechanism may occur through a second inhibitor binding site. Thus, ATH reacts directly with thrombin through a bridge mechanism and probably catalyzes the reaction of thrombin with antithrombin by a second binding sequence on its heparin chain.

    View details for Web of Science ID 000077719700014

    View details for PubMedID 9856996

  • Covalent heparin cofactor II-heparin and heparin cofactor II-dermatan sulfate complexes - Characterization of novel anticoagulants JOURNAL OF BIOLOGICAL CHEMISTRY Monagle, P., Berry, L., O'Brodovich, H., Andrew, M., Chan, A. 1998; 273 (50): 33566-33571

    Abstract

    Heparin cofactor II is a naturally occurring anticoagulant that acts by specifically inhibiting thrombin and is facilitated by the binding of glycosaminoglycans such as heparin and dermatan sulfate. In vivo, heparin cofactor II-glycosaminoglycan complexes dissociate, leaving the inhibitor less active in its ability to function as a component of the anticoagulation pathway. We have produced permanently activated heparin cofactor II molecules by covalent linkage to either heparin or dermatan sulfate. Covalent heparin cofactor II-heparin and heparin cofactor II-dermatan sulfate complexes had catalytic antithrombin activities similar to those of the corresponding starting heparin and dermatan sulfate (86% and 110% of standard heparin and dermatan sulfate activity, respectively). Both heparin cofactor II-heparin and heparin cofactor II-dermatan sulfate had fast bimolecular rate constants of 1.4 x 10(7) M-1 s-1 and 1.3 x 10(7) M-1 s-1, respectively, for reaction with thrombin. The intravenous half-life of the covalent complexes in rabbits was significantly longer than that of free heparin or dermatan sulfate (4.4, 3.4, 0.33, and 0.50 h for heparin cofactor II-heparin, heparin cofactor II-dermatan sulfate, heparin, and dermatan sulfate, respectively). Given their unique properties, these conjugates may have a clinical application for long term, selective inhibition of thrombin.

    View details for Web of Science ID 000077462500069

    View details for PubMedID 9837939

  • Relation between alpha, beta, and gamma human amiloride-sensitive epithelial Na+ channel mRNA levels and nasal epithelial potential difference in healthy men AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE Otulakowski, G., Flueckiger-Staub, S., Ellis, L., Ramlall, K., Staub, O., Smith, D., Durie, P., O'Brodovich, H. 1998; 158 (4): 1213-1220

    Abstract

    To analyze messenger RNA (mRNA) levels for the alpha, beta, and gamma subunits of the human amiloride-sensitive epithelial Na+ channel (hENaC) in respiratory epithelia, we developed a competitive quantitative reverse transcriptase-polymerase chain reaction (QRT-PCR) assay specific for each subunit, using two human respiratory epithelial-cell lines. We next determined the relation between hENaC mRNA levels and the biologic activity of the hENaC in the respiratory epithelium of eight normal men. The electrical potential difference (PD) between the epithelium of the inferior nasal turbinate and the subcutaneous space was measured, using control and amiloride (100 microM) solutions. QRT-PCR measurement of hENaC-subunit mRNAs and epithelial-specific cytokeratin 18 mRNA allowed us to normalize hENaC expression to epithelial-cell RNA. Respective values for alpha, beta, and gamma hENaC mRNA levels in epithelium obtained at the site of maximal PD were 39 +/- 4.0, 7.5 +/- 0.92, and 1.8 +/- 0.25 attomol/fmol cytokeratin mRNA, respectively. Respiratory epithelial PD exhibited a significant negative correlation with gamma hENaC (r2 = 0.72, p < 0.01), tended to increase with increasing alpha hENaC, and was unaffected by beta hENaC mRNA levels. Our results suggest that hENaC activity in vivo is influenced by expression of the gene for gamma hENaC. The assay used in the study provides a useful tool for evaluating Na+-channel expression in clinically relevant patient populations.

    View details for Web of Science ID 000076453300032

    View details for PubMedID 9769284

  • O-2-induced ENaC expression is associated with NF-kappa B activation and blocked by superoxide scavenger AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Rafii, B., Tanswell, A. K., O'Tulakowski, G., Pitkanen, O., Belcastro-Taylor, R., O'Brodovich, H. 1998; 275 (4): L764-L770

    Abstract

    Cultured rat fetal distal lung epithelial cells (FDLEs), when switched from fetal (3%) to postnatal (21%) O2 concentrations, have increased epithelial Na+ channel (ENaC) mRNA levels and amiloride-sensitive Na+ transport [O. Pitkänen, A. K. Tanswell, G. Downey, and H. O'Brodovich. Am. J. Physiol. 270 (Lung Cell. Mol. Physiol. 14): L1060-L1066, 1996]. The mechanisms by which O2 mediates these effects are unknown. After isolation, FDLEs were kept at 3% O2 overnight, then switched to 21% O2 (3-21% O2 group) or maintained at 3% O2 (3-3% O2 group) for 48 h. The amiloride-sensitive short-circuit current (Isc) in the 3-21% O2 group was double that in the 3-3% O2 group. Amiloride-sensitive Isc could not be induced by medium conditioned by 21% O2-exposed FDLEs but was reversed by returning the cells to 3% O2. Neither the cyclooxygenase inhibitor ibuprofen, liposome-encapsulated catalase, nor hydroperoxide scavengers (U-74389G or Trolox) blocked the O2-induced amiloride-sensitive Isc. In contrast, the cell-permeable superoxide scavenger tetramethylpiperidine-N-oxyl (TEMPO) eliminated the O2-induced increases in amiloride-sensitive Isc and ENaC mRNA levels. The switch from 3 to 21% O2 induced the transcription factor nuclear factor-kappaB, which could also be blocked by TEMPO. We conclude that 1) the O2-induced increase in amiloride-sensitive Isc is reversible and 2) the O2-induced increase in amiloride-sensitive Isc and ENaC mRNA levels is associated with activation of nuclear factor-kappaB and may be mediated, at least in part, by superoxide.

    View details for Web of Science ID 000076268200015

    View details for PubMedID 9755109

  • Influence of mechanical stretch on thrombin regulation by fetal mixed lung cells AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Chan, A. K., Baranowski, B., Berry, L., Liu, M. Y., Rafii, B., Post, M., O'Brodovich, H., Monagle, P., Andrew, M. 1998; 19 (3): 419-425

    Abstract

    Respiratory distress syndrome (RDS) is characterized by intrapulmonary fibrin deposition, which can adversely affect surfactant function, and stimulate fibroblast proliferation, which may contribute to the development of bronchopulmonary dysplasia (BPD). We speculated that the premature lung may have impaired regulation of thrombin, thus making preterm infants susceptible to fibrin formation within the lung. Therefore, we studied the effect of stretch, which simulates fetal breathing movements (FBMs), on the generation and inhibition of a key hemostatic enzyme-thrombin-by rat fetal mixed lung cells (FMLCs). Our results showed that stretch induced glycosaminoglycan production with increased antithrombin activity due to an increase in the concentration of active chondroitin sulfate. Stretch downregulated secretion of tissue factor procoagulant activity, which may lead to decreased thrombin generation on the surface of FMLCs. Overall, stretch enhanced the local control of thrombin by FMLCs. These results suggest that premature infants, who will have experienced less FBM, may have impaired thrombin regulation. Impaired thrombin regulation likely contributes to increased fibrin deposition and, potentially, the development of BPD.

    View details for Web of Science ID 000075976000009

    View details for PubMedID 9730869

  • Liposome-mediated transfection of fetal lung epithelial cells: DNA degradation and enhanced superoxide toxicity AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Tanswell, A. K., Staub, O., Iles, R., Belcastro, R., Cabacungan, J., Sedlackova, L., Steer, B., Wen, Y. X., Hu, J., O'Brodovich, H. 1998; 275 (3): L452-L460

    Abstract

    Cationic liposomes, 1:1 (mol/mol) 1,2-dioleoyldimethylammonium chloride-1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, were used to transfect primary cultures of distal rat fetal lung epithelial cells with pCMV4-based plasmids. A DNA-to-lipid ratio of 1:10 to 1:15 (wt/wt) optimized DNA uptake over a 24-h exposure. At a fixed DNA-to-lipid ratio of 1:15, chloramphenicol acetyltransferase (CAT) reporter gene expression declined at lipid concentrations > 2.5 nmol/cm2 cell surface area, whereas DNA uptake remained concentration dependent. CAT expression peaked 48 h after removal of the liposome-DNA complex, declining thereafter. Reporter gene expression was increased, and supercoiled cDNA degradation was reduced by the addition of 0.2 mM nicotinamide and 10 microM chloroquine. Rat fetal lung epithelial cells transfected with two different expression cassettes had an increased susceptibility to superoxide-mediated cytotoxicity. This could be attributed to a nonspecific delivery of exogenous DNA or some other copurified factor. The DNA-dependent increase in superoxide-mediated cytotoxicity, but not basal levels of cytotoxicity, was inhibited by the addition of 0.2 mM nicotinamide and 10 microM chloroquine.

    View details for Web of Science ID 000075745200004

    View details for PubMedID 9728039

  • Effect of a novel covalent antithrombin-heparin complex on thrombin generation on fetal distal lung epithelium AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Chan, A. K., Berry, L., Mitchell, L., Baranowski, B., O'Brodovich, H., Andrew, M. 1998; 274 (6): L914-L921

    Abstract

    Respiratory distress syndrome is characterized by fibrin deposition in the lung. Fibrin adversely affects surfactant function and stimulates proliferation of fibroblasts. There is evidence that these properties may be important to the development of bronchopulmonary dysplasia. Despite successful initial treatment of neonatal respiratory distress syndrome with surfactant, the incidence of bronchopulmonary dysplasia has not decreased. In previous studies, it has been demonstrated that rat fetal distal lung epithelium (FDLE) possesses both procoagulant and anticoagulant properties. In this report, we have demonstrated (using factor VII-deficient plasma) that tissue factor is expressed on the FDLE surface and promotes thrombin generation. To regulate thrombin within this procoagulant environment, we have developed a novel anticoagulant, antithrombin-heparin covalent complex (ATH) that can be retained within the lung after intrapulmonary instillation. We have demonstrated that ATH was superior to antithrombin plus standard heparin in suppressing thrombin generation (P < 0.001) and prothrombin consumption (P < 0.01) in recalcified defibrinated plasma on the surface of FDLE. Further studies with ATH in vivo need to be performed.

    View details for Web of Science ID 000073905400006

    View details for PubMedID 9609730

  • Modulation of aquaporin 4 and the amiloride-inhibitable sodium channel in perinatal rat lung epithelial cells AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Ruddy, M. K., Drazen, J. M., Pitkanen, O. M., Rafii, B., O'Brodovich, H. M., Harris, H. W. 1998; 274 (6): L1066-L1072

    Abstract

    During the perinatal period, a dramatic reversal of lung transepithelial ion and water transport occurs that involves the amiloride-inhibitable Na+ channel (ENaC). Aquaporin (AQP) water channel proteins facilitate cell membrane water transport. We now report that AQP-4, localized to basolateral membranes of airway epithelial cells, increases its mRNA expression in developing lung eightfold during the 2 days before birth to reach a peak on the first postnatal day in the lungs but not in brains or kidneys of neonatal rats. AQP-4 and the alpha-, beta-, and gamma-subunits of ENaC are both expressed by cultured rat fetal distal lung epithelial (FDLE) cells. AQP-4 and ENaC expression increase in FDLE cells cultured on uncoated permeant filters compared with matched control cells cultured on filters containing extracellular matrix derived from fetal lung epithelial cells. Similarly, AQP-4 expression increases in FDLE cells exposed to 21% O2 compared with cells exposed to 3% O2. These data demonstrate that AQP-4 expression is highest on the first day after birth in neonatal rat lungs. Exposure to ambient 21% O2 may contribute to increases in AQP-4 and ENaC expression to facilitate water transport across neonatal airway epithelia in the immediate postnatal period.

    View details for Web of Science ID 000073905400023

    View details for PubMedID 9609747

  • Significance of ion transport during lung development and in respiratory disease of the newborn ANNALS OF MEDICINE Pitkanen, O. M., O'Brodovich, H. M. 1998; 30 (2): 134-142

    Abstract

    Active ion transport plays a critical role in the liquid movement across the fetal and perinatal lung epithelium. The fetal lung liquid production is coupled with active secretion of Cl- into the luminal space. The potential for fluid absorbing mechanisms related to active Na+ transport from the apical to the basolateral side of the epithelium appears near the end of gestation. At birth there is a dramatic change of environment with commencement of air-breathing, sudden increase in oxygen partial pressure (PO2) and profound changes in the pulmonary circulation. A concurrent switch from fluid secretion to maintenance of low amounts of alveolar fluid is another major physiological adjustment taking place in the perinatal distal lung epithelium. The fluid-absorbing mechanism is a result of a well-synchronized co-operation between the basolateral membrane Na-K-ATPase and the apical membrane Na+ channels and it promotes salt and water movement from the airspace. Inability of the fetal lung epithelium to switch from fluid secretion to Na+ transport-dependent absorption seems to be an important factor adversely contributing to the respiratory distress of the newborn premature infant.

    View details for Web of Science ID 000073666600002

    View details for PubMedID 9667791

  • Inhibition of amiloride-sensitive sodium-channel activity in distal lung epithelial cells by nitric oxide AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Ding, J. W., Dickie, J., O'Brodovich, H., Shintani, Y., Rafii, B., Hackam, D., Marunaka, Y., Rotstein, O. D. 1998; 274 (3): L378-L387

    Abstract

    Distal lung epithelial cells (DLECs) play an active role in fluid clearance from the alveolus by virtue of their ability to actively transport Na+ from the alveolus to the interstitial space. The present study evaluated the ability of activated macrophages to modulate the bioelectric properties of DLECs. Low numbers of lipopolysaccharide (LPS)-treated macrophages were able to significantly reduce amiloride-sensitive short-circuit current (Isc) without affecting total Isc or monolayer resistance. This was associated with a rise in the flufenamic acid-sensitive component of the Isc. The effect was reversed by the addition of N-monomethyl-L-arginine to the medium, implying a role for nitric oxide. We hypothesized that macrophages exerted their effect by expressing inducible nitric oxide synthase (iNOS) in DLECs. The products of LPS-treated macrophages increased the levels of iNOS protein and mRNA transcripts in DLECs as well as causing a rise in iNOS activity. Immunofluorescence microscopy of LPS-stimulated macrophage-DLEC cocultures with anti-nitrotyrosine antibodies provided evidence for the generation of peroxynitrite in macrophages but not in DLECs. These data indicate that activated macrophages in the lung may contribute to impaired resolution of acute respiratory distress syndrome and suggest a novel mechanism whereby nitric oxide might alter cell function by altering its ion-transporting phenotype.

    View details for Web of Science ID 000072415800010

    View details for PubMedID 9530173

  • Female gender hormones regulate mRNA levels and function of the rat lung epithelial Na channel AMERICAN JOURNAL OF PHYSIOLOGY-CELL PHYSIOLOGY Sweezey, N., Tchepichev, S., Gagnon, S., Fertuck, K., O'Brodovich, H. 1998; 274 (2): C379-C386

    Abstract

    The epithelial Na channel (ENaC) plays a critical role in the active reabsorption of alveolar fluid at the time of birth or during pulmonary edema. Although rat (r) ENaC is regulated by glucocorticoids during fetal development, there are no data regarding the influence of gender hormones on ENaC expression or function. We report higher levels of mRNAs encoding the alpha-rENaC subunit or the cystic fibrosis transmembrane conductance regulator (CFTR) in the lungs of nonpregnant adult female relative to adult male Wistar rats. Combined, but not separate, administration of progesterone and 17 beta-estradiol increased mRNA levels encoding alpha-rENaC, gamma-rENaC, and CFTR within 24 h. We also found a dose-dependent increase in rENaC functional activity (as assessed by the amiloride-sensitive short-circuit current across primary monolayer cultures of alveolar epithelial cells mounted in Ussing chambers) after a 5-day incubation of cells in medium containing progesterone and 17 beta-estradiol. These findings suggest a gender-dependent influence on the lung's ability to recover from pulmonary edema and on the degree of airway fluid hydration in cystic fibrosis.

    View details for Web of Science ID 000071884000012

    View details for PubMedID 9486127

  • Comparison between intratracheal and intravenous administration of liposome-DNA complexes for cystic fibrosis lung gene therapy GENE THERAPY Griesenbach, U., Chonn, A., Cassady, R., Hannam, V., Ackerley, C., Post, M., Tanswell, A. K., Olek, K., O'Brodovich, H., TSUI, L. C. 1998; 5 (2): 181-188

    Abstract

    Intratracheal (i.t.) and intravenous (i.v.) delivery of DNA-vector formulations are two strategies to obtain gene transfer to the lung, it is still uncertain, however, which of these two modes of delivery will be more effective in the treatment of cystic fibrosis and other lung diseases. In this study, we attempted to optimize formulations of the cationic liposome DODAC:DOPE (dioleoyldimethylammonium-chloride: dioleoylphosphatidylethanolamine) complexed to plasmids encoding chloramphenicol acetyltransferase for i.t. and i.v. injection into CD-2 mice and compared the two methods. Our results showed that both methods conferred reporter gene expression in the lung that was significantly higher relative to injection of plasmid DNA alone. Expression using either mode of administration was maximal 24 h after injection and declined to around 10% of day 1 levels 2 weeks after injection. For i.v. delivery of DODAC. DOPE-DNA complexes multilamellar vesicles were more effective than large unilamellar vesicles in all organs investigated. Recombinant DNA could be detected in the distal lung region following either route of administration. However, i.t. administration predominantly led to DNA deposition in epithelial cells lining the bronchioles, e.g. in clara cells, whereas i.v. administration resulted in DNA deposition in the alveolar region of the lung including type II alveolar epithelial cells.

    View details for Web of Science ID 000072033100006

    View details for PubMedID 9578837

  • Development of an epithelium-specific expression cassette with human DNA regulatory elements for transgene expression in lung airways PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Chow, Y. H., O'Brodovich, H., Plumb, J., Wen, Y. X., Sohn, K. J., Lu, Z., Zhang, F. Y., LUKACS, G. L., Tanswell, A. K., Hui, C. C., Buchwald, M., Hu, J. 1997; 94 (26): 14695-14700

    Abstract

    The efficient expression of therapeutic genes in target cells or tissues is an important component of efficient and safe gene therapy. Utilizing regulatory elements from the human cytokeratin 18 (K18) gene, including 5' genomic sequences and one of its introns, we have developed a novel expression cassette that can efficiently express reporter genes, as well as the human cystic fibrosis transmembrane conductance regulator (CFTR) gene, in cultured lung epithelial cells. CFTR transcripts expressed from the native K18 enhancer/promoter include two alternative splicing products, due to the activation of two cryptic splice sites in the CFTR coding region. Modification of the K18 intron and CFTR cDNA sequences eliminated the cryptic splice sites without changing the CFTR amino acid sequence, and led to enhanced CFTR mRNA and protein expression as well as biological function. Transgenic expression analysis in mice showed that the modified expression cassette can direct efficient and epithelium-specific expression of the Escherichia coli LacZ gene in the airways of fetal lungs, with no detectable expression in lung fibroblasts or endothelial cells. This is the first expression cassette which selectively directs lung transgene expression for CFTR gene therapy to airway epithelia.

    View details for Web of Science ID 000071182800085

    View details for PubMedID 9405675

  • Lung disease in mice with cystic fibrosis JOURNAL OF CLINICAL INVESTIGATION Kent, G., Iles, R., Bear, C. E., Huan, L. J., Griesenbach, U., McKerlie, C., Frndova, H., Ackerley, C., Gosselin, D., Radzioch, D., O'Brodovich, H., TSUI, L. C., Buchwald, M., Tanswell, A. K. 1997; 100 (12): 3060-3069

    Abstract

    The leading cause of mortality and morbidity in humans with cystic fibrosis is lung disease. Advances in our understanding of the pathogenesis of the lung disease of cystic fibrosis, as well as development of innovative therapeutic interventions, have been compromised by the lack of a natural animal model. The utility of the CFTR-knockout mouse in studying the pathogenesis of cystic fibrosis has been limited because of their failure, despite the presence of severe intestinal disease, to develop lung disease. Herein, we describe the phenotype of an inbred congenic strain of CFTR-knockout mouse that develops spontaneous and progressive lung disease of early onset. The major features of the lung disease include failure of effective mucociliary transport, postbronchiolar over inflation of alveoli and parenchymal interstitial thickening, with evidence of fibrosis and inflammatory cell recruitment. We speculate that the basis for development of lung disease in the congenic CFTR-knockout mice is their observed lack of a non-CFTR chloride channel normally found in CFTR-knockout mice of mixed genetic background.

    View details for Web of Science ID 000071149100019

    View details for PubMedID 9399953

  • Covalent antithrombin-heparin complexes with high anticoagulant activity - Intravenous, subcutaneous, and intratracheal administration JOURNAL OF BIOLOGICAL CHEMISTRY Chan, A., Berry, L., OBRODOVICH, H., Klement, P., Mitchell, L., Baranowski, B., Monagle, P., Andrew, M. 1997; 272 (35): 22111-22117

    Abstract

    Although heparin has been used clinically for prophylaxis and treatment of thrombosis, it has suffered from problems such as short duration within compartments in vivo that require long term anticoagulation. A covalent antithrombin-heparin complex has been produced with high anticoagulant activity and a long half-life relative to heparin. The product had high anti-factor Xa and antithrombin activities compared with noncovalent mixtures of antithrombin and heparin (861 and 753 units/mg versus 209 and 198 units/mg, respectively). Reaction with thrombin was rapid with bimolecular and second order rate constants of 1.3 x 10(9) M-1 s-1 and 3.1 x 10(9) M-1 s-1, respectively. The intravenous half-life of the complex in rabbits was 2.6 h as compared with 0.32 h for similar loads of heparin. Subcutaneous injection of antithrombin-heparin resulted in plasma levels (peaking at 24-30 h) that were still detectable 96 h post-injection. Given the increased lifetime in these vascular and intravascular spaces, use of the covalent complex in the lung was investigated. Activity of antithrombin-heparin instilled into rabbit lungs remained for 48 h with no detection of any complex systemically. Thus, this highly active agent has features required for pulmonary sequestration as a possible treatment for thrombotic diseases such as respiratory distress syndrome.

    View details for Web of Science ID A1997XT85000066

    View details for PubMedID 9268354

  • The effect of brefeldin A on terbutaline-induced sodium absorption in fetal rat distal lung epithelium PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY Ito, Y., Niisato, N., OBRODOVICH, H., Marunaka, Y. 1997; 434 (4): 492-494

    Abstract

    We studied the effect of brefeldin A, which inhibits the intracellular trafficking of membrane proteins from the cytosolic pool to the cell surface, on terbutaline (a beta2-specific adrenergic agonist)-induced alterations in ion transport by primary monolayer cultures of fetal rat distal lung epithelium. The amiloride-sensitive short circuit current (Isc) increased 2.5-fold 50 min after application of terbutaline (10 microM) from basolateral side; this response was abolished by pretreatment with brefeldin A (1 microg/ml). Brefeldin A did not suppress the Na+/K+ pump capacity. Single channel patch clamp experiments demonstrated that terbutaline increased the density of amiloride-sensitive Na+-permeable nonselective cation channels on the apical cell membrane and this action was blocked by brefeldin A. These observations suggest that beta2-specific adrenergic agonists promote the trafficking of amiloride sensitive Na+-permeable nonselective cation channels to the apical cell surface.

    View details for Web of Science ID A1997XL55100020

    View details for PubMedID 9211817

  • The present and future role of gene therapy in the newborn. Current opinion in pediatrics Tanswell, A. K., O'Brodovich, H. M. 1997; 9 (2): 141-145

    Abstract

    There are currently a large number of ongoing gene therapy trials in North America and Europe. These trials have almost exclusively involved patients with inherited lethal disorders or malignancies. Although there are significant ethical and safety issues that remain unresolved, it seems inevitable that this technology will soon be adapted for use in lethal, or potentially lethal, fetal and neonatal diseases. If ethical and safety issues can be resolved, a wide spectrum of nonlethal acute and chronic diseases could also benefit from this form of therapy. The purpose of this brief review is to provide an overview of current approaches to gene delivery, their successes, and their limitations. Where possible, the discussion has focused on conditions that are recognized in fetal or neonatal life, to give the reader some sense of the potential scope for this form of therapy.

    View details for PubMedID 9204241

  • Respiratory distress syndrome: The importance of effective transport JOURNAL OF PEDIATRICS OBRODOVICH, H. M. 1997; 130 (3): 342-344

    View details for Web of Science ID A1997WM26400003

    View details for PubMedID 9063404

  • Immature epithelial nat channel expression is one of the pathogenetic mechanisms leading to human neonatal respiratory distress syndrome PROCEEDINGS OF THE ASSOCIATION OF AMERICAN PHYSICIANS OBRODOVICH, H. M. 1996; 108 (5): 345-355

    Abstract

    Noninfective acute respiratory disease develops in approximately 1% of all newborn infants and results in their admission to a critical care unit. Transient tachypnea of the newborn occurs as a result of a delay in the clearance of fetal lung liquid; however, respiratory distress syndrome, typically thought to be exclusively a problem of relative surfactant deficiency, is now suspected to be characterized by an even greater air space fluid burden from the inability to absorb fetal lung liquid. In vivo experiments have demonstrated that the lung epithelium secretes Cl and fluid throughout gestation and develops the ability to actively reabsorb Na+ only during late gestation. At birth, the mature lung switches from active Cl- (fluid) secretion to active Na+ (fluid) absorption in response to circulating catecholamines. Changes in oxygen tension augment the Na(+)-transporting capacity of the epithelium and increase gene expression for the epithelial Na+ channel (ENaC). The inability of the immature fetal lung to switch from fluid secretion to fluid absorption results, at least in large part, from an immaturity in the expression of ENaC, which can be upregulated by glucocorticosteroids. Both pharmacological blockade of the lung's epithelial Na+ channel and genetic knockout experiments using mice deficient in the ENaC pore-forming subunit have demonstrated the critical physiological importance of lung Na+ transport at birth. When Na+ transport is ineffective, newborn animals develop respiratory distress and hypoxemia, retain their fetal lung liquid and, in the case of the ENaC knockout mice, die. Bioelectrical studies of human infants' nasal epithelia demonstrate that both transient tachypnea of the newborn and respiratory distress syndrome have defective amiloride-sensitive Na+ transport. These results suggest that neonatal respiratory distress syndrome has, in addition to a relative deficiency in surfactant, defective Na+ transport, which plays a mechanistic role in the development of the disease.

    View details for Web of Science ID A1996VP84500001

    View details for PubMedID 8902878

  • Increased Po-2 alters the bioelectric properties of fetal distal lung epithelium AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Pitkanen, O., Tanswell, A. K., Downey, G., OBRODOVICH, H. 1996; 270 (6): L1060-L1066

    Abstract

    At birth the lung must efficiently clear the liquid from its air spaces and permanently convert from a fluid-secreting to a fluid-absorbing organ. When primary cultures of rat fetal distal lung epithelium (FDLE) grown on permeable supports were switched from a fetal (3%) to a postnatal (21%) oxygen environment, there was an increase in epithelial permeability as reflected by a dose-dependent decline in transepithelial resistance (Rt) 4 h later (3% = 239 +/- 19 omega.cm2; 21% = 170 +/- 28 omega.cm2; 50% = 98 +/- 20 omega.cm2; P < 0.05). The effect was transient, since monolayers initially maintained at 3% and switched to these higher oxygen concentrations subsequently had Rt values comparable to the 3% group at 48 h (3% = 153 +/- 17 omega.cm2; 21% = 181 +/- 19 omega.cm2; 50% = 192 +/- 21 omega.cm2; P = NS). Changes in Rt were associated with expected changes in the histological appearance of the interepithelial tight junctions, but intracellular actin content and distribution remained constant. Amiloride-sensitive equivalent short-circuit current increased within 18 h, with further increases after 48 h of exposure to postnatal oxygen concentrations. Ion substitution experiments suggested diminished FDLE Cl transport and increased Na transport. The amount of FDLE-alpha, -beta, and -gamma rat epithelial Na channel mRNA increased within 48 h of increasing the ambient oxygen concentration. These results suggest that the physiological increase in alveolar Po2 at birth is, at least in part, responsible for distal lung's permanent switch from Cl secretion to Na absorption at birth.

    View details for Web of Science ID A1996UT99800022

    View details for PubMedID 8764233

  • LUNG EPITHELIAL NA CHANNEL SUBUNITS ARE DIFFERENTIALLY REGULATED DURING DEVELOPMENT AND BY STEROIDS AMERICAN JOURNAL OF PHYSIOLOGY-CELL PHYSIOLOGY Tchepichev, S., Ueda, J., Canessa, C., Rossier, B. C., OBRODOVICH, H. 1995; 269 (3): C805-C812

    Abstract

    Because the alpha-subunit of the rat lung epithelial Na channel (rENaC) is not expressed until late fetal gestation, the developmental immaturity of alpha-rENaC may be involved in the premature fetal lung's inability to mount a Na-absorptive response to appropriate agonists. As previous work has shown that the beta- and gamma-rENaC subunits of the Na channel are required for maximal alpha-rENaC activity, we determined their developmental expression in the fetal lung. In addition, because thyroid and corticosteroid therapy can mature the in vivo fetal lamb lung's ability to transport Na, we wished to determine whether such treatment increased the expression of alpha-, beta-, and gamma-rENaC. Lungs were harvested from normal rat fetuses of 17 through 22 days gestation (term = 22 days), normal rat pups during the first week of life, and adult rats. Initial expression of alpha-rENaC was detected at 19 days gestation and progressively increased in utero. beta- and gamma-rENaC mRNA were not detected until 21 and 22 days gestation, and then only at very low levels. During the first week after birth, the levels of alpha-rENaC declined, whereas beta- and gamma-rENaC mRNA levels increased. This pre- and postnatal pattern of alpha-rENaC expression correlates with the endogenous glucocorticosteroid levels in the fetus and the rat pup's early postnatal corticosteroid resistance. Combined or separate treatment of pregnant rats (16 through 22 days gestational age) with thyroid-releasing hormone (TRH) and/or dexamethasone (Dex) for 48 h showed that Dex, but not TRH, could increase fetal lung alpha-rENaC mRNA levels.(ABSTRACT TRUNCATED AT 250 WORDS)

    View details for Web of Science ID A1995RV40300033

    View details for PubMedID 7573414

  • LUNG FLUID RESTRICTION AFFECTS GROWTH BUT NOT AIRWAY BRANCHING OF EMBRYONIC RAT LUNG INTERNATIONAL JOURNAL OF DEVELOPMENTAL BIOLOGY Souza, P., OBRODOVICH, H., Post, M. 1995; 39 (4): 629-637

    Abstract

    During the later stages of fetal life lung growth and development is dependent upon a variety of factors, including a normal amount of liquid within the lung's lumen. To investigate whether embryonic lung epithelium secretes fluid and whether lung liquid is essential for proper embryonic airway lung growth and branching, we incubated 12-day rat lung primordia (term=22 days) in submersion culture in serum-free medium for 48 h in room air (21% O2/5% CO2). Under these conditions, lung growth and branching proceeded but at a slower rate when compared to growth and branching in vivo. Neither addition of serum nor incubation in a fetal O2 concentration (=3% O2) changed the growth rate or the degree of branching in vitro. The luminal area of the explant increased progressively with time in culture. Inhibitors of active Cl- secretion (200 microM bumetanide and 1 mM furosemide) significantly reduced the lumen size compared with control. A similar effect was noted with lung explants of 13-15 days of gestation. Branching morphogenesis was not impaired by lung fluid reduction. Reduction of luminal liquid significantly increased DNA synthesis of 12-day embryonic lung explants, but this effect of bumetanide and furosemide on DNA synthesis was reversed when 13-15 day lung explants were used. These data suggest that embryonic lung epithelium secretes fluid and that the secretion is chloride dependent. Lung fluid is involved in controlling lung growth but not branching of the embryonic rat lung.

    View details for Web of Science ID A1995TD77600009

    View details for PubMedID 8619961

  • FETAL LUNG CELL-DERIVED MATRIX ALTERS DISTAL LUNG EPITHELIAL ION-TRANSPORT AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Pitkanen, O. M., Tanswell, A. K., OBRODOVICH, H. M. 1995; 268 (5): L762-L771

    Abstract

    Extracellular matrix (ECM) synthesized by the fetal mesenchymal cells provides a supporting structure for the growing airways and is important for airway branching and in the differentiation of the primitive epithelium. We studied whether ECM, in addition to its structural role in lung interstitium, influences the ion transport of rat fetal distal lung epithelial cells (FDLE). FDLE monolayers were cultured on two different fetal mixed lung cell (MLC)-derived matrix preparations and studied in Ussing chambers. FDLE on MLC matrix had an increased resting equivalent short-circuit current (Ieq). Amiloride (10(-4) M apically) decreased the Ieq significantly in all the FDLE monolayers. The residual Ieq was significantly larger in FDLE grown on MLC matrix (increased by 150 and 80% under baseline and beta 2-agonist-stimulated conditions, respectively) than on control filters and filters coated with type I collagen, and type IV collagen, laminin, or fibronectin. The matrix produced by MLC isolated at an earlier gestational stage decreased the FDLE's sensitivity to amiloride. The increased amiloride-insensitive Ieq was only modestly affected by the Na+/K+/Cl- cotransport inhibitor bumetanide (10(-4) M basally) but was abolished when the [Cl-] of the bathing solution was reduced to 10 mM. These observations demonstrated that MLC elaborated ECM is able to change the nature of the ion transport of FDLE. ECM may be an important factor governing the ion transporting phenotype of fetal type II alveolar epithelial cells.

    View details for Web of Science ID A1995QW58100008

    View details for PubMedID 7762679

  • The role of active Na+ transport by lung epithelium in the clearance of airspace fluid. New horizons (Baltimore, Md.) O'Brodovich, H. M. 1995; 3 (2): 240-247

    Abstract

    Clinical and laboratory-based studies of pulmonary edema have usually focused on the mechanisms responsible for the production of the edema and how therapeutic maneuvers can oppose or treat such processes. Recently, there has been increasing interest in the mechanisms involved in the clearance of airspace fluids. These studies have demonstrated that active transport of Na+ by the distal lung epithelium plays an important physiologic role in the clearance of pulmonary edema fluid. Specifically, the ability of the lung to clear its fluid by active transport processes correlates with survival from high-pressure or high-permeability pulmonary edema. Also, studies have shown that immaturity of Na+ transport processes and, specifically, inadequate expression of Na+ channels contribute to the pathogenesis of respiratory distress syndrome in the newborn.

    View details for PubMedID 7583165

  • AN SH3 BINDING REGION IN THE EPITHELIAL NA+ CHANNEL (ALPHA-RENAC) MEDIATES ITS LOCALIZATION AT THE APICAL MEMBRANE EMBO JOURNAL Rotin, D., BARSAGI, D., OBRODOVICH, H., Merilainen, J., LEHTO, V. P., Canessa, C. M., Rossier, B. C., Downey, G. P. 1994; 13 (19): 4440-4450

    Abstract

    The amiloride-sensitive Na+ channel constitutes the rate-limiting step for Na+ transport in epithelia. Immunolocalization and electrophysiological studies have demonstrated that this channel is localized at the apical membrane of polarized epithelial cells. This localization is essential for proper channel function in Na+ transporting epithelia. In addition, the channel has been shown to associate with the cytoskeletal proteins ankyrin and alpha-spectrin in renal epithelia. However, the molecular mechanisms underlying the cytoskeletal interactions and apical membrane localization of this channel are largely unknown. In this study we show that the putative pore forming subunit of the rat epithelial (amiloride-sensitive) Na+ channel (alpha ENaC) binds to alpha-spectrin in vivo, as determined by co-immunoprecipitation. This binding is mediated by the SH3 domain of alpha-spectrin which binds to a unique proline-rich sequence within the C-terminal region of alpha rENaC. Accordingly, the C-terminal region is sufficient to mediate binding to intact alpha-spectrin from alveolar epithelial cell lysate. When microinjected into the cytoplasm of polarized primary rat alveolar epithelial cells, a recombinant fusion protein containing the C-terminal proline-rich region of alpha rENaC localized exclusively to the apical area of the plasma membrane, as determined by confocal microscopy. This localization paralleled that of alpha-spectrin. In contrast, microinjected fusion protein containing the N-terminal (control) protein of alpha rENaC remained diffuse within the cytoplasm. These results suggest that an SH3 binding region in alpha rENaC mediates the apical localization of the Na+ channel. Thus, cytoskeletal interactions via SH3 domains may provide a novel mechanism for retaining proteins in specific membranes of polarized epithelial cells.

    View details for Web of Science ID A1994PL72000002

    View details for PubMedID 7925286

  • THROMBIN INHIBITION BY FETAL DISTAL LUNG EPITHELIUM IS DIFFERENT IN FETAL AND ADULT PLASMA AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Andrew, M., Berry, L., OBRODOVICH, H. 1994; 11 (1): 35-41

    Abstract

    Intra-alveolar fibrin deposition is a cardinal feature of neonatal respiratory distress syndrome and likely contributes to short-term and long-term morbidity. Previous studies have shown that fetal distal lung epithelial cell (FDLE) surfaces express procoagulant activity when incubated with adult plasma and may therefore provide one mechanism by which fibrin is generated. However, plasma concentrations of prothrombin and thrombin inhibitors differ significantly at birth and during the first weeks of life compared with adult values. Therefore, we measured thrombin-generating capacity and inhibitor complex formation in cord and adult plasma incubated in the presence of FDLE. Although starting cord plasma concentrations of prothrombin were 43% of adult values, the amount of thrombin generated was decreased by only 21%. When cord plasma concentrations of prothrombin were selectively increased to adult values, the amount of thrombin generated surpassed adult plasma by 89%. The latter observations suggested that thrombin inhibition was impaired in cord plasma compared with adult plasma and supplementation of cord plasma with antithrombin III (ATIII) as well as prothrombin returned thrombin generation to adult levels. However, the percentage of thrombin complexed to inhibitors (59%) at the completion of the experiments was similar in cord, cord plus prothrombin, cord plus prothrombin plus ATIII, and adult plasmas. Although a higher proportion of thrombin was inhibited by alpha 2-macroglobulin (alpha 2M) in cord plasma and cord plasma plus prothrombin, this did not compensate for the decreased amount of thrombin inhibited by ATIII. When cord plasma was supplemented with ATIII as well as prothrombin, the proportions of thrombin complexed by the different inhibitors were similar to those of adult plasma.(ABSTRACT TRUNCATED AT 250 WORDS)

    View details for Web of Science ID A1994NX17700005

    View details for PubMedID 7517142

  • ENDOTOXIN-STIMULATED ALVEOLAR MACROPHAGES IMPAIR LUNG EPITHELIAL NA+ TRANSPORT BY AN L-ARG-DEPENDENT MECHANISM AMERICAN JOURNAL OF PHYSIOLOGY COMPEAU, C. G., Rotstein, O. D., Tohda, H., Marunaka, Y., Rafii, B., Slutsky, A. S., OBRODOVICH, H. 1994; 266 (5): C1330-C1341

    Abstract

    The Na+ transport function of alveolar epithelium represents an important mechanism for air space fluid clearance after acute lung injury. We studied the effect of endotoxin-stimulated rat alveolar macrophages on lung epithelial ion transport and permeability in vitro. Cultured rat distal lung (alveolar) epithelial monolayers incubated with both endotoxin and macrophages demonstrated a 75% decline in transepithelial resistance and a selective 60% reduction in amiloride-sensitive short-circuit current (Isc). Single-channel patch-clamp analysis demonstrated a 60% decrease in the density of 25-pS nonselective cation (NSC) channels on the apical membrane of epithelium exposed to both endotoxin and macrophages. A concurrent reduction in epithelial F-actin content suggested a role for actin depolymerization in mediating this effect. Incubation of cocultures with the methylated L-arginine (Arg) derivative NG-monomethyl-L-arginine prevented the reduction in epithelial Isc, as did substitution of L-Arg with D-Arg or incubation in L-Arg-free medium. Furthermore, the stable and products of Arg metabolism were found to have no effect on epithelial ion transport. These studies show that endotoxin-stimulated alveolar macrophages impair distal lung epithelial ion transport by an L-Arg-dependent mechanism by inactivating amiloride-sensitive 25-pS NSC channels. This may represent a novel mechanism whereby local inflammatory cells regulate lung epithelial ion transport. This could affect the ability of the lung to clear fluid from the air space.

    View details for Web of Science ID A1994NP99500022

    View details for PubMedID 7515564

  • REGIONAL EXPRESSION OF CFTR IN DEVELOPING HUMAN RESPIRATORY TISSUES AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Tizzano, E. F., OBRODOVICH, H., Chitayat, D., Benichou, J. C., Buchwald, M. 1994; 10 (4): 355-362

    Abstract

    Morbidity and mortality in cystic fibrosis (CF) patients is strongly related to their respiratory disease. We have analyzed, by means of in situ hybridization, the localization and levels of CFTR mRNA in fetal, newborn, and infant respiratory tissues. Measurable levels of CFTR transcript are present in the fetal primordial epithelium of the pseudoglandular stage lung. During the following stages of lung development, CFTR expression decreases in cells of the future alveolar spaces and is gradually limited to the epithelium of the small airways. After birth, expression decreases in the small airways and is not detected in alveolar epithelia. In trachea and large bronchi, a differential pattern of expression is also observed. No CFTR expression is found in fetal submucosal glands during fetal development, but appears gradually in the newborn period. Since CFTR codes for a secretory Cl- channel, these data probably reflect the changes that occur in the lung transition from a fluid-secreting to an absorbing organ. The pattern of expression seems paradoxical in view of the clinical-pathological manifestations of CF. Although CFTR is expressed in the normal fetus and lung development is influenced by the amount of fetal lung liquid, newborns affected with CF have normal lungs. In addition, the earliest pathologic change described in CF lungs in hyperplasia of the submucosal glands, yet expression in these structures is seen only after birth. An improved understanding of the factors that alter the expected relationship between CFTR expression and pathologic lesions in the fetal lung may provide important insights into the pathogenesis and potential treatment of lung disease in CF patients.

    View details for Web of Science ID A1994NE36700002

    View details for PubMedID 7510983

  • CL- REGULATION OF A CA2+-ACTIVATED NONSELECTIVE CATION CHANNEL IN BETA-AGONIST-TREATED FETAL DISTAL LUNG EPITHELIUM AMERICAN JOURNAL OF PHYSIOLOGY Tohda, H., Foskett, J. K., OBRODOVICH, H., Marunaka, Y. 1994; 266 (1): C104-C109

    Abstract

    Nonselective cation (NSC) channels have been identified in the apical membrane of fetal distal lung epithelium (FDLE). However, their physiological role in Na+ transport is uncertain. Because terbutaline, a beta 2-agonist, increases Na+ transport by FDLE, we studied its effect and selected signal transduction mechanisms on NSC channel activity. Using patch-clamp and single-cell imaging techniques, we found that terbutaline activated the NSC channel by 1) increasing its sensitivity to cytosolic Ca2+ concentration ([Ca2+]c) by 100- to 1,000-fold, 2) increasing [Ca2+]c from 35 nM to 3.3 microM, 3) producing a dependency of the NSC channel activity on the cytosolic Cl- concentration ([Cl-]c) at a physiological [Ca2+]c, and 4) inducing a reduction in the [Cl-]c from 45 to 25 mM, which directly activates the beta 2-treated NSC channel. These observations indicate that a beta 2-agonist physiologically activates an amiloride-blockable NSC channel in FDLE through an increase in its sensitivity to [Ca2+]c, resulting in the development of a [Cl-]c dependency at a physiological [Ca2+]c associated with both an increase in [Ca2+]c and a reduction in [Cl-]c. A development of the [Cl-]c dependency and a reduction in [Cl-]c act as a second messenger of the beta-agonist signal transduction pathway in this Na(+)-transporting epithelium.

    View details for Web of Science ID A1994NA59100012

    View details for PubMedID 7508184

  • WHOLE-CELL CL- CURRENTS IN A HUMAN PERIPHERAL AIRWAY EPITHELIAL-CELL LINE CANADIAN JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY Wang, X. D., Fedorko, L., Marunaka, Y., OBRODOVICH, H. 1993; 71 (9): 662-670

    Abstract

    We have used the whole-cell patch-clamp technique to identify and characterize Cl- currents in a cell line derived from human peripheral airway epithelium (NCI-H-441-4). The permeability sequence and relative selectivity for different anions was Br- (1.4) approximately I- (1.3) > Cl- (1.0) > F- (0.6) > gluconate (0.4) > glutamate (0.2). The current-voltage relationship displayed rectification in the outward direction. Diphenylamine-2-carboxylate (10(-4) M) applied intracellularly blocked the outward-rectified current, while extracellularly applied diphenylamine-2-carboxylate had no effect on Cl- current. This current was also blocked by extracellularly applied 5-nitro-2-(3-phenylpropylamino)benzoate (NPPB), with an estimated IC50 of 15.2 microM. Dibutyryl-cyclic AMP (10(-4) M) increased outward current, whereas pretreatment with 100 ng/mL pertussis toxin almost completely abolished the Cl- current. Pertussis toxin inhibition of this current could be partially reversed by dialysis of the cell interior with the activated alpha i-2 subunit of Gi protein. This cell line provides an opportunity to study directly the regulation of Cl- channels in cells derived from the peripheral human lung airways.

    View details for Web of Science ID A1993MK60700006

    View details for PubMedID 8313229

  • ACTIVATION OF CL- CURRENTS BY INTRACELLULAR CHLORIDE IN FIBROBLASTS STABLY EXPRESSING THE HUMAN CYSTIC-FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR CANADIAN JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY Wang, X. D., Marunaka, Y., Fedorko, L., Dho, S., Foskett, J. K., OBRODOVICH, H. 1993; 71 (9): 645-649

    Abstract

    The Cl- conductance of a mouse fibroblast cell line (LTK- cells) that was stably transfected with the human CFTR (cystic fibrosis transmembrane conductance regulator) complementary DNA was studied. Single Cl- channel activity was observed only after treatment of the cells with forskolin, the single-channel conductance being 6.2 +/- 0.2 pS with a linear current-voltage relationship. In CFTR+ cells, the whole-cell current at +90 mV increased from 7.3 +/- 2.7 pA/pF (n = 12) to 46.1 +/- 11.2 pA/pF (n = 5) after addition of dibutyryl-cyclic AMP (10(-4) M) to the bath. Increasing the intracellular Cl- concentration to 150 mM activated linear Cl- currents in the absence of cyclic AMP in CFTR+ (n = 42) but not in CFTR- cells (n = 4). Similar Cl- current was also activated by high intracellular I- concentration. These results indicate that the CFTR-induced Cl- conductance in LTK- cells can be activated by either cyclic AMP or high intracellular halide concentrations.

    View details for Web of Science ID A1993MK60700003

    View details for PubMedID 7508815

  • EXPRESSION OF THE EPITHELIAL NA+ CHANNEL IN THE DEVELOPING RAT LUNG AMERICAN JOURNAL OF PHYSIOLOGY OBRODOVICH, H., Canessa, C., Ueda, J., Rafii, B., Rossier, B. C., Edelson, J. 1993; 265 (2): C491-C496

    Abstract

    The adult mature fetal, but not immature fetal, lung is capable of actively transporting Na+ from the alveolar space. The reason for the impaired Na+ transport in the immature lung is not known; however, the apical membrane Na+ channel is the rate-limiting step for epithelial Na+ transport. This study determined whether transcripts coding for the adult rat colonic epithelial Na+ channel (alpha rENaC) were present in the fetal and adult lung and whether they were developmentally regulated. Similarly sized alpha rENaC transcripts were identified in RNA isolated from fetal and adult whole rat lung, primary cultures of fetal and adult alveolar epithelium, and adult rat whole kidneys, suggesting that the lung alpha rENaC is a similar transcript to that found in the salt-deprived rat colonic epithelium. There were low mRNA levels in 17- to 18-day gestational age (GA) fetal lungs and epithelium (term GA = 22 days), but these levels increased markedly during the saccular stage of lung development (20 days GA) and remained high in adult lungs. The combined administration of thyroid-releasing hormone and dexamethasone to pregnant rats between 16 and 18 days GA induced the expression of lung alpha rENaC in their fetuses. We conclude that alpha rENaC is expressed in mature fetal and adult alveolar epithelium and that it is influenced by hormones known to alter maturation of the fetal lung.

    View details for Web of Science ID A1993LW03100024

    View details for PubMedID 7690185

  • EFFICACY OF ALBUTEROL ADMINISTERED BY NEBULIZER VERSUS SPACER DEVICE IN CHILDREN WITH ACUTE ASTHMA JOURNAL OF PEDIATRICS Kerem, E., Levison, H., Schuh, S., OBRODOVICH, H., Reisman, J., Bentur, L., Canny, G. J. 1993; 123 (2): 313-317

    Abstract

    The aim of this study was to compare the response to inhaled albuterol after administration by nebulizer with the response after administration by a metered-dose inhaler and spacer device (MDI-spacer) to children with acute asthma. In a double-blind fashion, 33 children (6 to 14 years of age) with forced expiratory volume in 1 second (FEV1) between 20% and 70% of predicted values, and who were seen in the emergency department with acute asthma, were studied. They were treated with aerosolized albuterol or placebo by MDI-spacer, followed immediately by albuterol or placebo administered by nebulizer with oxygen. The dose ratio for albuterol by MDI-spacer versus nebulizer was 1:5. Outcome measures included a clinical score, respiratory rate, arterial oxygen saturation, and FEV1, measured before and 10, 20, and 40 minutes after treatment. With the exception of heart rate (which increased in the nebulizer group and decreased in the MDI-spacer group (p < 0.05), no difference in the rate of improvement of clinical score, respiratory rate, arterial oxygen saturation, or FEV1 was noted during the 40-minute study period between children who received albuterol by nebulizer and those who received it by MDI-spacer. We conclude that spacers and nebulizers are equally effective means of delivering beta 2-agonists to children with acute asthma.

    View details for Web of Science ID A1993LR29500025

    View details for PubMedID 8345434

  • HINDLIMB AND LUNG LYMPH FLOWS DURING PROLONGED EXERCISE JOURNAL OF APPLIED PHYSIOLOGY Coates, G., OBRODOVICH, H., Goeree, G. 1993; 75 (2): 633-638

    Abstract

    We performed experiments to determine the effect of 2h of exercise on hindlimb lymph flow (QL) and protein concentration in sheep. We compared these results with the lung QL response to long-term exercise. Eleven sheep with catheters in an efferent duct of a prefemoral lymph node and 12 sheep with chronic lung lymph catheters exercised at 2.5-3.0 km/h for up to 2h (lung lymph: range 45-120 min, mean 80 min;hindlimb lymph: range 75-120 min, mean 110.5 min). Cardiac output approximately doubled. Pulmonary vascular resistance decreased by 42%, and systemic vascular resistance decreased by 35%. There were small increases in calculated pulmonary microvascular and arterial pressures. During steady-state exercise, lung QL doubled and the lung lymph-to-plasma protein concentration ratio decreased by 16%. There was an immediate fivefold increase in hindlimb QL, and the hindlimb lymph-to-plasma protein concentration ratio decreased by 26%. Hindlimb QL decreased to a constant 130% above baseline during the last 30 min of exercise. We conclude that the marked increase in hindlimb QL early in exercise is secondary to a massaging effect in working muscles. The steady-state increases in QL toward the end of the exercise period in both lung and hindlimb are secondary to both increased surface area and pressure in the pulmonary and systemic microvascular circulations. Our data suggest that in the lung the major factor determining QL is increased vascular surface area.

    View details for Web of Science ID A1993LT58200022

    View details for PubMedID 8226462

  • NOVOBIOCIN FORMS CATION-PERMEABLE ION CHANNELS IN RAT FETAL DISTAL LUNG EPITHELIUM AMERICAN JOURNAL OF PHYSIOLOGY OBRODOVICH, H., Wang, X. D., Li, C., RAFIL, B., Correa, J., Bear, C. 1993; 264 (6): C1532-C1537

    Abstract

    The antibiotic novobiocin has been previously reported to increase Na+ transport in frog skin, presumably by attenuation of Na+ self-inhibition of Na+ channels. To determine whether novobiocin had similar effects and utilized a similar mechanism in mammalian Na(+)-transporting tissues, we studied its effect on ion transport by primary cultures of fetal distal lung epithelium (FDLE) cultured from 20-day gestationally aged rats (term = 22 days). Novobiocin (10 mM) increased short-circuit current and markedly decreased the resistance in FDLE monolayers mounted in Ussing chambers. Fura-2 single-cell studies showed that novobiocin increased intracellular Ca2+ concentration and that this resulted from extracellular sources. Nystatin-perforated patch-clamp techniques demonstrated that novobiocin increased nonrectifying cation whole cell currents without inducing detectable anion currents. Novobiocin created nonrectifying monovalent cation-selective channels in lipid bilayers. These studies demonstrated that novobiocin affects the bioelectric properties of Na+ transporting lung epithelium and that this likely occurs by the formation of ion-permeant channels in their lipid membranes.

    View details for Web of Science ID A1993LL13200020

    View details for PubMedID 7687410

  • ONTOGENY OF ALPHA-1-ISOFORMS AND BETA-1-ISOFORMS OF NA+-K+-ATPASE IN FETAL DISTAL RAT LUNG EPITHELIUM AMERICAN JOURNAL OF PHYSIOLOGY OBRODOVICH, H., Staub, O., Rossier, B. C., Geering, K., KRAEHENBUHL, J. P. 1993; 264 (5): C1137-C1143

    Abstract

    Because immature, in contrast to mature, fetal lungs have ineffective Na transport, we wished to determine the ontogeny of Na(+)-K(+)-ATPase expression in fetal distal lung epithelium (FDLE). FDLE and fibroblasts (FLF) from 17- to 22-day gestational age fetal rats (term = 22 days) were grown in primary culture. Northern and slot-blot analyses utilizing isoform-specific cDNA probes determined that alpha 1- (3.7 kb) and beta 1- (2.7, 2.3, and 1.9 kb) transcripts were present in FDLE at levels approximately fivefold higher than in FLF. alpha 2-, alpha 3-, or beta 2-isoforms of Na(+)-K(+)-ATPase were not detected. In 17-day gestational age FDLE, only small amounts of alpha 1-mRNA levels were detectable, and there were approximately 10-fold less beta 1-isoform transcripts. By 20 days gestational age, the level of alpha 1-transcripts roughly doubled, whereas beta 1-levels increased approximately sixfold. Thus, during the transition from the canalicular to saccular stages of lung development, FDLE have a differentially regulated surge in mRNA levels of alpha 1- and beta 1-Na(+)-K(+)-ATPase isoforms and do not switch isoforms during lung development. Levels for both isoform transcripts then fell before birth, reaching values less than those seen for 17-day gestational age FDLE. FDLE vesicle Na(+)-K(+)-ATPase activity did not increase until 22 days gestational age.

    View details for Web of Science ID A1993LD34600007

    View details for PubMedID 7684557

  • EXOGENOUS SURFACTANT RAPIDLY INCREASES PAO2 IN MATURE RABBITS WITH LUNGS THAT CONTAIN LARGE AMOUNTS OF SALINE AMERICAN REVIEW OF RESPIRATORY DISEASE OBRODOVICH, H., Hannam, V. 1993; 147 (5): 1087-1090

    Abstract

    Neonatal respiratory distress syndrome (RDS) is characterized by a relative surfactant deficiency and air-space edema. We tested the hypothesis that exogenous surfactant would improve gas exchange in mature surfactant-replete lungs containing large amounts of saline. Healthy young rabbits weighing 550 to 1,000 g were anesthetized and tracheotomized, and they received assisted ventilation with a FlO2 = 1. Baseline PaO2 decreased when 20 ml/kg of warmed saline (n = 6) was instilled into the lung; average PaO2 was stable and remained less than 200 mm Hg during the next 180 min. When either 50 mg/kg of natural surfactant (n = 6) or lipid-extracted surfactant (LES) (n = 6) was included in the saline solution, the PaO2 was higher (p < 0.05) than in the saline-alone group. To evaluate the effect of delayed addition of the surfactant we performed six additional experiments; the PaO2 fell to 60 +/- 6.9 SEM mm Hg after saline instillation, but within 15 min of administering LES, the PaO2 increased to 246 +/- 44.1 mm Hg, and it rose to 469 +/- 29.6 mm Hg by the end of the experiment (t = 180 min). Similar ventilator settings maintained comparable PaCO2 values in untreated and surfactant-treated groups. Gravimetric lung water contents (ml/kg body weight or ml/total lung hydroxyproline content) were markedly increased, but similar, in all groups. These studies show that the increase in PaO2 after surfactant administration to mature lungs containing large amounts of saline is similar to the increase in PaO2 seen when surfactant is given to premature infants with RDS.(ABSTRACT TRUNCATED AT 250 WORDS)

    View details for Web of Science ID A1993LJ66600003

    View details for PubMedID 8484614

  • FETAL LUNG EPITELIAL CELLS CONTAIN 2 POPULATIONS OF AMILORIDE-SENSITIVE NA+ CHANNELS AMERICAN JOURNAL OF PHYSIOLOGY Matalon, S., Bauer, M. L., Benos, D. J., Kleyman, T. R., Lin, C. M., Cragoe, E. J., OBRODOVICH, H. 1993; 264 (4): L357-L364

    Abstract

    Active Na+ transport by the alveolar epithelium plays a major role in reabsorption of the fetal lung fluid after birth. We characterized the biochemical and physiological characteristics of Na+ conductive pathways in distal fetal lung epithelial (FLE) cells isolated from 20-day-old rat fetuses. We demonstrated that a polyclonal antibody to Na+ channel protein (NaAb) binds to the plasma membranes of FLE cells. In Western blot studies, this NaAb and an anti-idiotypic monoclonal antibody to the amiloride-binding subunit of the Na+ channel protein recognized 150- and 90-kDa polypeptides in plasma membrane vesicles of FLE. 22Na+ flux measurements across plasma membrane vesicles of FLE revealed the existence of electrogenic Na+ transport, which was twice as high as the corresponding adult value. One hundred micromolars of amiloride, benzamil, and 5-(N-ethyl-N-isopropyl)-2'-4'-amiloride inhibited 30, 40, and 70% of the electrogenic Na+ transport across plasma membrane vesicles of FLE cells, respectively. The half-maximum inhibition of electrogenic Na+ transport by these substances occurred between 0.3 and 1 microM. [3H]benzamil equilibrium binding studies in membrane vesicles of FLE cells revealed the existence of two binding sites that had dissociation constant values of 19 and 1,525 nM, respectively. These data indicate the presence of both high- and low-amiloride affinity Na+ conductive pathways (channels) in FLE cells.

    View details for Web of Science ID A1993KZ60300101

    View details for PubMedID 8386466

  • 5-(N-ETHYL-N-ISOPROPYL)AMILORIDE SENSITIVE NA+ CURRENTS IN INTACT FETAL DISTAL LUNG EPITHELIAL-CELLS CANADIAN JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY Wang, X., Kleyman, T. R., Tohda, H., Marunaka, Y., OBRODOVICH, H. 1993; 71 (1): 58-62

    Abstract

    To determine whether primary cultures of rat fetal distal lung epithelium (FDLE) possessed L-type Na+ channels on their plasma membrane we performed experiments with 5-(N-ethyl-N-isopropyl)amiloride (EIPA) and other amiloride analogs. Short-circuit current (Isc) was decreased by the apical application of amiloride and benzamil, but was unaffected by 10 microM dimethylamiloride (DMA). EIPA decreased Isc when added to either the apical or basal sides. Greatest effects were seen with bilateral EIPA, where half-maximal effects occurred in the micromolar range. Measurements of intracellular pH with the fluorescent dye BCECF demonstrated that DMA impaired (IC50 = 71 nM) the ability of FDLE to recover from intracellular acidification. Nystatin perforated patch clamp techniques showed that FDLE had nonrectifying Na+ currents but no detectable Cl- currents. The whole-cell currents were reversibly decreased by 20 microM concentrations of EIPA, benzamil, and amiloride but were unaffected by 20 microM DMA. These studies indicate that there are EIPA-sensitive Na+ conductances in intact FDLE and suggest the presence of L-type Na+ conductances on their apical membrane and EIPA-sensitive K+ channels on the basolateral membrane.

    View details for Web of Science ID A1993LA79000009

    View details for PubMedID 8390327

  • EFFECT OF K-CHANNEL BLOCKERS ON BASAL AND BETA-AGONIST STIMULATED ION-TRANSPORT BY FETAL DISTAL LUNG EPITHELIUM CANADIAN JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY OBRODOVICH, H., Rafii, B. 1993; 71 (1): 54-57

    Abstract

    To determine whether basolateral K channels play an important role in the basal and beta-agonist stimulated ion transport by fetal distal lung epithelium we harvested these cells from fetal rats (20 days gestation, term = 22 days) and studied them in Ussing chambers. Short-circuit current (Isc) fell with basal 3 mM BaCl2 (3.0 +/- 0.2 (+/- SEM) to 2.0 +/- 0.2 microA.cm-2, n = 18, p < 0.01) without affecting monolayer resistance (R = 693 +/- 57 omega.cm2). Basal quinine sulfate (1 mM) also decreased Isc (3.7 +/- 0.15 to 3.0 +/- 0.10 microA.cm-2; n = 4, p < 0.01). None of apical BaCl2 (3 mM), apical quinine (1 mM), nor bilaterally applied tetraethylammonium (10 mM), lidocaine (1 mM), or 4-aminopyridine (2 mM) decreased Isc. Cell monolayers treated with basal BaCl2 (3 mM) demonstrated an impaired ability to increase their Isc in response to the beta 2-agonist terbutaline (1 mM). Basal 3 mM BaCl2 also decreased Isc in amiloride (0.1 mM) and furosemide (1 mM) treated monolayers, indicating that barium also affected the previously described amiloride-insensitive Na transport by these cells (n = 8, p < 0.01). Together these experiments suggest that normal basolateral K channel function is required for normal and beta 2-stimulated Na transport in fetal distal lung epithelium.

    View details for Web of Science ID A1993LA79000008

    View details for PubMedID 8099845

  • BICARBONATE CONCENTRATION IN RHESUS-MONKEY AND GUINEA-PIG FETAL LUNG LIQUID AMERICAN REVIEW OF RESPIRATORY DISEASE OBRODOVICH, H., Merritt, T. A. 1992; 146 (6): 1613-1614

    Abstract

    Fetal lung liquid secretion is essential for the normal growth of the lung in utero. Previous studies of fetal lung liquid secretion, mostly performed in lambs, have demonstrated that it has high Cl and very low HCO3- concentrations relative to plasma values. Because it is unknown whether primates have a similar electrolyte profile in their lung liquid, we sampled the lung liquid from rhesus monkeys (Macaca mulatta) at 127 to 128 days gestation (0.8 gestation). Although we found that lung liquid Cl concentration was higher than plasma values (p < 0.05), the HCO3- concentration was the same as in the plasma. This indicates that nonhuman primates, relative to lambs, have different cellular mechanisms for regulating fetal lung liquid HCO3- concentrations.

    View details for Web of Science ID A1992KC19500045

    View details for PubMedID 1456586

  • ANALGESIA IN CHILDREN WITH SICKLE-CELL CRISIS - COMPARISON OF INTERMITTENT OPIOIDS VS CONTINUOUS INTRAVENOUS-INFUSION OF MORPHINE AND PLACEBO-CONTROLLED STUDY OF OXYGEN INHALATION PEDIATRIC HEMATOLOGY AND ONCOLOGY ROBIEUX, I. C., Kellner, J. D., Coppes, M. J., Shaw, D., Brown, E., Good, C., OBRODOVICH, H., Manson, D., Olivieri, N. F., Zipursky, A., Koren, G. 1992; 9 (4): 317-326

    Abstract

    The objectives of the study were to compare the efficacy and safety of a continuous infusion (CIV) of morphine and intermittent parenteral opioids (IPO) in children with sickle cell vaso-occlusive crises (VOCs); to determine whether 50% oxygen administration through a face mask can reduce the duration of severe pain in patients receiving CIV morphine; and to measure morphine concentration at steady state for pharmacokinetic and pharmacodynamic analysis in patients receiving CIV morphine. The study was designed as a prospective, controlled, "before-and-after" evaluation of two different analgesic regimens. For patients receiving CIV morphine, there was a randomized, double-blind, placebo-controlled study of O2 vs. air. The patients were 66 children with sickle cell disease, 3-18 years old, requiring opioid therapy for severe VOC (32 patients in phase A, 34 in phase B). The analgesic regimens were as follows: phase A: meperidine, morphine, or codeine IM or IV bolus every 3 or 4 hours; phase B: morphine sulfate, loading dose 0.15 mg followed by CIV 0.04 mg/kg/hr. The infusion rate was adjusted every 8 hours according to pain and/or symptoms of opioid toxicity. Pain assessment was by behavioral observation (BPS). In terms of results, the mean opioid dose (morphine equivalent) was similar in both groups (0.032 +/- 0.020 mg/kg/hr in phase A and 0.035 +/- 0.011 in phase B). However, the duration of severe pain was significantly shorter in phase B (0.9 +/- 1.0 days) than in phase A (2.0 +/- 1.8 days). No severe opioid toxicity was observed in either group. Oxygen did not shorten the duration of severe pain compared to the placebo group (0.94 +/- 1.08 and 0.95 +/- 1.19 days, respectively) nor did it prevent the appearance of new pain sites. Pharmacokinetic analysis was performed in 24 patients of phase B. Total body clearance (TBC) of morphine was greater in children before puberty than after (40.4 +/- 10 vs. 28 +/- 11 mL/kg/min; p < 0.05). In conclusion, in children with severe VOCs, continuous infusion of morphine provides better analgesia than intermittent opioid therapy. Fifty percent oxygen inhalation had no effect on the duration of pain.

    View details for Web of Science ID A1992JY46600003

    View details for PubMedID 1467165

  • CYTOSOLIC CA-2+-INDUCED MODULATION OF ION SELECTIVITY AND AMILORIDE SENSITIVITY OF A CATION CHANNEL AND BETA AGONIST ACTION IN FETAL LUNG EPITHELIUM BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Marunaka, Y., Tohda, H., Hagiwara, N., OBRODOVICH, H. 1992; 187 (2): 648-656

    Abstract

    The cytosolic Ca2+ concentration ([Ca2+]i) affects many cell functions, including the modulation of ion channel activity. In this study patch clamp experiments using primary cultures of fetal distal lung epithelium (FDLE) demonstrated that the elevation of [Ca2+]i modulated a 25pS amiloride-blockable non selective cation (NSC) channel's ion selectivity and sensitivity to amiloride. An elevation of [Ca2+]i from 0.1 microM to 1mM both increased open probability (Po) and decreased the ratio of the permeability to Na to the permeability to K (PNa/PK) from 1.96 +/- 0.11 (SEM, n = 6) to 0.88 +/- 0.04 (n = 6). Within the range of [Ca2+]i from 0.1 microM to 100 microM amiloride (0.5 microM) decreased Po, however amiloride (0.5 microM) no longer affected Po of the NSC channel when [Ca2+]i was increased to 1mM under physiologic membrane potentials. A beta adrenergic agonist (terbutaline, 10 microM) increased Po in cell-attached patches from almost 0 (Po less than 0.01; n = 9) to 0.39 +/- 0.09 (n = 9) and [Ca2+]i from 40 +/- 6nM (n = 9) to more than 1 microM. This suggested that amiloride-blockable NSC channel activity and ion permeability are modulated by changes in [Ca2+]i near physiologic membrane potentials and a beta adrenergic agonist increases [Ca2+]i to more than 1 microM (unlike other epithelial including adult alveolar cells) which is associated with activation the NSC channel.

    View details for Web of Science ID A1992JN58500014

    View details for PubMedID 1382419

  • EFFECTS OF INSULIN AND TYROSINE KINASE INHIBITOR ON ION-TRANSPORT IN THE ALVEOLAR CELL OF THE FETAL LUNG BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Hagiwara, N., Tohda, H., DOI, Y., OBRODOVICH, H., Marunaka, Y. 1992; 187 (2): 802-808

    Abstract

    We studied the effect of insulin and lavendustin-A (a tyrosine kinase inhibitor) on the short-circuit current (ISC) of primary cultures of fetal distal rat lung epithelium (FDLE). Insulin (2 microM) on the basolateral side of the monolayer increased ISC from 5.76 +/- 0.83 microA/cm2 (SEM, n = 7) to 7.23 +/- 1.00 microA/cm2 (p less than 0.01) under control conditions, and from 1.00 +/- 0.31 microA/cm1 to 1.53 +/- 0.34 microA/cm2 (p less than 0.05, n = 4) when amiloride (10 microM) was present on the apical side of the monolayer. Thus insulin increased both the amiloride-sensitive and insensitive ISC with the insulin-induced increase in ISC in the absence of amiloride (1.47 +/- 0.22 microA/cm2, n = 7) being significantly larger than that in the presence of 10 microM amiloride (0.53 +/- 0.14 microA/cm2, n = 4; p less than 0.025). Insulin's effect reached steady state in 1 hr. Lavendustin-A (10 microM), a tyrosine kinase inhibitor, applied to the apical side of the monolayer attenuated but did not completely block insulin's ability to increase in ISC; i.e., insulin increased ISC in lavendustin-A treated monolayers (0.63 +/- 0.09 microA/cm2, n = 5; p less than 0.0025) but the increase was significantly smaller than that without the pretreatment of lavendustin-A (p less than 0.05). In the presence of amiloride (10 microM) and lavendustin-A (10 microM) insulin was no longer able to increase ISC (change in ISC = 0.04 +/- 0.03 microA/cm2, n = 6), suggesting that lavendustin-A had blocked the insulin's effect on the amiloride-insensitive ISC. Lavendustin-A (10 microM) had no significant effect on the basal ISC in control and amiloride treated monolayers. Our studies demonstrate that insulin increases amiloride-insensitive ISC in FDLE via lavendustin-A sensitive tyrosine kinase and that insulin's action on the amiloride-sensitive ISC of FDLE is mediated through a lavendustin-A insensitive (and presumably tyrosine kinase-independent) pathway.

    View details for Web of Science ID A1992JN58500037

    View details for PubMedID 1382422

  • OXYGEN-THERAPY IN SICKLE-CELL DISEASE AMERICAN JOURNAL OF PEDIATRIC HEMATOLOGY ONCOLOGY Zipursky, A., ROBIEUX, I. C., Brown, E. J., Shaw, D., OBRODOVICH, H., Kellner, J. D., Coppes, M. J., Koren, G., Olivieri, N. F. 1992; 14 (3): 222-228

    Abstract

    The effect of oxygen therapy on the number of irreversibly (ISC) and reversibly (RSC) sickled cells was studied in patients with sickle cell anemia. Inhalation of 50% oxygen in patients who were not in crisis produced a significant fall in RSCs and a lesser fall in ISCs. Twenty-five subjects in sickle cell crisis were chosen at random to receive either oxygen (15 patients) or air (10 patients). Those who received oxygen showed a significant reduction in RSCs, but not in ISCs. No significant change in RSCs or ISCs occurred in the group who received air. Despite the reduction in RSCs in the oxygen-treated group, there was no significant difference between the air and oxygen groups in the duration of severe pain, opioid administration, and hospitalization. It was also observed that crisis was associated with arterial desaturation and a reduction in the number of RSCs. We conclude that, although RSCs may be involved in the pathophysiology of sickle cell crisis, reduction in RSCs by oxygen therapy in these studies did not result in any reduction in the duration of crisis.

    View details for Web of Science ID A1992JD36200007

    View details for PubMedID 1510191

  • ARGININE VASOPRESSIN AND ATRIAL-NATRIURETIC-PEPTIDE DO NOT ALTER ION-TRANSPORT BY CULTURED FETAL DISTAL LUNG EPITHELIUM PEDIATRIC RESEARCH OBRODOVICH, H., Rafii, B., PERLON, P. 1992; 31 (4): 318-322

    Abstract

    Previous studies have shown that i.v. arginine vasopressin (AVP) decreases but does not stop lung fluid secretion in term fetuses not in labor. Although it has been presumed that the response to AVP results from augmented sodium transport, there is controversy whether AVP actually does affect sodium transport in mammalian lung epithelium. To determine if AVP or aldosterone could alone or together augment sodium transport in the perinatal lung, we studied primary cultures of fetal rat distal lung epithelium in Ussing chambers. The short circuit current of these sodium-transporting cells was not affected by the application of either 30 or 300 mU/mL AVP whether or not they were previously exposed to aldosterone (10(-6) M). Aldosterone also did not affect the baseline bioelectric properties. Short circuit current increased in response to 8-bromo cAMP (10(-4) M) and 3-isobutyl-1-methylxanthine (10(-3) M) to levels 169 +/- 16 (SEM) and 172 +/- 7% of respective baseline values. AVP had no effect in cells pretreated with 3-isobutyl-1-methylxanthine. Monolayers also did not respond to atrial natriuretic peptide (10(-11) to 10(-8) M). Monolayers of Na-absorbing A6 renal epithelium did increase short circuit current with either aldosterone or AVP. AVP increased endogenous cAMP levels in A6 but not fetal rat distal lung epithelium cells, suggesting that fetal rat distal lung epithelium lacks V2 receptors. These studies demonstrate that AVP does not increase ion transport in cultured fetal distal lung epithelium although these cells possess the necessary second messenger system.

    View details for Web of Science ID A1992HL04500003

    View details for PubMedID 1315019

  • FATAL PULMONARY ARTERIAL OCCLUSIVE VASCULAR-DISEASE FOLLOWING CHEMOTHERAPY IN A 9-MONTH-OLD INFANT HUMAN PATHOLOGY Bentur, L., Cullinane, C., Wilson, P., Greenberg, M., OBRODOVICH, H., Silver, M. M. 1991; 22 (12): 1295-1298

    Abstract

    Fatal pulmonary hypertension developed in an infant during the 7-month period in which he received, via a central venous catheter, combination chemotherapy for stage IV neuroblastoma as well as intermittent parenteral feeding. In a lung biopsy and at autopsy, small pulmonary arteries showed diffuse medial hypertrophy and peripheral muscularization, very extensive concentric intimal fibrosis, and focal eccentric fibrosis evolving from organizing thrombi. Pulmonary veins were normal. Hypothetically, chemotherapeutic drug therapy (possibly potentiated either by the parenteral nutrition or simply by the vehicular fluids causing volume loading of the pulmonary circulation) could cause occlusive pulmonary arterial disease by several mechanisms, but the association has not been described previously, although use of such drugs has been reported with pulmonary veno-occlusive disease.

    View details for Web of Science ID A1991GX83700018

    View details for PubMedID 1748437

  • SODIUM-CHANNEL BUT NEITHER NA+-H+ NOR NA-GLUCOSE SYMPORT INHIBITORS SLOW NEONATAL LUNG WATER CLEARANCE AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY OBRODOVICH, H., Hannam, V., Rafii, B. 1991; 5 (4): 377-384

    Abstract

    Normal clearance of alveolar liquid following birth requires active Na transport; however, the contribution of Na channels, Na-H antiports, and Na-glucose symports is unknown. We demonstrated that intraalveolar instillation of amiloride (n = 6) or the more specific Na channel blockers benzamil (n = 13) or phenamil (n = 12) before the first breath impaired lung water clearance relative to control newborns (n = 34). Benzamil and phenamil were more potent than amiloride (P less than 0.05). Neither the Na-H antiport inhibitor dimethyl amiloride (n = 7) nor the Na-glucose symport inhibitor phloridzin (n = 7) impaired lung water clearance. Ion substitution experiments with fetal rat type II alveolar epithelia demonstrated that more than 95% of their resting or terbutaline-stimulated short circuit current (Isc) depended upon Na bathing their apical membrane. Isc was decreased by amiloride (IC50 of amiloride-sensitive Isc = 0.3 x 10(-6) M) and benzamil (IC50 of benzamil-sensitive Isc = 0.3 x 10(-7) M) but was unaffected by dimethyl amiloride (10(-4) M). We conclude that in vivo postnatal clearance of fetal lung liquid can be impaired by Na channel blockers and is unaffected by blockers of Na-H antiports and Na-glucose symports. Na transport in fetal type II cells has high affinity for amiloride, and these cells likely contribute to normal neonatal lung liquid clearance.

    View details for Web of Science ID A1991GJ51400011

    View details for PubMedID 1654956

  • INFLUENCE OF FETAL PULMONARY EPITHELIUM ON THROMBIN ACTIVITY AMERICAN JOURNAL OF PHYSIOLOGY OBRODOVICH, H., Berry, L., DCOSTA, M., Burrows, R., Andrew, M. 1991; 261 (4): L262-L270

    Abstract

    Epithelial injury and intra-alveolar fibrin are present in lung injury. To determine whether healthy fetal and neonatal lung epithelium could regulate thrombin activity (hence fibrin formation) we collected amniotic and postnatal endotracheal tube fluids from humans and directly sampled lung and amniotic fluids from fetal guinea piglets, rabbit pups, and lambs. The coagulant properties of the cell surface and media conditioned by rat fetal type II alveolar epithelium were assessed. All fluids contained glycosaminoglycans (GAGs), but mass and biological assays demonstrated only some (heparan sulfate and to a lesser extent dermatan sulfate) had antithrombin activity. The presence of proteoglycans (greater than 1,000 kDa) yielding active GAGs with less than 100 kDa after base elimination were demonstrated by Sepharose CL4B chromatography. Epithelial-derived fluids contained a factor VII-dependent procoagulant activity, but concentrated conditioned media overlying primary cultures of type II epithelium demonstrated a net antithrombin effect. These studies demonstrate that the lungs of human and nonprimate mammalian fetuses and fetal type II epithelium secrete GAGs, some of which possess antithrombin activity, which would oppose intra-alveolar fibrin formation.

    View details for Web of Science ID A1991GK86700064

    View details for PubMedID 1928360

  • EPITHELIAL ION-TRANSPORT IN THE FETAL AND PERINATAL LUNG AMERICAN JOURNAL OF PHYSIOLOGY OBRODOVICH, H. 1991; 261 (4): C555-C564

    Abstract

    The lung is a complex organ whose intrauterine development depends on many factors, one of which is a continuous secretion of large volumes of Cl(-)-enriched fluid by the pulmonary epithelium. At birth this fluid must be cleared, and it is now known that this process depends in large part on active Na+ transport by the pulmonary epithelium. Only recently has it been possible to culture some of the different lung epithelial cells so that it is possible to investigate the role of individual epithelial cell types, their individual cellular transport mechanisms, and how these are affected by developmental lung maturity.

    View details for Web of Science ID A1991GK86700002

    View details for PubMedID 1928320

  • CATION SELECTIVE CHANNEL IN FETAL ALVEOLAR TYPE-II EPITHELIUM BIOCHIMICA ET BIOPHYSICA ACTA Orser, B. A., BERTLIK, M., Fedorko, L., OBRODOVICH, H. 1991; 1094 (1): 19-26

    Abstract

    A cation selective channel was identified in the apical membrane of fetal rat (Wistar) alveolar type II epithelium using the patch clamp technique. The single channel conductance was 23 +/- 1.2 pS (n = 16) with symmetrical NaCl (140 mM) solution in the bath and pipette. The channel was highly permeable to Na+ and K+ (PNa/PK = 0.9) but essentially impermeant to chloride and gluconate. Membrane potential did not influence open state probability when measured in a high Ca2+ (1.5 mM) bath. The channel reversibly inactivated when the bath was exchanged with a Ca(2+)-free (less than 10(-9) M) solution. The Na+ channel blocker amiloride (10(-6) M) applied to the extracellular side of the membrane reduced P(open) relative to control patches; P(control) = 0.57 +/- 0.11 (n = 5), P(amiloride) = 0.09 +/- 0.07 (n = 4, p less than 0.01), however, amiloride did not significantly influence channel conductance (g); g(control) 19 +/- 0.9 pS (n = 5), 18 +/- 3.0 pS (n = 4). More than one current level was observed in 42% (16/38) of active patches; multiple current levels (ranging from 2 to 6) were of equal amplitude suggesting the presence of multiple channels or subconductance states. Channel activity was also evident in cell attached patches. Since monolayers of these cells absorb Na+ via an amiloride sensitive transport mechanism we speculate that this amiloride sensitive cation selective channel is a potential apical pathway for electrogenic Na+ transport in the alveolar region of the lung.

    View details for Web of Science ID A1991GD60500003

    View details for PubMedID 1653022

  • CARDIOPULMONARY FUNCTION DURING LONG-TERM CENTRAL VENOUS CATHETERIZATION AMERICAN JOURNAL OF PEDIATRIC HEMATOLOGY ONCOLOGY OBRODOVICH, H., Adams, M., Coates, G., Way, R. C., Andrew, M. 1991; 13 (2): 148-151

    Abstract

    A prospective study was performed in an ambulatory group of infants, children, and young adults with neoplastic disorders to determine the prevalence of significant cardiopulmonary complications during long-term central venous catheterization. A cohort of 20 consecutive patients who had central venous catheters in situ for a mean of 13.5 months underwent pulmonary function testing, chest radiography, ventilation perfusion lung scintigraphy, electrocardiography, and echocardiography. No significant complications were seen. Specifically, there was no evidence of localized thrombus formation or pulmonary emboli, and no indirect evidence of pulmonary hypertension. In addition, we reviewed retrospectively the autopsy reports of 15 patients who died with central venous catheters in place. Three of these patients had superficial endocardial reactions. One patient had a right atrial mural thrombus related to the catheter, but two other patients were noted to have left atrial mural thrombi. This study demonstrates that central venous catheters can be used safely on a long-term basis in ambulatory patients requiring frequent venous access for treatment of their neoplastic disorders.

    View details for Web of Science ID A1991FL39600007

    View details for PubMedID 1906247

  • A549 LUNG EPITHELIAL-CELLS SYNTHESIZE ANTICOAGULANT MOLECULES ON THE CELL-SURFACE AND MATRIX AND IN CONDITIONED MEDIA AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Berry, L., Andrew, M., Post, M., Ofosu, F., OBRODOVICH, H. 1991; 4 (4): 338-346

    Abstract

    To investigate mechanisms regulating intra-alveolar coagulation, we studied monolayers of the A549 human lung epithelial cell line. The surface of A549 cells delayed the onset of prothrombin-to-thrombin conversion and prevented total prothrombin consumption in normal plasma compared to plastic cell-free wells. Similar results were achieved with bovine pulmonary endothelial (CPAE) and rat intestinal epithelial (IEC-6) cell lines, whereas Madin-Darby canine kidney renal epithelial cell line accelerated thrombin formation. The A549 surface catalyzed antithrombin III-thrombin complex formation with no significant increase in thrombin inactivation from heparin cofactor II. The A549 cell surface effects were largely, but not completely, reversed to values obtained for plastic when protein C-deficient plasma was used. Pretreatment of the cell surface with chondroitinase ABC plus heparitinase prior to thrombin generation experiments had no effect on the total prothrombin consumed but decreased the initial delay. Heparan sulfate as well as dermatan sulfate and other chondroitin sulfates were detected on the A549 surface using alcian blue staining. Conditioned media from A549, CPAE, and IEC-6 cells delayed the clot time of recalcified plasma. Use of chondroitinase ABC and heparitinase were both required to obliterate the A549 conditioned media activity. After growing A549 cells in 35SO(2-)4-containing medium, the resultant conditioned medium was found to contain 2,000 kD and 300- to 1,000-kD proteoglycans that yielded chains of less than or equal to 100 kD on reductive elimination with base.(ABSTRACT TRUNCATED AT 250 WORDS)

    View details for Web of Science ID A1991FE90300007

    View details for PubMedID 2015100

  • BRONCHOPULMONARY DYSPLASIA - IMPROVEMENT IN LUNG-FUNCTION BETWEEN 7 AND 10 YEARS OF AGE JOURNAL OF PEDIATRICS Blayney, M., Kerem, E., Whyte, H., OBRODOVICH, H. 1991; 118 (2): 201-206

    Abstract

    To evaluate the natural history of bronchopulmonary dysplasia, we studied the same 32 patients at a mean age of 7 and 10 years. The group as a whole had normal height and weight percentiles, and each child grew along his or her established somatic growth curve. Although some children had abnormal values, the group maintained a normal mean total lung capacity and functional residual capacity. The mean residual volume and the residual volume/total lung capacity ratios were elevated at both ages. At age 7 years the 19 patients (59%) who had a forced expiratory volume in 1 second (FEV1) of less than 80% had "catch up" improvement by 10 years of age (65 +/- 11% to 72 +/- 16% of predicted value; p less than 0.05). All the children who had a normal FEV1 at 7 years of age continued to have a normal FEV1 at age 10 years. Resting single-breath carbon monoxide uptake by the lung was normal when measured at age 10 years. The majority of patients had a positive methacholine challenge test result at both ages, although there was a low incidence of clinically diagnosed asthma. This study demonstrates that patients with bronchopulmonary dysplasia who have normal lung function at age 7 have had normal lung growth and that those with evidence of mild to moderate lung disease have continued lung growth or repair, or both, during their school years.

    View details for Web of Science ID A1991EX30200006

    View details for PubMedID 1993945

  • INFLUENCE OF GESTATIONAL-AGE ON POTENTIAL DIFFERENCE ACROSS FETAL TYPE-II ALVEOLAR EPITHELIUM IN ALVEOLAR-LIKE STRUCTURES BIOLOGY OF THE NEONATE Orser, B., Fedorko, L., Rafii, B., Post, M., OBRODOVICH, H. 1991; 59 (2): 61-68

    Abstract

    The active transport of ions across fetal pulmonary epithelium results in lung fluid secretion. This study investigated the potential difference (PD) across fetal type II alveolar epithelium, one of the several epithelial cell types in the fetal lung. Aggregates of these cells in alveolar-like structures (ALS) underwent microelectrode impalement to determine the effect of gestational age, and drugs on transcellular PD. In 6 ALS, the intraluminal position of the electrode was confirmed by fluorescent imaging after iontophoretic injection of Lucifer Yellow VS dye. The average PD recorded from the lumens of ALS harvested from 20-day fetal rats was 11.8 +/- (SE) 0.48 mV (lumen-negative; n = 164). Exposure to ouabain 10(-3) M significantly reduced PD from control values of 12.3 +/- (SE) 0.87 to 8.1 +/- (SE) 0.95 mV (p less than 0.01, n = 107) in ALS obtained from 20-day fetal rats. Terbutaline 10(-3) M and furosemide 10(-3) M did not influence PD. Cells obtained from fetuses near term showed a significant reduction in PD. ALS from 18-day (n = 53), 20-day (n = 164) and 22-day (n = 56) fetuses measured 13.5 +/- 0.62, 11.8 +/- 0.48 and 9.3 +/- 0.49 mV, respectively (p less than 0.05 day 18 vs. day 22). These results demonstrate that fetal type II cells grown in organotypic culture maintain a larger transcellular electrical gradient than previously reported. PD decreases with increasing gestational age and can be reduced by the Na-K ATPase inhibitor ouabain.

    View details for Web of Science ID A1991FD52100001

    View details for PubMedID 2036469

  • WHEN THE ALVEOLUS IS FLOODING, ITS TIME TO MAN THE PUMPS AMERICAN REVIEW OF RESPIRATORY DISEASE OBRODOVICH, H. 1990; 142 (6): 1247-1248

    View details for Web of Science ID A1990EM38500002

    View details for PubMedID 2252239

  • EFFECT OF PENTOXIFYLLINE ON HEMODYNAMICS, ALVEOLAR FLUID REABSORPTION, AND PULMONARY-EDEMA IN A MODEL OF ACUTE LUNG INJURY AMERICAN REVIEW OF RESPIRATORY DISEASE SEEAR, M. D., HANNAM, V. L., Kaapa, P., Raj, J. U., OBRODOVICH, H. M. 1990; 142 (5): 1083-1087

    Abstract

    We investigated the effect of pentoxifylline (PTX) on the development of pulmonary edema in a model of adult respiratory distress syndrome in rabbits. Lung injury was induced by repeated saline lavages in adult rabbits weighing 2.5 to 3.5 kg. Rabbits pretreated with PTX (20 mg/kg bolus followed by 20 mg/kg/h infusion) developed significantly lower amounts of lung edema 4 h after saline lavage (extravascular lung water to dry weight ratio [W/D], 6.9 +/- 0.6 SD versus 8.9 +/- 0.5 in control animals). PTX produced a 25% increase in cardiac output, but there were no differences between treated and untreated groups in calculated pulmonary vascular resistance or microvascular pressure. To determine whether PTX could have lowered pulmonary venous resistance and thus lowered effective microvascular pressure for fluid filtration, we directly measured pulmonary artery and left atrial pressures, and measured by micropuncture the pressure in 20 to 40 microns subpleural venules in four open-chested rabbits 3 to 4 h after lavage. Venous resistance was low (venous pressure drop 0.9 +/- 0.1 mm Hg) and was unchanged by PTX infusion. To determine if PTX decreased lung water by accelerating active alveolar fluid reabsorption, a single 60-ml aliquot of saline was instilled into the lungs of normal rabbits treated with saline or PTX. Both groups had a similar decrease in lung water content 1 and 4 h later. Our data indicate that PTX reduces edema formation in rabbits after saline lavage, not by lowering microvascular pressures for fluid filtration or by acceleration alveolar fluid reabsorption, but possibly by its anti-inflammatory effect on neutrophil function.

    View details for Web of Science ID A1990EG08800019

    View details for PubMedID 2240831

  • EFFECT OF FIBRINOGEN DEGRADATION PRODUCTS AND LUNG GROUND SUBSTANCE ON SURFACTANT FUNCTION BIOLOGY OF THE NEONATE OBRODOVICH, H. M., Weitz, J. I., Possmayer, F. 1990; 57 (6): 325-333

    Abstract

    Acute lung injury syndromes have many characteristics including protein-rich alveolar edema, hyaline membranes, and abnormal surface tension at the alveolar air-liquid interface. Increased surface tension can occur because of a relative surfactant deficiency and/or dysfunction. It has been previously demonstrated that surfactant dysfunction occurs when plasma protein inhibitors leak into the alveolar space during the induction of the lung injury and edema formation. The present study investigated whether inhibitors that would be generated during the stage of repair from lung injury could impair surfactant function. We determined whether fibrinogen degradation products (FDP) which would be released during lysis of the fibrin(ogen)-containing alveolar exudate and hyaline membranes, and components of the lungs' ground substance could inhibit the in vitro function of a lipid extract surfactant preparation. FDP were prepared by incubating human fibrinogen with plasmin or neutrophil elastase for 4 min to 60 h and were characterized by SDS-PAGE. Early (fragment X and Y) and late (fragment D and E) plasmin-derived FDP (MW greater than 40,000) inhibited surfactant function as assessed by a bubble surfactometer. The early elastase-derived FDP also inhibited surfactant, but the later and much smaller fragments (MW less than 15,000) did not affect surfactant function. Laminin also inhibited surfactant in a dose-dependent manner. Neither hyaluronic acid nor heparan sulfate affected surfactant performance in vitro. We conclude that plasmin-induced lysis of intraalveolar fibrinogen and hyaline membranes will result in prolonged generation (i.e. days) of surfactant inhibitors.(ABSTRACT TRUNCATED AT 250 WORDS)

    View details for Web of Science ID A1990DJ78900001

    View details for PubMedID 2142607

  • Cation channels in apical membrane of fetal alveolar epithelium. Canadian journal of anaesthesia = Journal canadien d'anesthésie Orser, B., Bertlik, H., Fedorko, L., O'Brodovich, H. 1990; 37 (4): S3-?

    View details for PubMedID 2163293

  • BIOELECTRIC PROPERTIES OF FETAL ALVEOLAR EPITHELIAL MONOLAYERS AMERICAN JOURNAL OF PHYSIOLOGY OBRODOVICH, H., Rafii, B., Post, M. 1990; 258 (4): L201-L206

    Abstract

    To investigate the bioelectric properties on one of the cell types that line the distal lung unit, we isolated type II alveolar epithelium from 18- to 20-day gestation fetal rats (term = 22 days) and grew them on collagen-coated nitrocellulose filters. Amiloride impaired ion transport in a dose-dependent fashion (10(-4) to 10(-6) M) with 10(-4) M decreasing potential difference (PD) (mean +/- SE, 2.0 +/- 0.49 to 0.9 +/- 0.26 mV, P less than 0.01, lumen negative) and short-circuit current (Isc) (7.0 +/- 1.0 to 2.4 +/- 0.64 uA/cm2, P less than 0.01) without affecting resistance (R) (241 +/- 33 to 216 +/- 41 omega. cm2). Benzamil (10(-5) M) but not dimethylamiloride (10(-5) M) decreased Isc. Terbutaline (10(-3) M) increased PD from 1.2 +/- 0.13 to 3.3 +/- 0.40 mV (P less than 0.01), and application of amiloride (10(-4) M) after terbutaline reduced PD and Isc to less than initial base-line values. The Na(+)-K(+)-2Cl- cotransport inhibitors bumetanide (10(-4) M) and furosemide (10(-3) M) had no effect on PD and Isc either before or after terbutaline. Neither the Cl- channel blocker diphenylamine-2-carboxylate (10(-3) M) nor the Na(+)-glucose cotransport inhibitor phloridzin (10(-3) M) affected the bioelectric properties. Fetal type II alveolar epithelium in primary culture actively transport ions and, on the basis of inhibitor-agonist experiments, probably do not secrete Cl- but absorb Na+ through Na+ channels.

    View details for Web of Science ID A1990DB09200079

    View details for PubMedID 2159225

  • AMILORIDE IMPAIRS LUNG WATER CLEARANCE IN NEWBORN GUINEA-PIGS JOURNAL OF APPLIED PHYSIOLOGY OBRODOVICH, H., Hannam, V., Seear, M., Mullen, J. B. 1990; 68 (4): 1758-1762

    Abstract

    To determine whether epithelial ion transport is physiologically important for lung water clearance after birth, the sodium transport inhibitor amiloride or its vehicle saline was given intratracheally to newborn full-term guinea pigs before the first breath. Guinea pigs given saline intratracheally breathed normally and had arterial O2 saturations (SaO2) greater than 94%. In contrast, guinea pigs that had an estimated 10(-4) M intra-alveolar concentration of amiloride had chest wall retractions and 88 +/- 3.6% (SD) SaO2 (P less than 0.01). Extravascular lung water (EVLW) per gram of dry lung weight 4 h after birth was significantly greater in newborns that received amiloride (8.3 +/- 1.1, n = 5) than in those that received saline (5.6 +/- 0.9, n = 7, P less than 0.01). The degree of perivascular fluid cuffing at 25 cmH2O inflation was quantitatively similar in amiloride- and saline-treated animals. The effect of amiloride was dose dependent. Intratracheal amiloride did not affect EVLW in 9-day-old guinea pigs. This study demonstrates that intratracheal amiloride before the first breath results in respiratory distress, hypoxemia, and an abnormally high EVLW. Epithelial sodium transport contributes normal lung liquid clearance after birth.

    View details for Web of Science ID A1990DA92600063

    View details for PubMedID 2161411

  • SEQUENTIAL PULMONARY-FUNCTION MEASUREMENTS DURING TREATMENT OF INFANTILE CHRONIC INTERSTITIAL PNEUMONITIS JOURNAL OF PEDIATRICS Kerem, E., Bentur, L., England, S., Reisman, J., OBRODOVICH, H., Bryan, A. C., Levison, H. 1990; 116 (1): 61-67

    Abstract

    Three infants with histologically confirmed chronic interstitial pneumonitis were treated with monthly intravenously administered high doses of methylprednisolone with or without daily hydroxychloroquine therapy. We applied the multiple occlusion technique to measure the static respiratory system compliance, and the end-inspiratory occlusion technique to measure passive respiratory system compliance, resistance, and time constant. When assessed by clinical criteria and pulmonary function measurements, all three patients showed improvement with this treatment. Clinical improvement was associated with an increase in respiratory system compliance as measured by both techniques (60% to 100% increase in all patients). The passive respiratory resistance and the time constant did not closely reflect the clinical course. We conclude (1) that high doses (pulses) of methylprednisolone and daily oral doses of hydroxychloroquine are effective in the treatment of infantile chronic interstitial pneumonitis and (2) that the respiratory system compliance, measured by both pulmonary function techniques, correlates well with the response to treatment and change in clinical status.

    View details for Web of Science ID A1990CH03400011

    View details for PubMedID 2295964

  • SIMULTANEOUS MEASUREMENT OF LUNG CLEARANCE RATES FOR TC-DTPA AND IN-DTPA IN NORMAL AND DAMAGED LUNGS JOURNAL OF APPLIED PHYSIOLOGY OBRODOVICH, H., Coates, G., Kay, J., MUYSSON, D. 1989; 66 (5): 2293-2297

    Abstract

    We investigated the relative clearance rates for 99mTc-labeled diethylenetriamine-pentaacetate (Tc-DTPA) and 113mIn-labeled DTPA (In-DTPA) when they were inhaled and deposited together within the lungs of same animal. Submicronic aerosols containing Tc-DTPA and In-DTPA were simultaneously generated by different nebulizers and collected within the same anesthetic bag. The combined aerosols were insufflated into piglets. Clearances for both compounds were measured simultaneously in normal lungs and when the lungs were damaged by intravenous oleic acid or by a presumed oxidant agent, intravenous or intratracheal phorbol myristate acetate (PMA). A medium-energy collimator and a computer-assisted gamma camera were used to calculate clearances. Correction was made for downscatter from the In photopeak into the Tc window. Marked lung injury occurred as evidenced by increases in lung water content and decreases in arterial PO2. The clearance of In-DTPA was slightly but significantly slower than for Tc-DTPA in each group of animals. The correlation (r = 0.93) between clearances for Tc-DTPA and In-DTPA was good, even though in vitro studies demonstrated that Tc-DTPA, but not In-DTPA, slowly dissociated at room and body temperatures. Oleic acid increased, but surprisingly, PMA had no effect on clearance rates for both In-DTPA and Tc-DTPA. We recommend continued use of Tc-DTPA for these measurements in view of its lower cost, requirement for only low-energy collimation, better imaging characteristics, and widespread availability. The overlap between control and injured lungs and the lack of increased clearance rates after PMA suggest this technique does not always detect acute lung injury.

    View details for Web of Science ID A1989U606100038

    View details for PubMedID 2663817

  • THE RESPONSE OF THE PULMONARY CIRCULATION TO EXERCISE DURING NORMOXIA AND HYPOXIA FOLLOWING PNEUMONECTOMY IN THE ADULT SHEEP CANADIAN JOURNAL OF PHYSIOLOGY AND PHARMACOLOGY Smith, M., Coates, G., Kay, J. M., OBRODOVICH, H. 1989; 67 (3): 202-206

    Abstract

    Pneumonectomy approximately halves the available pulmonary vascular bed. It is unknown whether the remaining lung has sufficient vascular reserve to cope with increased blood flow under stressful conditions without demonstrating abnormal pulmonary hemodynamics. To investigate this question, unanesthetized ewes with vascular catheters had hemodynamics assessed before and after a left pneumonectomy. Subsequently, on different days, the sheep were exercised on a treadmill under normoxic and hypobaric hypoxic (430 mmHg) (1 mmHg = 133.3 Pa) conditions. Pneumonectomy itself increased mean pulmonary arterial pressure by 4 mmHg. During normoxic or hypoxic exercise, the pneumonectomized sheep demonstrated a pulmonary hemodynamic response similar to normal sheep with two lungs. The pressure-flow relation for the right lung suggested the vascular reserve of the lung was not exceeded during exercise in the pneumonectomized sheep. Eighteen to 70 days after pneumonectomy there was no evidence of right ventricular hypertrophy, but there were small increases in the number of muscularized vessels less than 50 microns diameter and in the amount of muscle in normally muscularized pulmonary arteries. This study demonstrates that pneumonectomy slightly increases mean pulmonary arterial pressure. However, there is sufficient vascular reserve in the remaining lung to permit a normal hemodynamic response to exercise-induced increased blood flow even under hypoxic conditions.

    View details for Web of Science ID A1989T784500005

    View details for PubMedID 2743207

  • FETAL LAMB COAGULATION SYSTEM DURING BIRTH ASPHYXIA AMERICAN JOURNAL OF HEMATOLOGY Andrew, M., OBRODOVICH, H., Mitchell, L. 1988; 28 (3): 201-203

    Abstract

    Levels of many coagulation factors are low in the healthy infant and even lower in the asphyxiated premature infant. We investigated whether a brief exposure to asphyxia at the time of birth causes the activation and consumption of coagulation factors. Following delivery by caesarean section, premature lambs were asphyxiated by occluding the endotracheal tube for 10 min and then resuscitated. Blood was obtained prior to and following birth for measurement of blood gases and the coagulation system. Birth asphyxia in the premature lamb resulted in thrombin generation and rapid consumption of specific coagulation factors.

    View details for Web of Science ID A1988P700500016

    View details for PubMedID 3407640

  • Lung clearance of 99mTc-DTPA in patients with acute lung injury and pulmonary edema. Journal of thoracic imaging Coates, G., O'Brodovich, H., Dolovich, M. 1988; 3 (3): 21-27

    Abstract

    Several acute and chronic conditions that alter the integrity of the pulmonary epithelium increased the rate of absorption or clearance into the circulation of small solutes deposited in the alveoli. Technetium 99m diethylenetriamine pentaacetic acid can be deposited in the lungs as a submicronic aerosol and its rate of clearance measured with a gamma camera or simple probe. This clearance technique is currently being used to evaluate patients who have developed pulmonary edema and also to detect those patients from a high risk group who are likely to develop adult respiratory distress syndrome (ARDS). Its role in the evaluation of patients with pulmonary edema is still under active investigation. It is clear that a single measurement in patients who smoke is not useful, but repeated measurements may provide important information. The lung clearance measurement is very sensitive to changes in epithelial integrity but is not specific for ARDS. It may be most useful in combination with other predictive tests or when the clearance rate is normal.

    View details for PubMedID 3292780

  • FETAL LAMB COAGULATION SYSTEM DURING NORMAL BIRTH AMERICAN JOURNAL OF HEMATOLOGY Andrew, M., OBRODOVICH, H., Mitchell, L. 1988; 28 (2): 116-118

    Abstract

    The contribution of the birth process to the unique coagulation system in the young was investigated using the fetal lamb model. Specific components of the coagulation system were measured prior to and immediately following delivery of the lamb. The results show that the birth process itself does not contribute to the physiologically low levels of coagulation factors present in the newborn.

    View details for Web of Science ID A1988P266500010

    View details for PubMedID 3394713

  • ANGIOTENSIN-II PREVENTS HYPOXIC PULMONARY-HYPERTENSION AND VASCULAR CHANGES IN RAT AMERICAN JOURNAL OF PHYSIOLOGY RABINOVITCH, M., Mullen, M., Rosenberg, H. C., Maruyama, K., OBRODOVICH, H., Olley, P. M. 1988; 254 (3): H500-H508

    Abstract

    Angiotensin II, a vasoconstrictor, has been previously demonstrated to produce a secondary vasodilatation due to release of prostaglandins. Because of this effect, we investigated whether infusion of exogenous angiotensin II via miniosmopumps in rats during a 1-wk exposure to chronic hypobaric hypoxia might prevent pulmonary hypertension, right ventricular hypertrophy, and vascular changes. We instrumented the rats with indwelling cardiovascular catheters and compared the hemo-dynamic and structural response in animals given angiotensin II, indomethacin in addition to angiotensin II (to block prostaglandin production), or saline with or without indomethacin. We then determined whether angiotensin II infusion also prevents acute hypoxic pulmonary vasoconstriction. We observed that exogenous angiotensin II infusion abolished the rise in pulmonary artery pressure, the right ventricular hypertrophy, and the vascular changes induced during chronic hypoxia in control saline-infused rats with or without indomethacin. The protective effect of angiotensin II was lost when indomethacin was given to block prostaglandin synthesis. During acute hypoxia, both angiotensin II and prostacyclin infusions similarly prevented the rise in pulmonary artery pressure observed in saline-infused rats and in rats given indomethacin or saralasin in addition to angiotensin II. Thus exogenous angiotensin II infusion prevents chronic hypoxic pulmonary hypertension, associated right ventricular hypertrophy, and vascular changes and blocks acute hypoxic pulmonary hypertension, and this is likely related to its ability to release vasodilator prostaglandins.

    View details for Web of Science ID A1988M591900014

    View details for PubMedID 3348429

  • THE EFFECT OF HYPOXIA ON PLATELET SURVIVAL AND SITE OF SEQUESTRATION IN THE NEWBORN RABBIT THROMBOSIS AND HAEMOSTASIS Castle, V., Coates, G., Mitchell, L. G., OBRODOVICH, H., Andrew, M. 1988; 59 (1): 45-48

    Abstract

    Thrombocytopenia occurs frequently in sick neonates that have experienced perinatal asphyxia. This study investigated the effect of one component of asphyxia, hypoxia, on platelet lifespan and site of sequestration. 111Indium oxine platelet survivals with scintigraphic imaging were performed in newborn and adult rabbits exposed to room air (normoxia) or following exposure to a 15 minute, severe hypoxic insult (FIO2 = 0.05). Platelet survivals in normoxic adults (n = 27) and newborn rabbits (n = 11) were similar (60 +/- 3.9 hr vs 64 +/- 8.0 hr, m +/- SEM). Inhalation of 5% oxygen for 15 minutes was not associated with an acidemia and did not produce thrombocytopenia but significantly shortened the platelet survival to 34 +/- 3 hr in the adult (n = 18) and 38 +/- 3 hr in the newborn rabbit (n = 7). Postmortem measurement of the sites of 111In-platelet accumulation showed that under normoxic conditions the platelets accumulated in the liver and spleen (23 +/- 4.3% and 8 +/- 1.0% of the total body counts) in the adult with even greater accumulation in the liver (58 +/- 6.8%) and spleen (19 +/- 4.9%) of the newborn (p less than 0.001). The latter observation was likely due to the relatively increased size of the liver and spleen in the newborn compared to the adult. Hypoxia did not alter the site of platelet sequestration in adults or newborns. Our results suggest that the newborn has the same platelet survival as the adult and that acute, severe hypoxia significantly shortens the survival of platelets in both groups. Although the sites of sequestration are qualitatively the same in the newborn, there is greater sequestration in the liver and spleen when compared to the adult.

    View details for Web of Science ID A1988M016200009

    View details for PubMedID 3363533

  • AIRWAY EPITHELIAL DAMAGE IN PREMATURE-INFANTS WITH RESPIRATORY-FAILURE AMERICAN REVIEW OF RESPIRATORY DISEASE Lee, R. M., OBRODOVICH, H. 1988; 137 (2): 450-457

    Abstract

    To evaluate the evolution of airway epithelial damage in premature infants developing bronchopulmonary dysplasia (BPD), a detailed quantitative light and electron microscopic evaluation was performed of the major airways in 3 infants who succumbed from respiratory failure. The 3 infants ranged from 24 to 28 wk gestational age and died after 14 h, 10 days, and 6 months of life. The tracheal epithelium was severely damaged with up to half of the epithelium being denuded in each infant. The infants who died after 14 h and after 10 days demonstrated minimal epithelial denudation in more distal bronchial divisions. However, gross and microscopic abnormalities of the ciliated component of the epithelium were common. In the infant with severe established BPD, there was extensive epithelial denudation and ciliary abnormalities out to the fourth bronchial division. The percentage of goblet cells in the total cell population decreased from proximal to distal major airways, with the lowest percentage being seen in the infant with established BPD. These studies demonstrate the respiratory failure and assisted ventilation in the human neonate is associated with severe epithelial damage in the trachea and proximal bronchi. It suggests that the difficulty in clearing airway secretion results from a defect in mucociliary transport system rather than from an increase in airway secretion.

    View details for Web of Science ID A1988L993400033

    View details for PubMedID 3341633

  • PULMONARY CLEARANCE OF TC-99M-DTPA IN INFANTS WHO SUBSEQUENTLY DEVELOP BRONCHOPULMONARY DYSPLASIA AMERICAN REVIEW OF RESPIRATORY DISEASE OBRODOVICH, H., Coates, G. 1988; 137 (1): 210-212

    Abstract

    The lung clearance of inhaled and deposited 99mTc-DTPA is biphasic, with a very rapid initial clearance in premature infants with acute hyaline membrane disease (HMD). Infants who rapidly recover from HMD revert to a monophasic slower clearance rate prior to successful extubation. This raises the possibility that a persistently fast clearance might identify infants who will not recover and in whom bronchopulmonary dysplasia (BPD) will subsequently be diagnosed. To examine this possibility, we reviewed our experience using this technique and identified infants who had subsequently developed BPD. These 8 intubated, ventilator-dependent, premature infants inhaled a submicronic aerosol containing 99mTc-DTPA on 14 occasions during the second, third, and fourth weeks of life. Clearance was biphasic, with very rapid initial lung clearance (T1/2 less than 2 min) on 5 occasions in 4 infants during their second or third week of life. The other infants had monophasic clearances that were much slower (T1/2 = 52 +/- 26 SD min). At Day 28 of life, all infants with BPD were ventilator-and/or oxygen-dependent. We conclude that lung clearance of 99Tc-DTPA is very rapid during acute HMD, but that it can normalize before 28 days of life in infants in whom BPD is subsequently diagnosed.

    View details for Web of Science ID A1988L634100040

    View details for PubMedID 3276254

  • LONG-TERM OUTCOME OF PRETERM INFANTS WITH RESPIRATORY-DISEASE CLINICS IN PERINATOLOGY Saigal, S., OBRODOVICH, H. 1987; 14 (3): 635-650

    Abstract

    Improved survival of preterm ventilated infants has resulted in an increase in the incidence of bronchopulmonary dysplasia, particularly in very tiny infants. In this article, the authors have reviewed the current literature on the pulmonary function and long-term neurodevelopmental outcome of VLBW infants to determine the true morbidity of respiratory disease in the neonatal period.

    View details for Web of Science ID A1987J999300010

    View details for PubMedID 2444378

  • OPERATION EVEREST .2. COAGULATION SYSTEM DURING PROLONGED DECOMPRESSION TO 282 TORR JOURNAL OF APPLIED PHYSIOLOGY Andrew, M., OBRODOVICH, H., Sutton, J. 1987; 63 (3): 1262-1267

    Abstract

    Thromboembolic phenomena may occur as humans ascend to high altitude. To investigate the role of the coagulation cascade and its inhibitors in these disorders, venous blood was obtained from eight subjects who participated in the Operation Everest II project. Samples were obtained before and 5 min after completion of a progressive incremental exercise test to exhaustion at sea level and atmospheric pressures of 380 (18,000 ft) and 282 Torr (25,000 ft). Plasma was analyzed for the activity or concentration of factors II, V, VII, VIII complex, IX-XIII, prekallikrein, high-molecular-weight kininogen, fibrinogen, antithrombin III, alpha 2-macroglobulin, alpha 2-antiplasmin, C1-esterase inhibitor, alpha 1-antitrypsin, and protein C. Prolonged exposure to simulated high altitude did not alter the concentration of any of the coagulation factors or inhibitors. Exercise increased the circulating concentrations of the factor VIII complex at sea level, 380, and 282 Torr. However, the increment was less at the simulated high altitudes. The increase in the factor VIII complex was inversely related to arterial O2 saturation and directly related to the work load achieved and blood pH and plasma lactate concentrations. These studies suggest that the gradual development of marked chronic hypoxia does not affect the coagulation cascade.

    View details for Web of Science ID A1987K088300052

    View details for PubMedID 3115952

  • ASSESSMENT OF POSTMORTEM RESPIRATORY CILIARY MOTILITY AND ULTRASTRUCTURE AMERICAN REVIEW OF RESPIRATORY DISEASE Lee, R. M., ROSSMAN, C. M., OBRODOVICH, H. 1987; 136 (2): 445-447

    Abstract

    Acquired and congenital abnormalities in the mucociliary transport system result in respiratory dysfunction and clinical disease. Although upper and lower respiratory epithelium can be analyzed premortem, it is uncertain whether postmortem analyses provide a valid indicator of the premortem status of the ciliary apparatus. To address this question, we studied nasal ciliary motility and ultrastructure from 2 patients 16 h after death. After placement of the epithelial samples in Krebs-Henseleit solution and warming to body temperature, normal ciliary motility and frequency were observed in both samples. The specimen then could be stored within the solution at 4 degrees C for subsequent reanalysis on later days for as long as 60 h after collection with no deterioration in ciliary motility, as assessed by video motion analysis. Quantitative assessment of the ciliary ultrastructure did not reveal any abnormality in microtubular arrangements, and good tissue preservation was seen. These studies demonstrate that postmortem collection of nasal epithelium samples can provide a valid assessment of premortem ciliary structure and function.

    View details for Web of Science ID A1987J583200037

    View details for PubMedID 3619203

  • VENTILATION SCINTIGRAPHY WITH SUBMICRONIC RADIOAEROSOL AS AN ADJUNCT IN THE DIAGNOSIS OF CONGENITAL LOBAR EMPHYSEMA JOURNAL OF NUCLEAR MEDICINE Loewy, J., OBRODOVICH, H., Coates, G. 1987; 28 (7): 1213-1217

    Abstract

    When clinical examination and routine chest x-ray do not adequately explain neonatal respiratory distress, lung scintigraphy using a submicronic aerosol particle may be most helpful. Three cases illustrating this point are presented. Discussion centers around the diagnosis of an atypical case of congenital lobar emphysema (CLE), and differentiating between CLE, foreign body aspiration and compensatory hyperinflation in neonates with respiratory distress. Conservative and surgical treatment options for CLE are also illustrated.

    View details for Web of Science ID A1987J068600021

    View details for PubMedID 3598706

  • EXTRAPULMONARY RADIOACTIVITY IN LUNG PERMEABILITY MEASUREMENTS JOURNAL OF NUCLEAR MEDICINE Coates, G., OBRODOVICH, H. 1987; 28 (5): 903-906

    Abstract

    The pulmonary clearance rate of aerosolized and deposited [99mTc]DTPA is used to assess pulmonary epithelial permeability to solutes. To evaluate whether it is necessary to correct these measurements for radioactivity in the chest wall and pulmonary vasculature five patients with no ventilation to one hemithorax were studied. After inhaling a submicronic aerosol of [99mTc]DTPA a gamma camera measured count rates over both hemithoraces for 20 min. The observed T1/2 for clearance from the normal hemithorax was 56 min (range 18-115 min), and when this was corrected for chest wall contribution (derived from the abnormal hemithorax) the average T1/2 was 52 min (range 17-107 min). To simulate infinitely permeable lungs we measured thoracic radioactivity in two adults and six children who were injected i.v. with a known amount of [99mTc]DTPA. The count rate over the thorax was only 8.1% (range 2.7%-11%) of that obtained when a similar amount of aerosolized [99mTc]DTPA was deposited in the lungs during a subsequent study. We conclude that it is not necessary to correct for nonpulmonary epithelial radioactivity during the measurement of [99mTc]DTPA clearance rate from the lungs.

    View details for Web of Science ID A1987H287600020

    View details for PubMedID 3553464

  • PULMONARY CLEARANCE OF TC-99M-DTPA - A NONINVASIVE ASSESSMENT OF EPITHELIAL INTEGRITY LUNG OBRODOVICH, H., Coates, G. 1987; 165 (1): 1-16

    View details for Web of Science ID A1987F767500001

    View details for PubMedID 3104696

  • PULMONARY HEMODYNAMICS DURING EXERCISE AFTER LOW AND HIGH-DOSES OF ACETYLSALICYLIC-ACID IN SHEEP RESPIRATION Coates, G., OBRODOVICH, H., Jefferies, A. L. 1987; 52 (2): 94-100

    Abstract

    Despite comparable increases in cardiac output there is a greater reduction in pulmonary vascular resistance (PVR) in sheep during exercise than in dog or man. The mechanism for this marked reduction in PVR in sheep is unknown. To assess the role of arachidonic acid metabolites in producing this low PVR we measured the effect of intravenous acetylsalicylic acid (ASA, 10 mg/kg) on PVR at rest and during exercise in sheep. ASA caused a slight rise in resting PVR (p less than 0.05), but did not affect the exercise-induced decrease in PVR. High-dose ASA (100 mg/kg), presumably sufficient to block the lipoxygenase pathway produced the same responses as low-dose ASA which should only block the cyclo-oxygenase pathway. Products of the cyclo-oxygenase and lipoxygenase pathways which include vasodilator prostaglandins, have only a minor role in maintaining low pulmonary vascular resistance at rest and have no demonstrable role in the reduced PVR that occurs during exercise in sheep.

    View details for Web of Science ID A1987K377200003

    View details for PubMedID 3118440

  • PULMONARY DEPOSITION SITES OF AN INHALED RADIOLABELED SUBMICRONIC AEROSOL PEDIATRIC RESEARCH Kay, J., Coates, G., OBRODOVICH, H. 1986; 20 (12): 1297-1300

    Abstract

    The clearance rate of aerosolized and deposited 99mtechnetium diethylenetriamine pentacetate provides an index of lung epithelial permeability. However, the location of the epithelium being assessed is uncertain. We determined the percentage of submicronic aerosol deposited on ciliated and nonciliated airways in healthy and damaged lungs using 99mTc-sulphur (99mTc-S) colloid. Colloidal particles can be cleared by mucociliary transport but not through the epithelium. Piglets aged 12-72 h, weighing 0.7-3.3 kg (average 1.6 kg) were anesthetized and underwent endotracheal intubation. 99mTc-S labeled submicronic aerosol (Syntevent II) was collected in anesthetic bags and either inhaled spontaneously (n = 4), or insufflated by hand (n = 5) at 30 breaths per min and a peak airway pressure of 15 cm H2O. Piglets were immediately extubated and scanned on a gamma camera. Twenty-four h later a repeat scan was performed. The residual radioactivity represents the amount of aerosol deposited on nonciliated airways. In six other piglets pulmonary damage was produced by an intravenous infusion of air microemboli for 2 to 3 h (0.14 to 0.38 ml/min). The maximal decrease in PaO2 while breathing room air ranged from 24-59 mm Hg. During the last 15 min of infusion they were intubated, insufflated by hand with the 99mTc-S aerosol, and scanned as above. The results demonstrate that approximately three-fourths of the submicronic aerosol is deposited distally to ciliated airways in both healthy and damaged newborn piglet lungs. This suggests that the clearance rate of 99mtechnetium diethylenetriamine pentacetate predominantly reflects epithelial permeability of terminal lung units when a submicronic aerosol is used for delivery to the lung.

    View details for Web of Science ID A1986F145800017

    View details for PubMedID 3797121

  • EFFECT OF EXERCISE ON LUNG LYMPH-FLOW IN UNANESTHETIZED SHEEP WITH INCREASED PULMONARY VASCULAR-PERMEABILITY AMERICAN REVIEW OF RESPIRATORY DISEASE OBRODOVICH, H., Coates, G. 1986; 134 (5): 862-866

    Abstract

    To investigate the effect of increased cardiac output on lung water and solute movement in abnormally permeable lungs, experiments were performed in sheep with chronic vascular catheters and lung lymph fistulas. Pulmonary permeability was increased by infusing air microemboli intravenously for 3 h. Once stable lymph flow was observed after stopping the air embolization sheep were either exercised on a treadmill to increase cardiac output (n = 10) or merely continuously monitored (control experiments, n = 6). In control experiments, lung lymph flow remained at markedly elevated but stable values. Sheep exercised for a median time of 45 min. Cardiac output increased 85%, whereas lung lymph flow, in terms of baseline flow rates, increased 296%. The lymph to plasma concentration ratio for total protein was unchanged, and left atrial pressure remained stable and normal during exercise. During exercise, pulmonary arterial pressure increased by 3 mm Hg. The data, when compared with previous observations in normal sheep, demonstrate that exercise-induced increases in cardiac output result in disproportionate increases in lung lymph flow when pulmonary vascular permeability is increased.

    View details for Web of Science ID A1986E792100006

    View details for PubMedID 3777682

  • MEASUREMENT OF PULMONARY EPITHELIAL PERMEABILITY WITH TC-99M-DTPA AEROSOL SEMINARS IN NUCLEAR MEDICINE Coates, G., OBRODOVICH, H. 1986; 16 (4): 275-284

    Abstract

    The rate at which inhaled aerosol of 99mTc-diethylenetriamine pentaacetate (DTPA) leaves the lung by diffusion into the vascular space can be measured with a gamma camera or simple probe. In normal humans, 99mTc-DTPA clears from the lung with a half time of about 80 minutes. Many acute and chronic conditions that alter the integrity of the pulmonary epithelium cause an increased clearance rate. Thus cigarette smoking, alveolitis from a variety of causes, adult respiratory distress syndrome (ARDS), and hyaline membrane disease (HMD) in the infant have all been shown to be associated with rapid pulmonary clearance of 99mTc-DTPA. Rapid clearance is also promoted by increased lung volume and decreased surfactant activity. Although the mechanism of increased clearance in pathological states is not known, the 99mTc-DTPA lung-clearance technique has great potential clinically, particularly in patients at risk from ARDS and HMD and in the diagnosis and follow-up of alveolitis.

    View details for Web of Science ID A1986E973300004

    View details for PubMedID 3541208

  • INCREASES IN FACTOR-VIII COMPLEX AND FIBRINOLYTIC-ACTIVITY ARE DEPENDENT ON EXERCISE INTENSITY JOURNAL OF APPLIED PHYSIOLOGY Andrew, M., Carter, C., OBRODOVICH, H., Heigenhauser, G. 1986; 60 (6): 1917-1922

    Abstract

    Components of the factor VIII complex increase and activation of the fibrinolytic system occur during exercise. The relation between the duration and intensity of exercise and the relative changes in the VIII complex and fibrinolytic system have not been previously examined. Five healthy male subjects were exercised with three protocols: a graded progressive exercise test to exhaustion on a cycle ergometer with 50-W increments every 4 min, steady-state exercise, 15 min at 5 and 125 W each, and an acute 30-s maximal exercise test on a cycle ergometer. Venous blood samples were drawn at base line, during the last 30 s of each power output in the graded exercise, at 5-min intervals for the steady-state exercise, and for up to 1 h after completion of exercise in all three protocols. At the maximum exercise intensities, increases in plasma lactate concentration ([La]), O2 uptake, and [H+] were observed. Components of the VIII complex [VIII procoagulant, VIII procoagulant antigen, VIII-related antigen (VIIIR:Ag), VIII ristocetin cofactor activity] abruptly rose at only the highest work intensities, whereas the whole blood clot lysis time began to gradually shorten much earlier at low work intensities. There were no qualitative changes in the factor VIIIR:Ag on crossed immunoelectrophoresis nor was there evidence of thrombin generation as determined by fibrinopeptide A generation. We conclude that during exercise the changes observed in the coagulation and fibrinolytic systems are related to the intensity of the exercise, which is reflected by increases in plasma [La] and [H+], and that the fibrinolytic system is activated before the changes in the VIII complex are observed.

    View details for Web of Science ID A1986C850400016

    View details for PubMedID 3087936

  • EFFECT OF INSPIRATORY RESISTANCE AND PEEP ON TC-99M-DTPA CLEARANCE JOURNAL OF APPLIED PHYSIOLOGY OBRODOVICH, H., Coates, G., Marrin, M. 1986; 60 (5): 1461-1465

    Abstract

    Experiments were performed to determine the effect of markedly negative pleural pressure (Ppl) or positive end-expiratory pressure (PEEP) on the pulmonary clearance (k) of technetium-99m-labeled diethylenetriaminepentaacetic acid (99mTc-DTPA). A submicronic aerosol containing 99mTc-DTPA was insufflated into the lungs of anesthetized intubated sheep. In six experiments k was 0.44 +/- 0.46% (SD)/min during the initial 30 min and was unchanged during the subsequent 30-min interval [k = 0.21 +/- 12%/min] when there was markedly increased inspiratory resistance. A 3-mm-diam orifice in the inspiratory tubing created the resistance. It resulted on average in a 13-cmH2O decrease in inspiratory Ppl. In eight additional experiments sheep were exposed to 2, 10, and 15 cmH2O PEEP (20 min at each level). During 2 cmH2O PEEP k = 0.47 +/- 0.15%/min, and clearance increased slightly at 10 cmH2O PEEP [0.76 +/- 0.28%/min, P less than 0.01]. When PEEP was increased to 15 cmH2O a marked increase in clearance occurred [k = 1.95 +/- 1.08%/min, P less than 0.001]. The experiments demonstrate that markedly negative inspiratory pressures do not accelerate the clearance of 99mTc-DTPA from normal lungs. The effect of PEEP on k is nonlinear, with large effects being seen only with very large increases in PEEP.

    View details for Web of Science ID A1986C311700001

    View details for PubMedID 3519565

  • AEROSOL DELIVERY TO THE RABBIT LUNG WITH AN INFANT VENTILATOR PEDIATRIC PULMONOLOGY Flavin, M., MacDonald, M., Dolovich, M., Coates, G., OBRODOVICH, H. 1986; 2 (1): 35-39

    Abstract

    Little is known about delivery of aerosolized medications to infants undergoing assisted positive pressure ventilation. To assess delivery of medications to the lung with an infant ventilator (Bournes LS104), 27 experiments were performed on anesthetized adult rabbits. Lung deposition was determined by using aerosol radiolabeled with 99m technetium sulfur colloid. Initially a traditional nebulizer was studied and very inefficient delivery to the lung was observed (0.19 +/- 0.10 SD% of the initial nebulizer dose). Subsequent experiments using a nebulizer that generates submicronic aerosol particles, at equivalent aerosol output, achieved a highly significant increase in delivery (1.96 +/- 1.19 SD%, P less than 0.001). Our experiments demonstrated that modifications to to traditional nebulizer systems can enhance delivery of aerosolized medication to the lungs of rabbits.

    View details for Web of Science ID A1986A582500007

    View details for PubMedID 3513104

  • EFFECT OF ISOPROTERENOL OR EXERCISE ON PULMONARY LYMPH-FLOW AND HEMODYNAMICS JOURNAL OF APPLIED PHYSIOLOGY OBRODOVICH, H., Coates, G. 1986; 60 (1): 38-44

    Abstract

    Experiments were performed to determine whether different methods of increasing cardiac output would have similar effects on lung lymph flow, and to assess the contribution of the microvasculature (fluid-exchanging vessels) to the total calculated pulmonary vascular resistance. Yearling unanesthetized sheep with chronic vascular catheters and lung lymph fistulas underwent intravenous infusions of isoproterenol at 0.2 micrograms X kg-1. min-1 (n = 8) or were exercised on a treadmill (n = 16). Both isoproterenol and exercise increased cardiac output, lowered calculated total pulmonary and systemic vascular resistances, and had no effect on the calculated pulmonary microvascular pressure. Isoproterenol infusions did not affect lung lymph flow, whereas exercise increased lung lymph flow in proportion to the increase in cardiac output. We conclude that 1) the sheep has a different pulmonary hemodynamic response to exercise than dogs and man, 2) the microvasculature is recruited during exercise-induced but not isoproterenol-induced increases in cardiac output, and 3) the microvasculature represents only a small proportion of the total calculated pulmonary vascular resistance.

    View details for Web of Science ID A1986AXV0900006

    View details for PubMedID 3944045

  • THE CONTRIBUTION OF LYMPHATIC DRAINAGE TO THE CLEARANCE OF INHALED TC-99M-DTPA FROM THE LUNGS CLINICAL AND INVESTIGATIVE MEDICINE-MEDECINE CLINIQUE ET EXPERIMENTALE Coates, G., OBRODOVICH, H. M. 1986; 9 (1): 15-20

    Abstract

    When inhaled as an aerosol, 99mTc labelled diethylene triamine pentacetate (99mTc-DTPA) moves rapidly from the airspace to the vascular space. The rate at which it leaves the lungs is being used to measure the integrity of the pulmonary epithelium. In order to determine what part the lymphatic system plays in the clearance of 99mTc-DTPA from the lungs, we measured the rate of appearance in plasma and lymph of inhaled 99mTc-DTPA and intravenously injected Indium-113m labelled (113mIn-DTPA) in 5 sheep with chronic lung lymph fistalae. Inhaled 99mTc-DTPA was detected in the plasma and lymph after 1 minute. This suggests that the inhaled sub-micronic aerosol of 99mTc-DTPA was deposited predominately in a region of the lung with a large vascular surface area, ie. the terminal lung units. The lymph/plasma concentration (1/p) ratio for injected 113mIn-DTPA became greater than 1 by 4 minutes whereas the 1/p ratio for inhaled 99mTc-DTPA did not reach 1 until 25 minutes. This suggests that lymph drainage has very little part to play in the clearance of inhaled 99mTc-DTPA from the lungs.

    View details for Web of Science ID A1986A337400004

    View details for PubMedID 3514019

  • Retraction of two articles: Bradykinin. Journal of applied physiology STALCUP, S. A., LIPSET, J. S., LEGANT, P. M., LEUENBERGER, P. J., Pang, L. M., O'Brodovich, H. M., Mellins, R. B. 1985; 59 (4): 1333-?

    View details for PubMedID 3902780

  • On pulmonary epithelial permeability. Pediatric pulmonology O'Brodovich, H. 1985; 1 (5): 291-?

    View details for PubMedID 4069819

  • PULMONARY EPITHELIAL PERMEABILITY IN HYALINE-MEMBRANE DISEASE - REPLY NEW ENGLAND JOURNAL OF MEDICINE Jefferies, A. L., OBRODOVICH, H. M., Coates, G. 1985; 312 (9): 586-586
  • HYPOXIA-INDUCED VASOPRESSIN RELEASE AND COAGULOPATHY IN A NORMAL SUBJECT AVIATION SPACE AND ENVIRONMENTAL MEDICINE Andrew, M., OBRODOVICH, H., Coates, G., Robertson, G. L., Gray, G. W. 1985; 56 (12): 1220-1223

    Abstract

    Humans exposed to hypoxia usually increase their plasma procoagulant VIII activity (VIII:C) with no change in the concentration of VIII related antigen (VIIIR:Ag). This case report describes an apparently normal subject who developed marked qualitative and quantitative changes in all components of the factor VIII complex while inhaling an 11% oxygen/balance nitrogen gas mixture for 2 h. Blood from fresh venepunctures was drawn at baseline, during and after exposure to hypoxia for the following: a partial thromboplastin time, a prothrombin time, fibrin monomer, factor VIII:C, VIII procoagulant antigen (VIII:CAg); ristocetin cofactor activity (VIIIR:Co); VIII von Willebrand factor (VIII:vWF) multimer pattern; and arginine vasopressin. During hypoxia VIII:C, VIII:CAg, VIIIR:Ag and VIIIR:Co increased 4 to 5 fold; the VIII:vWF multimer pattern showed increasing low molecular weight complexes, fibrin monomer appeared and arginine vasopressin (AVP) levels increased from 5.5 pg . ml-1 to 73.8 pg . ml-1. These changes are compatible with both the release of the VIIIR:Ag by AVP and protease induced fragmentation of the VIII complex.

    View details for Web of Science ID A1985AVP3900016

    View details for PubMedID 3936469

  • PULMONARY THROMBOEMBOLI AFTER NEONATAL ASPHYXIA JOURNAL OF PEDIATRICS Arnold, J., OBRODOVICH, H., Whyte, R., Coates, G. 1985; 106 (5): 806-809

    Abstract

    Two surviving neonates with pulmonary thromboembolism, diagnosed by ventilation-perfusion lung scan, are described. Both infants had respiratory distress, and one had features consistent with persistent pulmonary hypertension of the neonate. These reports demonstrate that substantive pulmonary emboli can occur in neonates and may not be recognized without an appropriate level of clinical suspicion.

    View details for Web of Science ID A1985AHF6000024

    View details for PubMedID 3998922

  • BRADYKININ IS DEGRADED IN HYPOXIC LUNGS AND DOES NOT AFFECT EPITHELIAL PERMEABILITY JOURNAL OF APPLIED PHYSIOLOGY OBRODOVICH, H., Kay, J., Coates, G. 1985; 59 (4): 1185-1190

    Abstract

    To investigate the effect of intravenous infusions of bradykinin (BK) on the permeability of the hypoxic pulmonary epithelium to small solutes, experiments (n = 7) were performed in yearling sheep with chronic vascular catheters. Sheep were anesthetized, intubated, paralyzed, and ventilated. After establishing stable and normal base-line pulmonary hemodynamics and blood gas tensions, the lungs were insufflated with a submicronic aerosol of technetium-99m-labeled diethylenetriaminepentaacetate (99mTc-DTPA, mol wt = 492). Radioactivity arising from the right hemithorax was measured by an NaI probe with a parallel-holed collimator. The base-line pulmonary clearance rate (k) for 99mTc-DTPA was 0.51 +/- 0.09% (SE)/min, while the sheep were ventilated with a fractional concentration of inspired O2 (FIO2) of 0.5 [arterial partial pressure of O2 (PaO2) = 196 +/- 11.4 (SE) Torr]. Clearance of 99mTc-DTPA was unaffected by hypoxia alone or BK infusions in nonhypoxic lungs. The combination of an intravenous infusion of BK at either 1.2 (n = 3) or 2.4 micrograms . kg-1 . min-1 (n = 4) and alveolar hypoxia [FIO2 = 0.11, PaO2 = 28 +/- 1.6 (SE) Torr] did not affect pulmonary clearance of 99mTc-DTPA [k = 0.43 +/- 0.08% (SE)/min]. In contrast, a 0.05-ml/kg intravenous infusion of oleic acid increased clearance 10-fold in one sheep. During combined hypoxia and BK infusion the pulmonary arterial BK concentration (radioimmunoassay) increased from 0.82 +/- 0.16 (SE) to 7.05 +/- 1.86 ng/ml (P less than 0.001), but the systemic arterial concentrations were unchanged [0.67 +/- 0.19 (SE) to 0.66 +/- 0.09 ng/ml].(ABSTRACT TRUNCATED AT 250 WORDS)

    View details for Web of Science ID A1985ASP6600022

    View details for PubMedID 3902777

  • BRONCHOPULMONARY DYSPLASIA - UNRESOLVED NEONATAL ACUTE LUNG INJURY AMERICAN REVIEW OF RESPIRATORY DISEASE OBRODOVICH, H. M., Mellins, R. B. 1985; 132 (3): 694-709

    View details for Web of Science ID A1985AQL8900042

    View details for PubMedID 3898946

  • FLUOSOL-DA CAUSES PULMONARY-HYPERTENSION AND INCREASED LUNG LYMPH-FLOW IN UNANESTHETIZED SHEEP CLINICAL AND INVESTIGATIVE MEDICINE-MEDECINE CLINIQUE ET EXPERIMENTALE OBRODOVICH, H., BELBECK, L., Andrew, M., Coates, G. 1985; 8 (1): 15-21

    Abstract

    Test doses (0.5 ml) of the perfluorochemical blood substitute Fluosol-DA causes pulmonary hypertension in some patients. To investigate this phenomenon we infused Fluosol-DA into unanesthetized sheep with chronic vascular catheters on ten occasions. In six of these experiments lung lymph flow was measured. Since other fluorochemical blood substitutes alter the coagulation cascade we also sampled blood to perform coagulation screening tests and look for evidence of thrombin generation as assessed by the survival of 125I labelled sheep fibrinogen. Test doses (0.5 ml) of Fluosol-DA resulted in transient but marked pulmonary hypertension in nine of ten experiments (17 +/- 1.2 SE to 43 +/- 3.9 SE torr). Cardiac output decreased by an average of 30%, and lung lymph flow transiently increased without a change in the lymph to plasma protein concentration (L/P) ratio in five of six experiments. When 50 ml of Fluosol-DA was administered one hour later the pulmonary hypertension was more prolonged but less severe (18 +/- 1.0 SE to 34 +/- 3.2 SE torr). Lymph flow again increased without a change in the L/P ratio for protein. The activated partial thromboplastin and prothrombin times, and survival in plasma of 125I labelled sheep fibrinogen were unaffected by administration of Fluosol-DA. We conclude that Fluosol-DA causes pulmonary hypertension in sheep and increases lung lymph flow. This results from vasoconstriction of the pulmonary vessels and subsequent recruitment of the pulmonary microvasculature and not from thromboembolic phenomena.

    View details for Web of Science ID A1985AGV6100004

    View details for PubMedID 3987115

  • PULMONARY EPITHELIAL PERMEABILITY IN HYALINE-MEMBRANE DISEASE INNERE MEDIZIN Jefferies, A. L., Coates, G., OBRODOVICH, H. 1985; 12 (4): 204-205
  • TRYPAN BLUE METHOD FOR THE IDENTIFICATION OF AREAS OF DAMAGE TO AIRWAY EPITHELIUM DUE TO MECHANICAL TRAUMA SCANNING ELECTRON MICROSCOPY Lee, R. M., Chambers, C., OBRODOVICH, H., Forrest, J. B. 1984: 1267-1271

    Abstract

    A method has been developed for locating the regions of pulmonary epithelial damage due to mechanical trauma, based on the inability of the injured cells to exclude vital stains such as trypan blue. Scanning electron microscopy was used to confirm the sites, as well as the specific type of mechanical damage. The damaged area on rabbit tracheal epithelium due to gentle stroking was characterized by denudation of the ciliated epithelial cells, leaving only a layer of flat, non-ciliated cells. Repeated gentle stroking of the epithelial surface removed all the epithelial cells, exposing the underlying connective tissue layer. The trypan blue method provides a fast and sensitive means of locating damaged airway epithelium due to mechanical trauma. Because the dye is non-toxic, the repair process of the epithelium can also be studied, following the initial assessment of the extent of epithelial damage.

    View details for Web of Science ID A1984TM34700027

    View details for PubMedID 6505613

  • CILIARY DEFECTS ASSOCIATED WITH THE DEVELOPMENT OF BRONCHOPULMONARY DYSPLASIA - CILIARY MOTILITY AND ULTRASTRUCTURE AMERICAN REVIEW OF RESPIRATORY DISEASE Lee, R. M., ROSSMAN, C. M., OBRODOVICH, H., Forrest, J. B., Newhouse, M. T. 1984; 129 (1): 190-193

    Abstract

    We have on several occasions studied the nasal respiratory epithelium of an infant with hyaline membrane disease that evolved into bronchopulmonary dysplasia and observed an association between the clinical status and naso-ciliary motion and ultrastructure. At 4 months of age, when the patient had significant respiratory disease, few cilia were present and they beat with a slow dyskinetic motion. The specimens contained primarily necrotic and squamous epithelial membranes; the occasional cilia present had swollen or ruptured membranes. Partial recovery of the epithelium was noted at 4.5 months, with 45% of the cilia having normal ultrastructure. The beat frequency was 15.2 +/- 1.5 Hz (mean +/- SD), and although some degree of dyskinesia was evident, primarily normal ciliary motion was observed. By 10 months of age, significant clinical improvement had occurred and the nasal epithelium had regenerated; 96% of the cilia had normal ultrastructure, and the ciliary beat frequency (12.4 +/- 1.2 Hz) and motion were normal.

    View details for Web of Science ID A1984RY55000039

    View details for PubMedID 6703479

  • PULMONARY EPITHELIAL PERMEABILITY IN HYALINE-MEMBRANE DISEASE NEW ENGLAND JOURNAL OF MEDICINE Jefferies, A. L., Coates, G., OBRODOVICH, H. 1984; 311 (17): 1075-1080

    Abstract

    Neonatal hyaline-membrane disease is complicated by pulmonary edema, yet left atrial pressures are normal. Alveolar-capillary-membrane permeability may therefore be increased. To assess pulmonary epithelial permeability, we measured the pulmonary clearance and half-life of aerosolized 99mTc-diethylenetriamine pentacetate (99mTc-DTPA) on 31 occasions in 15 intubated premature infants with hyaline-membrane disease. Three infants with respiratory failure due to other diseases were studied on four occasions. All studies of infants with hyaline-membrane disease that were performed in the first 72 hours of life demonstrated a biphasic clearance curve with a rapid-phase half-life of 1.6 +/- 0.6 minutes (mean +/- S.D.). As these infants recovered, the curve became monophasic with a half-life of 56.0 +/- 32.1 minutes. Two infants remained dependent on oxygen and ventilator support and had persistent biphasic curves with a rapid-phase half-life of 1.5 +/- 0.7 minutes. All infants without hyaline-membrane disease had monophasic curves with a half-life of 65.4 +/- 33.6 minutes. Using a similar technique, we observed that newborn lambs and piglets have a monophasic pulmonary clearance of 99mTc-DTPA (114 +/- 59 minutes in lambs and 52.5 +/- 16.3 minutes in piglets). We conclude that the lungs of neonates with hyaline-membrane disease are abnormally permeable to small solutes and that this abnormality persists in infants with subsequent chronic lung disease.

    View details for Web of Science ID A1984TP17700003

    View details for PubMedID 6384782

  • EFFECTS OF EXERCISE ON LUNG LYMPH-FLOW IN SHEEP AND GOATS DURING NORMOXIA AND HYPOXIA JOURNAL OF CLINICAL INVESTIGATION Coates, G., OBRODOVICH, H., Jefferies, A. L., Gray, G. W. 1984; 74 (1): 133-141

    Abstract

    Vigorous exercise causes a marked increase in cardiac output with only a minimal increase in measureable pulmonary vascular pressures. These changes in pulmonary hemodynamics should affect lung water and solute movement. On nine occasions, we measured the effect of normoxic exercise on lung lymph flow in four sheep and two goats with chronic lymph fistulas (wt = 15-25 kg). In addition, lymph flow was also measured on five occasions in sheep during exercise at reduced barometric pressures (430 and 380 mmHg). During normobaria, the animals ran at 3-5 km/h with 0-10% elevation of the treadmill for 15 to 85 min. Exercise on average caused a 100% increase in cardiac output, a 140% increase in lung lymph flow, and a slight but significant reduction in lymph to plasma concentration ratio (l/p) for total protein and albumin (mol wt = 70,000). There was a significant linear correlation between lymph flow and cardiac output (r = 0.87, P less than 0.01). There was no change in l/p for IgG (mol wt = 150,000) or IgM (mol wt = 900,000) and no significant change in mean pulmonary arterial (Ppa) or mean left atrial (Pla) pressures. Transition from normobaria to hypobaria caused an increase in Ppa but no change in Pla, cardiac output, or lymph flow. Exercise during hypobaria caused increases in lymph flow that were qualitatively similar to changes observed during normobaric exercise: there was a 60% increase in cardiac output, a 90% increase in lymph flow, and an 11% reduction in l/p for total protein. There was no change in l/p for albumin, IgG, or IgM, and no further change in Ppa. The increased lymph flow during normoxic and hypobaric exercise is best explained by an increase in pulmonary vascular surface area for fluid and protein exchange. Our results suggest that the normal ovine lung has the potential to nearly triple the amount of perfused microvascular surface area. This speculation is relevant to the interpretation of lymph flow data from other experiments.

    View details for Web of Science ID A1984SY22400016

    View details for PubMedID 6736245

  • HYPOXIA ALTERS BLOOD-COAGULATION DURING ACUTE DECOMPRESSION IN HUMANS JOURNAL OF APPLIED PHYSIOLOGY OBRODOVICH, H. M., Andrew, M., Gray, G. W., Coates, G. 1984; 56 (3): 666-670

    Abstract

    Acute decompression is associated with a shortening of the activated partial thromboplastin time (aPTT). This study was performed to examine whether this change in aPTT results from hypoxia or hypobaria. We exposed healthy adults on three separate occasions to 2 h of 1) hypoxic hypobaria (410 Torr, n = 5), 2) hypoxic normobaria (fractional inspired O2 tension = 0.11, n = 4), or 3) normoxic hypobaria (410 Torr breathing supplemental O2, n = 5). The aPTT shortened during hypoxic hypobaria and hypoxic normobaria (P less than 0.05) but was unchanged during normoxic hypobaria. The prothrombin and thrombin times, hematocrit, and concentrations of fibrinogen, total plasma protein, and fibrinogen-fibrin fragment E were unchanged. During hypoxic hypobaria biologic levels of prekallikrein, high-molecular-weight kininogen, and factors XII, XI, X, VII, V, and II were unchanged, but procoagulant VIII (VIII:C) increased 50% without an increase in VIII-related antigen levels (VIIIR:Ag). Fibrin monomer was not detected in any group. In one subject who became ill after 1.5 h of hypoxic normobaria aPTT shortened by 10 s; the platelet count decreased by 93,000/mm3; VIII:C increased fivefold, but VIIIR:Ag only increased three-fold. We conclude that it is the hypoxia which shortens aPTT during acute decompression to 410 Torr and speculate that it results from an increase in plasma VIII:C-like activity.

    View details for Web of Science ID A1984SH56300017

    View details for PubMedID 6423590

  • MEASUREMENT OF PULMONARY CLEARANCE OF RADIOAEROSOL USING A PORTABLE SODIUM-IODIDE PROBE JOURNAL OF APPLIED PHYSIOLOGY Jefferies, A. L., Coates, G., Webber, C. E., OBRODOVICH, H. M. 1984; 57 (6): 1908-1912

    Abstract

    To determine whether a portable sodium iodide (NaI) probe could provide a valid measure of the pulmonary half-life (T1/2) of aerosolized technetium-99m-diethylenetriaminepentaacetate (99mTc-DTPA, mol wt = 492) in small chests, we measured pulmonary clearance in rabbits using a gamma-scintillation camera and the portable probe. In 10 experiments the lungs of New Zealand White rabbits were insufflated with aerosolized 99mTc-DTPA (0.6 mum aerodynamic mass median diameter) and then simultaneously imaged with the gamma-camera and the probe positioned over the upper right lung. In an additional 12 experiments, alveolar-capillary membrane permeability was increased by either intratracheal instillation of 0.1 N hydrochloric acid (HCl) or intravenous injection of 100 mg/kg of oleic acid. All animals tolerated the procedure. There was a significant decrease in pulmonary T1/2 in both the HCl group (53.4 +/- 10.4 min, mean +/- SE) and the oleic acid group (14.7 +/- 2.3 min) when compared with control (127.5 +/- 18.1 min). When we compared the T 1/2 of the right lung determined by the gamma-camera with that measured by the probe, the correlation coefficient was 0.95. Potential nonpulmonary contributions to thoracic radioactivity were not significant. We conclude that a portable NaI probe is a valid means of determining T 1/2 of 99mTc-DTPA in small chests when compared with a gamma-camera and can detect increases in the permeability of the alveolar-capillary membrane to small solutes.

    View details for Web of Science ID A1984TX58800044

    View details for PubMedID 6392229

  • QUANTITATIVE VENTILATION-PERFUSION LUNG SCANS IN INFANTS AND CHILDREN - UTILITY OF A SUBMICRONIC RADIOLABELED AEROSOL TO ASSESS VENTILATION JOURNAL OF PEDIATRICS OBRODOVICH, H. M., Coates, G. 1984; 105 (3): 377-383

    Abstract

    The quantitative assessment of regional pulmonary ventilation and perfusion provides useful information regarding lung function. Its use in infants and young children, however, has been minimal because of practical and technical limitations when the distribution of ventilation is assessed by radioactive gases. In 16 infants and children we used an inexpensive commercially available nebulizer to produce a submicronic aerosol labeled with 99mtechnetium-diethylenetriamine pentacetic acid to assess ventilation quantitatively, and intravenous injections of 99mtechnetium-labeled macroaggregates of albumin to assess pulmonary perfusion quantitatively. Studies were safely completed in both ambulatory and critically ill patients, including two premature infants who had endotracheal tubes in place for ventilatory support. No sedation or patient cooperation is required. This technique enables any department of nuclear medicine to measure regional pulmonary ventilation and perfusion in infants and children.

    View details for Web of Science ID A1984TH00600006

    View details for PubMedID 6236292

  • PLASMA-CATECHOLAMINES DURING HYPOXEMIA AND HYPERCAPNIA AMERICAN JOURNAL OF PHYSIOLOGY OBRODOVICH, H. 1984; 247 (2): H341-H341

    View details for Web of Science ID A1984TE85900022

    View details for PubMedID 6465336

  • ASSESSMENT OF COAGULATION CASCADE DURING AIR MICROEMBOLIZATION OF THE LUNG JOURNAL OF APPLIED PHYSIOLOGY OBRODOVICH, H., Andrew, M., Silver, R., Coates, G. 1983; 55 (6): 1743-1747

    Abstract

    Experiments were performed to determine whether activation of the coagulation cascade was required for pulmonary vascular permeability to increase during microembolization of the lung. For 30-45 min air microemboli were intravenously infused (0.05-0.10 ml X kg-1 X min-1) into awake sheep with chronic lung-lymph fistulas and anesthetized mongrel dogs. During embolization the pulmonary arterial pressure increased, and O2 partial pressure (PaO2) fell by more than 20 Torr (P less than 0.01). Subsequently lymph flow nearly tripled without a change in the lymph-to-plasma protein concentration ratio. Partial thromboplastin and prothrombin times, biological activity of antithrombin III, and circulating concentration of 125I-labeled dog or sheep fibrinogen did not change during or following air infusion. In two additional sheep an intravenous infusion of thrombin at 0.6 U X kg-1 X min-1 for 15 min resulted in a 20% decrease in 125I-labeled sheep fibrinogen concentration without a change in pulmonary arterial pressure or PaO2. We conclude that air microembolization can increase permeability to water and protein without a detectable activation of the coagulation cascade in the sheep or dog.

    View details for Web of Science ID A1983RV39800015

    View details for PubMedID 6662765

  • EFFECT OF HIGH-FREQUENCY OSCILLATION ON LUNG LYMPH-FLOW JOURNAL OF APPLIED PHYSIOLOGY Jefferies, A. L., Hamilton, P., OBRODOVICH, H. M. 1983; 55 (5): 1373-1378

    Abstract

    We investigated the effect of high-frequency oscillation (HFO) on lung lymphatic function under normal conditions and when lung lymph flow was increased by air microembolization. In six experiments, sheep and goats with chronic lung lymph fistulas and vascular catheters were anesthetized, paralyzed, intubated, and ventilated according to the following protocol: 1) intermittent positive-pressure ventilation (IPPV) for 1 h, 2) HFO with a frequency of 15 Hz and an estimated tidal volume of 1-2 ml/kg for 1-2 h, and 3) IPPV for 0.5 h. Ventilator settings were adjusted to maintain arterial Po2 above 100 Torr and a normal arterial Pco2. Vascular, esophageal, and mean airway pressures were monitored continuously. Lymph flow and cardiac output were recorded every 15 min. With this protocol, there were no changes in pulmonary vascular or esophageal pressures, and lymph flow remained stable throughout the experiment. In an additional five experiments, air microemboli were infused for approximately 30 min during HFO. Left atrial pressure was unchanged and lymph flow tripled. This response was qualitatively and quantitatively similar to that previously reported for unanesthetized spontaneously breathing sheep. We conclude that HFO does not impair lymphatic function under resting conditions and that lymphatics retain their ability to increase water and protein clearance during HFO.

    View details for Web of Science ID A1983RQ80600001

    View details for PubMedID 6358161

  • ROLE OF LYMPHATICS IN REMOVAL OF SHEEP LUNG SURFACTANT LIPID JOURNAL OF APPLIED PHYSIOLOGY Tarpey, M. M., OBRODOVICH, H. M., Young, S. L. 1983; 54 (4): 984-988

    Abstract

    To study the role of lung lymphatics in the removal of surfactant lipid from the sheep lung, we injected [1-14C]palmitate intravenously into six animals previously fitted with a cannula draining the caudal mediastinal lymph node. Lung lymph was collected for 100 h after injection of radiolabel. We obtained alveolar lavage material through a tracheostomy in four other animals after intravenous injection of [9,10-3H]palmitate. We measured radioactivity at several time points in lipid extracts from lymph, lavage fluid, and lung tissue. Alveolar lavage disaturated phosphatidylcholine (DSPC) specific activity peaked at about 40 h and was reduced to 30% of this value by 82 h. About 2% of the injected radiolabel was incorporated into lung tissue lipids. Only 4% of the level of labeling achieved in lung tissue lipids was found in lung lymph lipid during 100 h of lymph collection. Sixty-three percent of radiolabel in lymph lipid was recovered in phospholipids, and 29% of phospholipid radiolabel was found in DSPC. The distribution of phosphorus and palmitate radiolabel in lung lymph phospholipid did not closely resemble that of surfactant lipid. No rise in lung lymph DSPC specific activity was observed following the peak in lavage specific activity. If surfactant lipid is removed from the alveolar compartment without extensive recycling, then we conclude that the lung lymphatics do not play a major role in the clearance of surfactant lipid from the alveolar surface.

    View details for Web of Science ID A1983QL11600019

    View details for PubMedID 6687883

  • NON-INVASIVE DIAGNOSIS OF PULMONARY HEMORRHAGE IN RHEUMATOID-ARTHRITIS PEDIATRICS OBRODOVICH, H. M., Way, R. C., Andrew, M., Dent, P. B. 1983; 72 (5): 720-723

    View details for Web of Science ID A1983RQ01700023

    View details for PubMedID 6634278

  • BRADYKININ-INDUCED INCREASE IN PULMONARY VASCULAR-PERMEABILITY IN HYPOXIC SHEEP JOURNAL OF APPLIED PHYSIOLOGY Pang, L. M., OBRODOVICH, H. M., Mellins, R. B., STALCUP, S. A. 1982; 52 (2): 370-375

    Abstract

    Bradykinin (BK) is a potent edematogenic agent in systemic tissues. It is degraded by angiotensin-converting enzyme (ACE), which is located on the surface of all vascular endothelia. We hypothesized that since oxygen tension modulates ACE activity, the high pulmonary oxygen tension and hence high ACE activity protects the lung from the edematogenic effects of BK. We therefore studied the effect of exogenous BK in unanesthetized sheep with surgically created lymph fistulas and vascular catheters during normoxia and hypoxia. BK significantly elevated lung lymph flow and protein flux only when the sheep were made hypoxic and the lung's ability to degrade BK was impaired. This increase could not be attributed to recruitment of vascular surface area or to an increase in the driving force for fluid exchange because there were no changes in pulmonary arterial or left atrial pressures, cardiac output, or pulmonary vascular resistance. We conclude that BK increases water and protein movement in the lung by increasing vascular permeability.

    View details for Web of Science ID A1982ND15300013

    View details for PubMedID 7037714

  • HEMODYNAMIC AND VASOACTIVE MEDIATOR RESPONSE TO EXPERIMENTAL RESPIRATORY-FAILURE JOURNAL OF APPLIED PHYSIOLOGY OBRODOVICH, H. M., STALCUP, S. A., Pang, L. M., Mellins, R. B. 1982; 52 (5): 1230-1236

    Abstract

    We determined the changes in hemodynamics and circulating concentrations of endogenous vasoactive mediators during acute respiratory failure in chronically catheterized unanesthetized sheep. Inhalation of a hypercarbic-hypoxic gas mixture for 2 h (pHa = 7.16, PaCO2 = 84 Torr, PaO2 = 48 Torr, n = 5) resulted in a doubling of cardiac output (thermodilution) and systemic and pulmonary hypertension. Systemic vascular resistance decreased, whereas pulmonary vascular resistance gradually increased throughout the 2-h experimental gas period. Pulmonary hypertension persisted for 45 min after return to room air breathing. Plasma renin activity tripled, and circulating bradykinin concentration increased 25-fold (radioimmunoassay). Plasma norepinephrine and epinephrine concentrations (radioenzymatic assay) dramatically increased with respective initial values (15 min) of 23.9 +/- 9.5 (SE) and 26.7 +/- 13.3 ng/ml (base line less than 0.2). Inhalation of hypoxic (n = 4) or hypercarbic-enriched O2 (n = 7) gas mixtures did not produce similar findings. We concluded that the hemodynamic response to experimental respiratory failure results from the combination of hypercarbic acidosis and hypoxia. These changes were mediated in part by increased sympathoadrenal activity and altered concentrations of H+, O2, and endogenous vasoactive mediators.

    View details for Web of Science ID A1982NP44200021

    View details for PubMedID 7047470

  • EFFECTS OF GROWTH ON THE DIFFUSION CONSTANT FOR CARBON-MONOXIDE AMERICAN REVIEW OF RESPIRATORY DISEASE OBRODOVICH, H. M., Mellins, R. B., Mansell, A. L. 1982; 125 (6): 670-673

    Abstract

    To investigate factors determining the diffusion constant for carbon monoxide (KCO) in normal children, we used the single breath method in 33 healthy children and young adults, 6 to 30 yr of age. In the sitting position, KCO decreased with increasing height (r = -0.59, p less than 0.001), suggesting greater recruitment of pulmonary vascular bed at the apexes in the shorter subjects. Increase in the KCO after taking the supine position was significantly less in the shorter subjects (8% at 120 cm) than in the taller subjects (27% at 170 cm), confirming this mechanism. Ten subjects with pulmonary involvement from cystic fibrosis had normal supine KCO but did not show a normal decrease in the upright position. From an analysis of present and previous data we conclude that (1) decrease in the conventionally measured KCO during childhood reflects a gravity-dependent decrease in recruitment of the pulmonary vascular bed and a decrease in the ratio of alveolar surface area to alveolar volume during growth and (2) diseases associated with hypoxic pulmonary vasoconstriction tend to recruit the pulmonary vascular bed at the apexes, increasing the conventionally measured KCO.

    View details for Web of Science ID A1982NS89400010

    View details for PubMedID 7091872

  • FAMILIAL LYMPHOID INTERSTITIAL PNEUMONIA - LONG-TERM FOLLOW-UP PEDIATRICS OBRODOVICH, H. M., Moser, M. M., Lu, L. 1980; 65 (3): 523-528

    Abstract

    We describe two brothers who developed chronic pulmonary disease in early childhood. Lung biopsies were diagnostic of lymphoid interstitial pneumonia (LIP). Familial LIP has not been previously reported, and the natural history is unknown. The elder brother experienced progressive respiratory disability and died 10 years after the onset of symptoms. The younger brother age 13, has been observed for 11 years and despite progression of pathological changes revealed in his second lung biopsy, he has had few symptoms and leads an active life. Current pulmonary function tests reveal decreased lung volumes, increased maximal expiratory flow rates and decreased lung compliance. Arterial PO2 is 75 mm Hg at rest and falls to 56 mm Hg with exercise. These findings, consistent with restrictive lung disease, contrast with the obstructive ventilatory pattern seen in some adult patients.

    View details for Web of Science ID A1980JJ82400013

    View details for PubMedID 7360539

  • FIELD STUDIES ON USE OF COATED POROUS POLYURETHANE PLUGS AS INDWELLING MONITORS OF ORGANOCHLORINE PESTICIDES AND POLYCHLORINATED BIPHENYL CONTENTS OF STREAMS ENVIRONMENTAL LETTERS UTHE, J. F., Reinke, J., OBRODOVI, H. 1974; 6 (2): 103-115

    View details for Web of Science ID A1974S407300004

    View details for PubMedID 4206135

  • DETERMINATION OF POLYCHLORINATED BIPHENYLS IN PRESENCE OF ORGANOCHLORINE PESTICIDES BY THIN-LAYER CHROMATOGRAPHY ENVIRONMENTAL LETTERS Reinke, J., UTHE, J. F., OBRODOVI, H. 1973; 4 (3): 201-210

    View details for Web of Science ID A1973O680800003

    View details for PubMedID 4119800

Conference Proceedings


  • Active Na-22(+) transport by the intact lung during early postnatal life OBRODOVICH, H. M., Mullen, B., HANNAM, V. L., Goodman, B. E. NATL RESEARCH COUNCIL CANADA-N R C RESEARCH PRESS. 1997: 431-435

    Abstract

    The lung relies upon epithelial active transport of Na+ to aid in the clearance of fluid from its air spaces. Because it is unknown whether the rate of active Na+ transport by the distal lung epithelium varies during early postnatal age, we performed studies in young guinea pigs (7 and 30 days after birth). We used a single pass isolated perfused lung model in which a Krebs Ringer bicarbonate solution containing 22Na+, [14C]sucrose, and FITC-dextran was placed into the air spaces of the lungs, and apparent permeability-surface area (PS) products were calculated after determining the changes in lung weight and the concentrations of the isotopes in the vascular effluent. The PS product for 22Na+, but not [14C]sucrose, decreased significantly at both ages when amiloride was infused (final concentration of 10(-4) M). Amiloride also decreased the rate of fluid clearance, as assessed by changes in organ weight, at both ages. Although the absolute rate of amiloride-sensitive 22Na+ transport increased with age, morphometric measurement of the alveolar region demonstrated that the rate of amiloride-sensitive 22Na+ transport per unit alveolar surface area was similar. These data indicate that although the guinea pig lung undergoes significant growth shortly after birth, the rate of amiloride-sensitive active Na+ transport per unit surface area remains constant. Since a component of weight loss was insensitive to amiloride, these in vivo studies suggest that the amiloride-insensitive Na+ transport pathways previously identified in cultured lung epithelium exist in the intact lung.

    View details for Web of Science ID A1997XM24600011

    View details for PubMedID 9250377

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