Emeritus Faculty, Acad Council, Pathology
Work in our laboratory is focused on the study of the human major histocompatibility (HLA) complex. We are seeking to characterize the genes and gene products of this complex and to understand their role in determining transplantation and transfusion compatibility. Laboratory work on this topic ranges from clinical studies correlating graft rejection with HLA antigens identified by classical serology or DNA typing. New approaches to enhancing engraftment are also sought using genetic engineering to convert membrane-bound HLA antigens into soluble products for the purpose of inducing tolerance; and by characterization of endothelial cells antigens. Identification and characterization of HLA genes related to disease susceptibility, e.g., narcolepsy, is also under study.
The lack of reliable human proxies for minor (ie, non-HLA) histocompatibility loci hampers the ability to leverage these factors toward improving transplant outcomes. Despite conflicting reports of the effect of donor-recipient sex mismatch on renal allografts, the association between acute rejection of renal allografts and the development of human alloantibodies to the male H-Y antigen suggested to us that donor-recipient sex mismatch deserved re-evaluation.To evaluate whether the relationships between donor sex and allograft failure differed by recipient sex.We studied recipients of deceased-donor (n = 125,369) and living-donor (n = 63,139) transplants in the United States Renal Data System. Using Cox proportional hazards models stratified by donor type, we estimated the association between donor-recipient sex mismatch and death-censored allograft failure with adjustment for known risk factors, with and without the use of multiple imputation methods to account for potential bias and/or loss of efficiency due to missing data.The advantage afforded by male donor kidneys was more pronounced among male than among female recipients (8% vs 2% relative risk reduction; interaction P < 0.01). This difference is of the order of magnitude of several other risk factors affecting donor selection decisions.Donor-recipient sex mismatch affects renal allograft survival in a direction consistent with immune responses to sexually determined minor histocompatibility antigens. Our study provides a paradigm for clinical detection of markers for minor histocompatibility loci.
View details for DOI 10.1016/j.genm.2012.07.004
View details for Web of Science ID 000310170000005
View details for PubMedID 22906727
Transplanted nephron mass is an important determinant of long-term allograft survival, but accurate assessment before organ retrieval is challenging. Newer radiologic imaging techniques allow for better determination of total kidney and cortical volumes.Using volume measurements reconstructed from magnetic resonance or computed tomography imaging from living donor candidates, we characterized total kidney (n=312) and cortical volumes (n=236) according to sex, age, weight, height, body mass index (BMI), and body surface area (BSA).The mean cortical volume was 204 mL (range 105-355 mL) with no significant differences between left and right cortical volumes. The degree to which existing anthropomorphic surrogates predict nephron mass was quantified, and a diligent attempt was made to derive a better surrogate model for nephron mass. Cortical volumes were strongly associated with sex and BSA, but not with weight, height, or BMI. Four prediction models for cortical volume constructed using combinations of age, sex, race, weight, and height were compared with models including either BSA or BMI.Among existing surrogate measures, BSA was superior to BMI in predicting renal cortical volume. We were able to construct a statistically superior proxy for cortical volume, but whether relevant improvements in predictive accuracy could be gained needs further evaluation in a larger population.
View details for DOI 10.1097/TP.0b013e31823705ef
View details for Web of Science ID 000298149200012
View details for PubMedID 22011765
Pre-transplant screening of a woman with end-stage renal disease (ESRD) showed no anti-human leukocyte antigen (HLA) alloantibodies by anti-human globulin-complement-dependent cytotoxicity (AHG-CDC; class I) or enzyme-linked immunosorbent assay (class II). Following a negative AHG-CDC crossmatch, an HLA*01:01+ deceased donor (DD) kidney was transplanted in September 2005. Subsequent screening of pre-transplant serum by LABScreen Single Antigen (SA) array showed strong reactivity versus A*01:01. Despite that reactivity, at 5 years post-transplant, the patient has a serum creatinine of 1.6 mg/dl and has never experienced humoral or cellular rejection. Retrospective flow-cytometric crossmatch of pre- and post-transplant sera versus DD cells was negative. Rescreening of multiple pre- and post-transplant sera revealed anti-A1 reactivity persisting from the first through the last samples tested. The patient's anti-A1 was almost two fold more reactive with denatured A*01:01 FlowPRA SA beads after denaturation with acid treatment (pH 2.7) than with untreated beads. Parallel results were observed with pH 2.7 treated versus untreated A1+ T cells in FXM. These data highlight the difficulty in interpreting screening results obtained using bead arrays, because of antibodies that appear to recognize denatured but not native class I HLA antigens. We suggest that such bead-positive, flow cytometric crossmatch negative antibodies are not associated with humoral rejection, may not necessarily be detrimental to a graft, and deserve further evaluation before becoming a barrier to transplantation.
View details for DOI 10.1016/j.humimm.2011.02.012
View details for Web of Science ID 000291138900005
View details for PubMedID 21396421
Clonal chromosomal abnormalities are often found in the tumor cells of patients with malignancies. These abnormalities can cause downregulation of human leukocyte antigen (HLA) and instability of short tandem repeat (STR) DNA sequences, confounding HLA typing and/or engraftment analysis in hematopoietic stem cell transplants (HSCT). We describe here the abnormalities observed during testing of 600 HSCT patients. HLA molecular typing was performed by reference strand conformational analyses and/or sequence-based typing. STR testing was performed with 10 to 16 STR primer sets, following 1 to 4 informative loci in each patient. Eight patients exhibited either loss of heterozygosity (4 STR, 3 HLA) or STR length mutation (n = 1), and 5 of the 8 exhibited correlative cytogenetic abnormalities. Diagnoses were acute myelogenous leukemia (AML; n = 7) or myelofibrosis (MFIB: n = 1), yielding an 11% incidence of these chromosomal abnormalities among the subset of 72 AML/MFIB HSCT patients. These results highlight some of the problems encountered and the possibility for interpretive errors that can arise when analyzing molecular typing and engraftment data, particularly among AML/MFIB patients.
View details for DOI 10.1016/j.humimm.2011.03.003
View details for Web of Science ID 000291138900007
View details for PubMedID 21463659
Human minor histocompatibility antigens (mHA) and clinically relevant immune responses to them have not been well defined in organ transplantation. We hypothesized that women with male kidney transplants would develop antibodies against H-Y, the mHA encoded on the Y-chromosome, in association with graft rejection.We tested sera from 118 consecutive transplant recipients with kidney biopsies. Antibodies that specifically recognized the recombinant H-Y antigens RPS4Y1 or DDX3Y were detected by IgG enzyme-linked immunosorbent assay and western blotting. Immunogenic epitopes were further identified using overlapping H-Y antigen peptides for both the H-Y proteins.In the 26 female recipients of male kidneys, H-Y antibody development posttransplant (1) was more frequent (46%) than in other gender combinations (P<0.001), (2) showed strong correlation with acute rejection (P=0.00048), (3) correlated with plasma cell infiltrates in biopsied kidneys (P=0.04), and (4) did not correlate with C4d deposition or donor-specific anti-human leukocyte antigen (HLA) antibodies. Of the two H-Y antigens, RPS4Y1 was more frequently recognized (P=0.005).This first demonstration of a strong association between H-Y antibody development and acute rejection in kidney transplant recipients shows that in solid organ allografts, humoral immune responses against well defined mHA have clear clinical correlates, can be easily monitored, and warrant study for possible effects on long-term graft function.
View details for DOI 10.1097/TP.0b013e31817352b9
View details for Web of Science ID 000257790400014
View details for PubMedID 18622281
The association of narcolepsy with HLA-DQB1*0602 is established in Japanese, African-Americans, European, and North American Caucasians. We examined DRB1, DRB3, DRB4, DRB5, DQA1, and DQB1 in 163 patients with centrally mediated daytime sleepiness (100 with narcolepsy) and 211 Korean controls. In this population, the DQB1*0602 association was always evident in the context of the DRB1*1501-DQA1*0102-DQB1*0602 haplotype. The DQB1*0602 association was highest in cases with hypocretin deficiency (100% vs 13% in controls), most of which had narcolepsy-cataplexy (81%). A weaker DQB1*0602 (45%) association was present in cases without cataplexy. No human leukocyte antigen (HLA) association was present in idiopathic hypersomnia or in cases with normal cerebrospinal fluid (CSF) hypocretin-1. As in other populations, DQB1*0602 homozygosity increased risk in cases with cataplexy and/or hypocretin deficiency (odds ratio = 2.0 vs heterozygotes). Non-DQB1*0602 allelic effects were also observed but could not be interpreted in the context of DQB1*0602 overabundance and linkage disequilibrium. We therefore next analyzed compound heterozygote effects in 77 subjects with either hypocretin deficiency or cataplexy and one copy of DRB1*1501-DQA1*0102-DQB1*0602, a sample constructed to maximize etiologic homogeneity. In this analysis, we found additional predisposing effects of DQB1*0301 and protective effects for DQA1*0103-DQB1*0601. Unexpectedly, the predisposing effects of DQB1*0301 were present in the context of various DQA1-bearing haplotypes. A predisposing effect of DQA1*0303 was also suggested. These results indicate a remarkable consistency in the complex HLA association present in narcolepsy across multiple ethnic groups.
View details for DOI 10.1016/j.humimm.2006.10.006
View details for Web of Science ID 000243570200007
View details for PubMedID 17207713
TRALI is a challenging diagnosis for both the transfusion specialist and the clinician. A Canadian consensus panel has recently proposed guidelines to better define TRALI and its implications. The guidelines recommend classifying each suspected case in one of the following 3 categories: (1) "TRALI," (2) "Possible TRALI," or (3) "Not TRALI." We report the clinical presentation, laboratory evaluation, and management of 3 patients with respiratory failure (RF) following allogeneic blood transfusions. These patients all experienced RF within 6 hr post-transfusion. Based on a review of the clinical and laboratory data and applying the Canadian guidelines, the first patient, a 67-yr-old man with chronic myelomonocytic leukemia, was diagnosed as "TRALI" due to the sudden onset of RF requiring intensive resuscitation. The second patient, a 55-yr-old man with aplastic anemia, was diagnosed as "Possible TRALI" due to pre-existing RF that worsened after blood transfusion. The third patient, a 1-yr-old male, was diagnosed as transfusion associated circulatory overload (TACO) and "Possible TRALI," although his RF improved after treatment with diuretics. In all 3 cases, the blood donor center was informed of the suspected TRALI reactions. The remaining blood products from the donors associated with these reactions were quarantined. After review of the clinical data, the donors associated with cases #1 and #3 were screened by the blood center for granulocyte and HLA antibodies. Using a Luminex flow bead array, the following class I and class II antibodies specific for patient #1 were identified in the respective donor: anti-A25, B8, B18, and anti-DR15, DR 17. Subsequently, donor #1 was permanently deferred. A non-specific IgM anti-granulocyte antibody was identified in the donor associated with case #3, and this donor was subsequently disqualified from plasma and platelet donations. In conclusion, the Canadian guidelines to categorize patients suspected of TRALI provide a useful framework for evaluation of these patients and their respective blood donors.
View details for Web of Science ID 000236733400008
View details for PubMedID 16501237
Conditioning with total lymphoid irradiation plus antithymocyte serum protects mice against acute graft-versus-host disease (GVHD) after hematopoietic-cell transplantation. We tested this strategy in humans.Thirty-seven patients with lymphoid malignant diseases or acute leukemia underwent an experimental conditioning regimen with 10 doses of total lymphoid irradiation (80 cGy each) plus antithymocyte globulin, followed by an infusion of HLA-matched peripheral-blood mononuclear cells from related or unrelated donors who received granulocyte colony-stimulating factor.Of the 37 transplant recipients, only 2 had acute GVHD after hematopoietic-cell transplantation. Potent antitumor effects in patients with lymphoid malignant diseases were shown by the change from partial to complete remission. In the transplant recipients who underwent conditioning with total lymphoid irradiation and antithymocyte globulin, the fraction of donor CD4+ T cells that produced interleukin-4 after in vitro stimulation increased by a factor of five, and the proliferative response to alloantigens in vitro was reduced, as compared with normal control subjects and control subjects who underwent conditioning with a single dose of total-body irradiation (200 cGy).A regimen of total lymphoid irradiation plus antithymocyte globulin decreases the incidence of acute GVHD and allows graft antitumor activity in patients with lymphoid malignant diseases or acute leukemia treated with hematopoietic-cell transplantation.
View details for Web of Science ID 000232146200004
View details for PubMedID 16192477
Strategies to induce donor-specific allograft tolerance are best tested in preclinical models developed in nonhuman primates (NHPs). Most protocols prepare the recipient by infusing hematopoietic cells from the donor. We report here a procedure to isolate and characterize large numbers of bone marrow cells (BMCs) from cynomolgus monkeys (cynos) that can then successfully be transplanted into conditioned recipients.Vertebral columns of five cynos were excised en bloc and separated into individual vertebrae. The cancelous bone was extracted with a core puncher, fractionated, filtered, centrifuged, and resuspended in transplantation media before being analyzed by flow cytometry. In two instances, the collected BMCs were reinfused into allogeneic recipients preconditioned with a nonmyeloablative regimen. Chimerism was monitored using short-tandem repeat analysis.The mean total BMCs yield was 25.5 x 10(9) (range of 4.00 x 10(9) to 59 x 10(9)) with mean cell viability of 93.4% (range: 90-96%). CD34+ cells and CD3+ cells averaged 0.34 and 3.91% of total BMCs, respectively. This resulted in absolute cell number yields of 1.02 x 10(8) and 1.15 x 10(9) for CD34+ and CD3+ cells, respectively. Graft-versus-host disease was absent in both bone marrow infused animals, and a maximum level of chimerism of 18% was detected at 3 weeks after BMCs infusion.We present here the first detailed report of a procedure to retrieve and characterize large numbers of BMCs from vertebral bodies of cynos and demonstrate that cells collected with this technique have the capability of engrafting in allogenic recipients.
View details for DOI 10.1016/j.jss.2004.09.018
View details for Web of Science ID 000228275800018
View details for PubMedID 15820259
The influence of graft composition on clinical outcomes after reduced-intensity conditioning is not well-characterized. In this report we prospectively enumerated CD34+, CD3+, CD4+, and CD8+ cell doses in granulocyte colony-stimulating factor-mobilized peripheral blood mononuclear cell (G-PBMC) allografts in 63 patients who received transplants following non-myeloablative conditioning with total body irradiation 200 cGy plus fludarabine as treatment for malignant diseases. Donors were HLA-identical siblings (n = 38) or HLA-matched unrelated individuals (n = 25). By univariate analyses G-PBMC CD8+ T-cell dose in at least the 50th percentile favorably correlated with full donor blood T-cell chimerism (P = .03), freedom from progression (P = .001), and overall survival (P = .01). No G-PBMC cell dose influenced grade II to IV acute or extensive chronic graft-versus-host disease. In multivariate analysis only G-PBMC CD8+ T-cell dose (P = .003; RR = 0.2, 95% CI = 0.1-0.6) was associated with improved freedom from progression. Infusion of low G-PBMC CD8+ T-cell dose for reduced-intensity allografting may adversely affect T-cell engraftment and survival outcome.
View details for DOI 10.1182/blood-2004-04-1473
View details for Web of Science ID 000227630500020
View details for PubMedID 15572597
We analyzed the kinetics of donor engraftment among various peripheral blood cell subpopulations and their relationship to outcomes among 120 patients with hematologic malignancies given hematopoietic cell transplantation (HCT) after nonmyeloablative conditioning consisting of 2 Gy total body irradiation (TBI) with or without added fludarabine. While patients rapidly developed high degrees of donor engraftment, most remained mixed donor/host chimeras for up to 180 days after HCT. Patients given preceding chemotherapies and those given granulocyte colony-stimulating factor-mobilized peripheral blood mononuclear cell (G-PBMC) grafts had the highest degrees of donor chimerism. Low donor T-cell (P = .003) and natural killer (NK) cell (P = .004) chimerism levels on day 14 were associated with increased probabilities of graft rejection. High T-cell chimerism on day 28 was associated with an increased probability of acute graft-versus-host disease (GVHD) (P = .02). Of 93 patients with measurable malignant disease at transplantation, 41 achieved complete remissions a median of 199 days after HCT; 19 of the 41 were mixed T-cell chimeras when complete remissions were achieved. Earlier establishment of donor NK-cell chimerism was associated with improved progression-free survival (P = .02). Measuring the levels of peripheral blood cell subset donor chimerisms provided useful information on HCT outcomes and might allow early therapeutic interventions to prevent graft rejection or disease progression.
View details for Web of Science ID 000224378300015
View details for PubMedID 15226174
Chimerism assessment following bone marrow transplantation (BMT) in cynomolgus monkeys (cynos) has been hampered by the lack of good engraftment markers. In human BMT, such markers have been provided by short tandem repeat (STR) loci. We tested the idea that techniques effective for detecting human STR could be readily adapted to cynos. Genomic DNA was extracted from cyno unseparated blood or peripheral cell subsets. With only slight modifications, reagents for detecting human STR alleles were used to amplify and detect cyno STRs and to quantitate allelic mixtures on an automated sequencer. Of the 15 STR loci tested, only CSF1PO, D18S51, and FGA successfully amplified, with seven, seven and two alleles, respectively. CSF1PO and D18S51 heterozygosity (80% and 55%, respectively) allowed use of these two loci for chimerism quantitation after BMT. The successful adaptation of human STR reagents to monitor chimerism in transplanted cynos will facilitate the use of this species in preclinical tolerance studies.
View details for DOI 10.1111/j.1600-6143.2004.00529.x
View details for Web of Science ID 000223283900021
View details for PubMedID 15307845
MHC typing for human hematopoietic cell transplantation (HCT) from unrelated donors is currently performed by using a combination of serologic and molecular techniques. It has been determined that allelic differences in human MHC molecules, revealed by nucleotide sequencing but not by serologic typing, substantially influence graft rejection and graft-versus-host disease, two serious complications of clinical HCT. We studied transplantation of purified hematopoietic stem cells in a series of mouse strains that were matched at the MHC but had different background genes, and we observed striking differences in engraftment resistance and graft-versus-host disease severity, both factors depending on the donor-recipient strain combination. The individual mouse lines studied here were established nearly a century ago, and their MHC types were determined exclusively by serologic techniques. We considered the possibility that serologically silent MHC polymorphisms could account for our observations and, therefore, we performed DNA sequencing of the class I and II MHC alleles of our mouse strains. At each locus, exact homology was found between serologically MHC-matched strains. Our results likely extend to all serologically MHC-matched mouse strains used in modern research and highlight the profound and variable influence that non-MHC genetic determinants can have in dictating outcome after HCT.
View details for DOI 10.1073/pnas.2035077100
View details for Web of Science ID 000185685700072
View details for PubMedID 14504392
Rodents and dogs conditioned with total-lymphoid irradiation (TLI), with or without antithymocyte globulin (ATG), have been shown to develop mixed chimerism and immune tolerance without graft-versus-host disease (GVHD) after the infusion of major histocompatability complex (MHC)-mismatched donor bone marrow cells given alone or in combination with an organ allograft.Four human leukocyte antigen (HLA)-mismatched recipients of living donor kidney transplants were conditioned with TLI and ATG posttransplantation and infused with cyropreserved donor granulocyte colony-stimulating factor (G-CSF) "mobilized" hematopoietic progenitor (CD34+) cells (3-5x10(6) cells/kg) thereafter. Maintenance prednisone and cyclosporine dosages were tapered, and recipients were monitored for chimerism, GVHD, graft function, T-cell subsets in the blood, and antidonor reactivity in the mixed leukocyte reaction (MLR).Three of the four patients achieved multilineage macrochimerism, with up to 16% of donor-type cells among blood mononuclear cells without evidence of GVHD. Prolonged depletion of CD4+ T cells was observed in all four patients. Rejection episodes were not observed in the three macrochimeric recipients, and immunosuppressive drugs were withdrawn in the first patient by 12 months. Prednisone was withdrawn from a second patient at 9 months, and cyclosporine was tapered thereafter.Multilineage macrochimerism can be achieved without GVHD in HLA-mismatched recipients of combined kidney and hematopoietic progenitor transplants. Conditioning of the host with posttransplant TLI and ATG was nonmyeloablative and was not associated with severe infections. Recipients continue to be studied for the development of immune tolerance.
View details for Web of Science ID 000175933100002
View details for PubMedID 12023614
Regeneration of hematopoiesis after allogeneic hematopoietic cell transplantation (HCT) involves conversion of the recipient's immune system to donor type. It is likely that distinct cell lineages in the recipient reconstitute at different rates. Dendritic cells (DCs) are a subset of hematopoietic cells that function as a critical component of antigen-specific immune responses because they modulate T-cell activation, as well as induction of tolerance. Mature DCs are transferred with hematopoietic grafts and subsequently arise de novo. Little information exists about engraftment kinetics and turnover of this cell population in patients after allogeneic HCT. This study examined the kinetics of DC chimerism in patients who underwent matched sibling allogeneic HCT. T-cell, B-cell, and myelocytic and monocytic chimerism were also studied. Peripheral blood cells were analyzed at defined intervals after transplantation from 19 patients with various hematologic malignancies after treatment with myeloablative or nonmyeloablative preparatory regimens. Cell subsets were isolated before analysis of chimerism. Despite the heterogeneity of the patient population and preparatory regimens, all showed rapid and consistent development of DC chimerism. By day +14 after transplantation approximately 80% of DCs were of donor origin with steady increase to more than 95% by day +56. Earlier time points were examined in a subgroup of patients who had undergone nonmyeloablative conditioning and transplantation. These data suggest that a major proportion of blood DCs early after transplantation is donor-derived and that donor chimerism develops rapidly. This information has potential implications for manipulation of immune responses after allogeneic HCT.
View details for Web of Science ID 000173787600049
View details for PubMedID 11830498
Blastocyst transfer of just one or two embryos has been used to help limit the number of high-order gestations. In this case report we describe the occurrence of a quadruplet pregnancy after the transfer of only two blastocysts during IVF. Sonographic examination showed four fetuses and what appeared to be quadriamniotic/quadrichorionic sacs, suggesting that a concomitant spontaneous conception had occurred. Definite confirmation of zygosity was obtained by genetic testing using DNA microsatellite polymorphism determinations after the birth of one boy and three girls at 32 weeks gestation. Although this event has not been reported previously, the possibility of its occurrence should be kept in mind. IVF patients with patent Fallopian tubes should be cautioned against intercourse late in their controlled ovarian stimulation, especially if they would decline multifetal reduction.
View details for Web of Science ID 000172097200017
View details for PubMedID 11679513
Toxicities have limited the use of allogeneic hematopoietic cell transplantation (HCT) to younger, medically fit patients. In a canine HCT model, a combination of postgrafting mycophenolate mofetil (MMF) and cyclosporine (CSP) allowed stable allogeneic engraftment after minimally toxic conditioning with low-dose (200 cGy) total-body irradiation (TBI). These findings, together with the known antitumor effects of donor leukocyte infusions (DLIs), led to the design of this trial. Forty-five patients (median age 56 years) with hematologic malignancies, HLA-identical sibling donors, and relative contraindications to conventional HCT were treated. Immunosuppression involved TBI of 200 cGy before and CSP/MMF after HCT. DLIs were given after HCT for persistent malignancy, mixed chimerism, or both. Regimen toxicities and myelosuppression were mild, allowing 53% of eligible patients to have entirely outpatient transplantations. Nonfatal graft rejection occurred in 20% of patients. Grades II to III acute graft-versus-host disease (GVHD) occurred in 47% of patients with sustained engraftment. With median follow-up of 417 days, survival was 66.7%, nonrelapse mortality 6.7%, and relapse mortality 26.7%. Fifty-three percent of patients with sustained engraftment were in complete remission, including 8 with molecular remissions. This novel allografting approach, based on the use of postgrafting immunosuppression to control graft rejection and GVHD, has dramatically reduced the acute toxicities of allografting. HCT with the induction of potent graft-versus-tumor effects can be performed in previously ineligible patients, largely in an outpatient setting. Future protocol modifications should reduce rejection and GVHD, thereby facilitating studies of allogeneic immunotherapy for a variety of malignancies. (Blood. 2001;97:3390-3400)
View details for Web of Science ID 000168927900011
View details for PubMedID 11369628
Human narcolepsy-cataplexy, a sleep disorder associated with a centrally mediated hypocretin (orexin) deficiency, is tightly associated with HLA-DQB1*0602. Few studies have investigated the influence that additional HLA class II alleles have on susceptibility to this disease. In this work, 1,087 control subjects and 420 narcoleptic subjects with cataplexy, from three ethnic groups, were HLA typed, and the effects of HLA-DRB1, -DQA1, and -DQB1 were analyzed. As reported elsewhere, almost all narcoleptic subjects were positive for both HLA-DQA1*0102 and -DQB1*0602. A strong predisposing effect was observed in DQB1*0602 homozygotes, across all ethnic groups. Relative risks for narcolepsy were next calculated for heterozygous DQB1*0602/other HLA class II allelic combinations. Nine HLA class II alleles carried in trans with DQB1*0602 were found to influence disease predisposition. Significantly higher relative risks were observed for heterozygote combinations including DQB1*0301, DQA1*06, DRB1*04, DRB1*08, DRB1*11, and DRB1*12. Three alleles-DQB1*0601, DQB1*0501, and DQA1*01 (non-DQA1*0102)-were found to be protective. The genetic contribution of HLA-DQ to narcolepsy susceptibility was also estimated by use of lambda statistics. Results indicate that complex HLA-DR and -DQ interactions contribute to the genetic predisposition to human narcolepsy but that additional susceptibility loci are also most likely involved. Together with the recent hypocretin discoveries, these findings are consistent with an immunologically mediated destruction of hypocretin-containing cells in human narcolepsy-cataplexy.
View details for Web of Science ID 000166994200013
View details for PubMedID 11179016
Graft-versus-host disease (GVHD) complicating allogeneic bone marrow transplantation (BMT) is often attributed to mismatched minor histocompatibility antigens (mHags), which are poorly defined in humans. CD31 is a candidate human mHag relevant to acute GVHD, but reports disagree about its level of significance, the role of HLA restriction, and the relative importance of different polymorphic codons within the molecule. We therefore examined in greater detail the impact of CD31-matching on BMT outcome in a prospective study from a single institution. Samples of recipient and donor DNA were collected pretransplantation for all patients receiving unmanipulated bone marrow from an HLA-identical sibling over a 45-month period at our institution. CD31 DNA typing of alleles at the 3 polymorphic codons 125 (L or V), 563 (N or S), and 670 (R or G) was performed for 118 patient-donor pairs plus 2 additional pairs who had codon 125 typing only. Donor-recipient CD31 nonidentity was tested for correlation with BMT clinical outcome measures of severe acute GVHD, chronic GVHD, relapse, and survival. Gene frequencies of approximately 0.5 for each allele at all 3 codons were comparable to previous reports. Because complete association was seen for 563N with 670G and for 563S with 670R, nonidentity for those codons was analyzed as a single genetic marker designated codon 563/670. Donor-recipient CD31 nonidentity was a significant risk factor for overall survival, both at codon 563/670 (hazard ratio [hr] = 2.58, P = .005) and at codon 125 (hr = 1.07, P = .036). Similar results held for disease-free survival. Nonidentity at codon 563/670 was also a significant risk factor (odds ratio [OR] = 11.15, P = .011) for severe (grades III, IV) versus no (grade 0) acute GVHD. Nonidentity at codon 125 posed less but still significant risk (OR = 9.30, P = .030). When the comparison group without severe acute GVHD was expanded to include grade I as well as grade 0 patients, the risk from CD31 nonidentity increased for both codon 563/670 (OR = 12.31, P = .010) and codon 125 (OR = 11.24, P = .011). CD31 nonidentity remained a significant independent risk factor for survival and for severe acute GVHD when tested in multivariate analysis with the covariates of adulthood, recipient-donor sex difference, ethnic group, disease, pretransplantation risk category, HLA-A2 type, B44-like types, and GVHD prophylactic regimen. CD31 nonidentity showed a trend but failed to achieve statistical significance as a risk factor for relapse and for chronic GVHD. In conclusion, donor-recipient CD31 nonidentity is a significant risk factor for survival and for severe acute GVHD in HLA-identical sibling BMT. The stronger associations with codon 563/670 suggest that polymorphism may be more important than the linked polymorphism at codon 125.
View details for Web of Science ID 000171449200004
View details for PubMedID 11669217
Previous studies showed the feasibility of inducing transplantation tolerance to cadaveric renal allografts in patients given pretransplant total lymphoid irradiation (TLI). Microchimerism has been theorized to be an important or necessary factor in long-term graft acceptance and tolerance in humans.A cadaveric renal transplant recipient given pretransplant total lymphoid irradiation and withdrawn from immunosuppressive drugs more than 12 years ago was tested for microchimerism using a sensitive nested polymerase chain reaction technique, and for anti-donor reactivity using the mixed leukocyte reaction and an ELISA screen for anti-HLA antibodies. Donor and recipient were mismatched for all HLA-A, B, and DR antigens.The "tolerant" recipient had good graft function, no detectable donor-type cells in the blood by polymerase chain reaction analysis, vigorous reactivity to donor stimulator cells in the mixed leukocyte reaction, and no detectable serum anti-HLA antibodies.Operational tolerance to HLA-A, B, and DR mismatched organ allografts can be induced prospectively in humans for at least 12 years after withdrawal of immunosuppressive drugs. The allograft can be maintained in the absence of detectable donor microchimerism and in the presence of anti-donor reactivity in the mixed leukocyte reaction, suggesting that neither chimerism nor clonal deletion or anergy of recipient T cells to alloantigens presented by donor Class II HLA molecules is required for persistence of the tolerant state using this total lymphoid irradiation protocol.
View details for Web of Science ID 000086910700005
View details for PubMedID 10836360
View details for PubMedID 10681265
Associations between Human Leukocyte Antigen (HLA) (i.e. human major histocompatibility complex [MHC]) genes and susceptibility to infections and inflammatory processes have been described, but causal relationships have not been proven. We characterized effects of HLA-DQ alleles on outcome of congenital toxoplasma infection and found that among Caucasians, the DQ3 gene frequency was significantly higher in infected infants with hydrocephalus (0.783) than infected infants without hydrocephalus (0.444) or published normal controls (0.487). We then developed a novel animal model to definitively determine the effect of these HLA DQ molecules on the severity of toxoplasmosis. Human MHC-Class II transgenes reduced parasite burden and necrosis in brains of mice infected with Toxoplasma gondii. Consistent with the observed association between DQ3 and hydrocephalus in human infants, in the murine model the DQ3(DQ8; DQB1*0302) gene protected less than DQ1 (DQ6; DQB1*0601). Our findings definitively prove a cause and effect relationship between human MHC genes and resistance to infection, provide novel means to characterise human immune responses that are protective or pathogenic in infections, and are important for vaccine development.
View details for Web of Science ID 000083484200001
View details for PubMedID 10579423
An index patient with pseudohomozygosity for factor V Leiden was identified. Each of his two children inherited a different paternal factor V allele; a daughter was heterozygous for factor V Leiden, with 100% factor V activity, and a son was heterozygous for factor V deficiency, with 50% factor V activity. Genomic DNA was obtained from family members, and the 25 factor V exons and flanking intronic regions were sequenced in the proband and confirmed in the children. Within exon 13 of factor V, a 4 base insertion was found at NT 2856 in the proband and son. but not the daughter. This mutation, here designated factor V Stanford, results in a frameshift with loss of a thrombin activation site (R1545V) and premature termination of translation at amino acid 1560.
View details for Web of Science ID 000082421100018
View details for PubMedID 10494770
We studied cadaver kidney transplant recipients to determine if their serum levels of donor-specific class I sHLA correlated with graft outcome. Testing of sHLA was performed by an ELISA sandwich assay using allospecific monoclonal trapping antibodies and anti-beta2-mu detecting antibody. Sufficient sHLA sensitivity (<1 ng/ml) was achieved by using two synergistic trapping antibodies. Suitable antibodies were available for A2 and B7, and data were collected for these two antigens. Stability of these sHLA was determined in plasma and serum as were ranges of normal and background levels. Background levels varied substantially. Five A2- recipients of A2+ grafts and 5 B7- recipients of B7+ grafts were studied with appropriate sHLA levels measured pre-transplant and at intervals post-transplant. Graft outcome was assessed by serum creatinines, renal biopsies and/or therapy for rejection. In the 5 patients (3 A2- and 2 B7-) whose post-transplant donor-specific sHLA never exceeded immunological complications (e.g., post-operative ATN, ureteral obstruction) did not affect the correlation. In the 5 patients with post-transplant levels exceeding pre-transplant levels, subsequent evidence of rejection was observed. Periodic measurement of donor-specific sHLA should be a useful instrument for monitoring renal allograft rejection.
View details for Web of Science ID 000080730100008
View details for PubMedID 10447402
Narcolepsy is a neurological disorder known to be tightly associated with HLA-DQA1*0102 and DQB1*0602. In this study, we have examined if homozygosity for DQB1*0602 increases disease susceptibility and/or severity. Patients diagnosed at Stanford University (n=160) or enrolled in a multicenter clinical trial (n=509) were included in this analysis. In both African-Americans and Caucasian-Americans with or without cataplexy, a significantly higher than expected number of subjects were DQB1*0602 homozygotes. Relative risks were 2-4 times higher in DQB1*0602 homozygotes vs heterozygotes across all patient groups. In contrast, symptom severity did not differ between DQB1*0602 homozygous and heterozygous subjects. These results indicate that HLA-DQB1*0602 homozygosity increases susceptibility to narcolepsy but does not appear to influence disease severity.
View details for Web of Science ID 000071134000012
View details for PubMedID 9459509
Narcolepsy is considered a homogeneous clinical entity when excessive daytime sleepiness and cataplexy are present. Cataplexy is a polymorphic symptom that can be very mild and is thus subjectively defined. The Multiple Sleep Latency Test (MSLT) is widely used as a diagnostic test for narcolepsy. A short mean sleep latency and multiple sleep onset REM periods (SOREMPs) are typically observed in narcoleptic patients. The discovery of a tight association of narcolepsy with HLA class II antigens offers a unique opportunity to explore the respective value of the MSLT or of the presence of clear-cut cataplexy in defining an etiologically homogeneous group of narcoleptic patients. In this study, we carried out HLA typing for DR15(DR2) and DQB1*0602 in 188 narcoleptic patients with cataplexy in three ethnic groups (24 Asians, 61 Blacks, and 103 Caucasians). These results confirm the importance of DQB1*0602 typing rather than DR15 (DR2) typing in Black narcoleptic patients and demonstrate that the presence of clear-cut cataplexy is a better predictor for DQB1*0602 positivity than the presence of abnormal MSLT results.
View details for Web of Science ID A1997XE09100015
View details for PubMedID 9191765
Narcolepsy is a sleep disorder that has been shown to be tightly associated with HLA DR15 (DR2). In this study, 58 non-DR15 patients with narcolepsy-cataplexy were typed at the HLA DRB1, DQA1 and DQB1 loci. Subjects included both sporadic cases and narcoleptic probands from multiplex families. Additional markers studied in the class II region were the promoters of the DQA1 and DQB1 genes, two CA repeat polymorphisms (DQCAR and DQCARII) located between the DQA1 and DQB1 genes, three CA repeat markers (G51152, T16CAR and G411624R) located between DQB1 and DQB3 and polymorphisms at the DQB2 locus. Twenty-one (36%) of these 58 non-DR15 narcoleptic patients were DQA1*0102 and DQB1*0602, a DQ1 subtype normally associated with DRB1*15 in DR2-positive narcoleptic subjects. Additional microsatellite and DQA1 promoter diversity was found in some of these non-DR15 but DQB1*0602-positive haplotypes but the known allele specific codons of DQA1*0102 and DQB1*0602 were maintained in all 21 cases. The 37 non-DQA1*0102/DQB1*0602 subjects did not share any particular HLA DR or DQ alleles. We conclude that HLA DQA1*0102 and DQB1*0602 are the most likely primary candidate susceptibility genes for narcolepsy in the HLA class II region.
View details for Web of Science ID A1997WV41100002
View details for PubMedID 9151385
View details for Web of Science ID 000071716700074
Narcolepsy is a sleep disorder associated with HLA DR15 (DR2) and DQB1*0602. We HLA typed 509 patients enrolled in a clinical trial for the drug modafinil and analyzed the results in relation to cataplexy, a symptom of narcolepsy characterized by muscle weakness triggered by emotions. The patients were either subjects with cataplexy who had a mean sleep latency (SL) of less than 8 minutes and two or more sleep onset rapid eye movement (REM) periods (SOREMPs) during a multiple sleep latency test, or narcoleptic patients without cataplexy but with a mean SL shorter than 5 minutes and two or more SOREMPs. The respective values of DRB1*15 (DR2) and DQB1*0602 as markers for narcolepsy were first compared in different ethnic groups and in patients with and without cataplexy. DQB1*0602 was found to be a more sensitive marker for narcolepsy than DRB1*15 across all ethnic groups. DQB1*0602 frequency was strikingly higher in patients with cataplexy versus patients without cataplexy (76.1% in 421 patients versus 40.9% in 88 patients). Positivity was highest in patients with severe cataplexy (94.8%) and progressively decreased to 54.2% in patients with the mildest cataplexy. A voluntary 50-item questionnaire focusing on cataplexy was also analyzed in 212 of the 509 HLA-typed patients. Subjects with definite cataplexy as observed by an experienced clinician were more frequently HLA DQB1*0602-positive than those with doubtful cataplexy, and the manifestations of cataplexy were clinically more typical in DQB1*0602-positive patients. These results show that the HLA association is as tight as previously reported (85-95%) when cataplexy is clinically typical or severe. We also found that patients with mild, atypical, or no cataplexy have a significantly increased DQB1*0602 frequency (40-60%) in comparison with ethnically matched controls (24%). These results could be explained by increased disease heterogeneity in the noncataplexy group or by a direct effect of the HLA DQB1*0602 genotype on the clinical expression of narcolepsy.
View details for Web of Science ID 000071355600011
View details for PubMedID 9456467
Graft-versus-host disease (GVHD) caused by poorly defined minor (i.e., other than HLA) histocompatibility antigens remains a serious problem in recipients of bone marrow transplants. We sought to determine whether the CD31 adhesion molecule is a minor alloantigen.We directly sequenced samples of complementary DNA (cDNA) encoding CD31 molecules from 21 unrelated normal subjects. Sequence-specific primers were then designed to amplify alleles by the polymerase chain reaction, thereby permitting CD31 typing of genomic DNA from additional normal subjects. To assess the relevance of CD31 matching to bone marrow transplantation, we performed CD31 typing of 46 recipients of bone marrow (32 without GVHD and 14 with severe [grade III or IV] acute GVHD) and their HLA-identical sibling donors. The immunoreactivity of CD31 phenotypes with anti-CD31 monoclonal antibodies was compared by flow cytometry.Direct sequencing of cDNA for CD31 from the 21 normal subjects identified a single polymorphism, CTG-->GTG (Leu-->Val), at codon 125; we designated the resulting alleles CD31.L and CD31.V, respectively. The CD31 genotypes of these and 142 other unrelated subjects were of the expected frequencies. Among the transplant recipients, 71 percent of those with acute GVHD had CD31 genotypes that were not identical to the donor's genotype, as compared with 22 percent of the recipients without GVHD (P = 0.004). The binding of anti-CD31 monoclonal antibodies as measured by fluorescence-activated cell sorting correlated with the CD31 types of homozygous cell lines.The adhesion molecule CD31 is polymorphic. When donor and recipient genotypes are not identical, the risk of GVHD increases. Prospective CD31 typing may reduce the risk of acute GVHD.
View details for Web of Science ID A1996TV69500002
View details for PubMedID 8532023
The frequency of HLA-DQ antigens in AIDS patients with toxoplasmic encephalitis (TE) were examined. HLA-DQ3 was significantly more frequent in white North American AIDS patients with TE (85.0%) than in the general white population (51.8%; P = .007, corrected P = .028) or randomly selected control AIDS patients who had not developed TE (40.0%; P = .016). In contrast, the frequency of HLA-DQ1 was lower in TE patients than in healthy controls (40.0% vs. 66.5%, P = .027), but this difference did not reach statistical significance when corrected for the number of variables tested (corrected P = .108 for the general white population). HLA-DQ3 thus appears to be a genetic marker of susceptibility to development of TE in AIDS patients, and DQ1 may be a resistance marker. These HLA associations with disease indicate that development of TE in AIDS patients is affected by a gene or genes in the HLA complex and that HLA-DQ typing may help in decisions regarding TE prophylaxis.
View details for Web of Science ID A1996TM34300042
View details for PubMedID 8537674
DQCAR is a very polymorphic CA repeat microsatellite located between the HLA DQA1 and DQB1 gene. Previous studies have shown that specific DQCAR alleles are in tight linkage disequilibrium with known HLA DR-DQ haplotypes. Of special interest was the fact that haplotypes containing long CA repeat alleles (DQCAR > 111) were generally more polymorphic within and across ethnic groups. In these latter cases, several DQCAR alleles were found even in haplotypes containing the same flanking DQA1 and DQB1 alleles. In this work, three HLA class II associated diseases were studied using the DQCAR microsatellite. The aim of this study was to test if DQCAR typing could distinguish haplotypes with the same DRB1, DQA1 and DQB1 alleles in control and affected individuals. To do so, patients with selected HLA DR-DQ susceptibility haplotypes were compared with HLA DR and DQ matched controls. This included: Norwegian subjects with Celiac disease and the HLA DRB1*0301, DQA1*05011, DQB1*02 haplotype; Japanese subjects with Type 1 (insulin-dependent) Diabetes Mellitus and the HLA DRB1*0405, DQA1*0302, DQB1*0401 haplotype; and French patients with corticosensitive Idiopathic Nephrotic Syndrome and the HLA DRB1*0701, DQA1*0201, DQB1*0202 haplotype. These specific haplotypes were selected from our earlier work to include one haplotype bearing a short DQCAR allele (celiac disease and DR3,DQ2-DQCAR99) and two haplotypes bearing long DQCAR alleles (Diabetes Mellitus and DR4,DQ4-DQCAR 113 or 115 Idiopathic Nephrotic syndrome and DR7,DQ2-DQCAR 111-121). Additional DQCAR diversity was found in both control and patients bearing haplotypes with long CA repeat alleles. The results indicate that DQCAR typing did not improve specificity in combination with high resolution DNA HLA typing as a marker for these three disorders.
View details for PubMedID 8560448
A highly polymorphic (CA)n microsatellite marker (DQCAR), located between the DQA1 and the DQB1 genes, was characterized in four ethnic groups. Based on length polymorphism, 12 alleles could be defined. The marker is located 1- to 2-kb telomeric to the DQB1 gene and 10 kb centromeric to the DQA1 gene and was shown to be in tight linkage disequilibrium with HLA-DQ. Analysis of the linkage disequilibrium pattern revealed little additional diversity in DQ1-associated haplotypes. Almost all DQ1 subjects examined were DQCAR 103 or DQCAR 107 (13 and 15 CA repeats, respectively). In contrast, significant haplotypic diversity was observed for most DQ2-, DQ3-, and DQ4-associated haplotypes. These haplotypes often had longer allele sizes (DQCAR > 111, more than 17 CA repeats) and more DQCAR alleles per haplotype. These haplotypes also carried DQCAR alleles of different sizes, even though they bore the same DQA1 and DQB1 alleles, and sometimes the same DRB1 allele as well. These results indicate that DQCAR could be a useful marker to better define disease associations with HLA. Our results are also consistent with the hypothesis that CAR alleles with higher numbers of repeats have higher mutation rates and that recombination within the HLA-DR/DQ region is haplotype dependent.
View details for Web of Science ID A1995QL39300004
View details for PubMedID 7759308
Narcolepsy is a neurological disorder known to be associated with human leukocyte antigen (HLA)-DQB1*0602 in humans. In a canine model, the disorder is also genetically linked to a gene of high homology with the human mu-switch-like immunoglobulin (Ig) gene (current LOD score 13.6 at 0% recombination). Since association with HLA or other immune function polymorphic genes (T cell receptor of Ig, mainly) is a hallmark of most autoimmune diseases, it is proposed that autoimmunity may also play a role in the development of narcolepsy. Arguments for and against this hypothesis are reviewed. It is shown that both on the basis of the most recent molecular studies, and because of some of its clinical features, narcolepsy may be an autoimmune disorder. However, neither systemic nor central nervous system (CNS) evidence of any autoimmune abnormality have ever been found. To reconcile this discrepancy, it is suggested that the pathological immune process involved in narcolepsy could be difficult to detect because it is restricted to a very small region of the brain or targets a low abundance neuroeffector. Alternatively, it is possible that a more fundamental relationship is involved between sleep generation and immune regulation. The pathophysiology of narcolepsy may then involve new CNS-immune mechanisms that may shed new light on the sleep process itself.
View details for Web of Science ID A1995QP30900004
View details for PubMedID 7795891
The identification of HLA class I alloantibodies is important for organ transplantation and platelet transfusion in alloimmunized patients. Because microcytotoxicity testing against frozen trays of lymphocyte panels is rapid and efficient for determining specificities of unknown antibodies, a simple method was devised to increase test sensitivity to weak antibodies. Standard anti-human globulin (AHG)-facilitated microcytotoxicity was modified by the insertion of a double addition-of-serum (DAS) step, and reagent and patient's sera were evaluated by both methods. DAS modification increased antibody titers and, more significantly, made the identification of weak specificities easier because of the twofold to threefold increase in reactivity rates (29-42% for AHG vs. 75-82% for DAS) of panel cells that were expected to be positive, while low (approx. 1%) "extra" reaction rates were maintained for cells that were expected to be negative. DAS was relatively unaffected by variations in serum volumes or target cell preparation, and its use did not significantly increase test time or costs. In a program of platelet donor selection driven by donor antibody rather than donor-recipient antigen matching, DAS greatly facilitated platelet transfusion support for alloimmunized patients.
View details for Web of Science ID A1992KC09400012
View details for PubMedID 1471248
Human narcolepsy is a genetically determined disorder of sleep strongly associated with the human leucocyte antigens (HLA) DR2 and DQw1. In black narcoleptic patients, susceptibility for narcolepsy is more closely related to a specific gene subtype of DQw1, DQB1-0602, than to DR2. About 30% of black narcoleptic patients are nonDR2, but all carry the HLA DQB1-0602 gene. In the present study, we have tested caucasian nonDR2 cataplectic patients (6 sporadic cases and 7 familial cases from 3 multiplex families) for the presence of the HLA DQB1-0602 and DQA1-0102 (DQw1) using a specific polymerase chain reaction (PCR)-oligotyping technique. None of the patients was DQB1-0602 or DQA1-0102 positive, thus proving that, in caucasians, DQB1-0602 and DQA1-0102 (DQw1) are not prerequisites for the diagnosis of narcolepsy. Further studies with more patients are warranted to exclude the possibility that a few caucasian patients carry rare haplotypes with DQB1-0602 independently of DR2.
View details for Web of Science ID A1992JT08400005
View details for PubMedID 1455124
Identification of genes determining narcolepsy susceptibility is important not only for understanding that disorder but also for possible clues to general sleep-control mechanisms. Studies in humans reveal at least one such gene related to the major histocompatibility complex and in dog an as-yet-unmapped single, autosomal recessive gene canarc-1. Gene markers for canarc-1 were therefore sought by DNA restriction fragment length polymorphisms in our colony of narcoleptic dogs. A human mu-switch immunoglobulin probe and the enzyme Hae III identified a gene cosegregating with canarc-1 in backcrossed animals (logarithm of odds scores: m = 24, Z max = 7.2 at theta = 0%). canarc-1 was also shown not to be tightly linked with the dog major histocompatibility complex (m = 40, Z less than -2 at theta less than 4.8%). These results represent the mapping of a non-major histocompatibility complex narcolepsy gene and strongly suggest involvement of the immune system in the pathophysiology of that disease.
View details for Web of Science ID A1991FG91300106
View details for PubMedID 1673032
Familial patterns of narcolepsy were investigated in a clinic population of 334 unrelated narcoleptic patients. 40% of probands had at least 1 family member with an isolated daytime sleepiness complaint and 6% had a positive family history of narcolepsy. Multicase families were rare; only two families were found with 3 or more affected relatives. Family members often shared the same HLA-DR2 haplotype as the proband but did not have narcolepsy. However, the risk of disease for first-degree relatives was six to eighteen times greater than that for unrelated individuals. Although most patients were HLA-DR2+, 2 new HLA-DR2- individuals were found. The data predict that as many as 9% of unrelated North-American white patients with narcolepsy will be DR2-. Analysis of these and other data indicates that although strongly associated with disease, the HLA-DR2 haplotype is neither sufficient nor necessary for the development of narcolepsy.
View details for Web of Science ID A1989CD35200012
View details for PubMedID 2574313
Two soluble, secreted forms of HLA-B7 were engineered by the creation of hybrid human/mouse molecules containing the polymorphic 5' region of the HLA-B7 gene and the secretory 3' region of the mouse Q10d gene. The hybrid, designated F1, is the first construct with only human extracytoplasmic domains, consisting of exons for the leader peptide and the three extracellular domains (alpha 1, alpha 2, alpha 3) of B7 spliced to the exons for the Q10d truncated transmembrane and 3' untranslated (3'UT) sequences. The second construct, designated C2, is similar but has the human alpha 3 replaced by the Q10 alpha 3 domain. Protein product from each construct was best demonstrated after gene transfection into the J27.2 cell line. In particular, secretion of the F1 product proves that the Q10 alpha 3 domain is not necessary for secretion of class I/Q10 hybrids. Moreover, the two soluble B7 forms, which differ only in their alpha 3 domain, are similarly recognized by monoclonal antibodies W6/32 (anti-HLA-ABC), BBM.1 (anti-human beta 2 microglobulin), and allo-B7-antibody, but differentially recognized by monoclonal antibody Q1/28 (anti-HLA class I heavy chain). Production of such soluble hybrid class I molecules in large amounts should allow critical structural and functional studies of these proteins.
View details for Web of Science ID A1989AC04000006
View details for PubMedID 2670852
Human narcolepsy is almost exclusively associated with the major histocompatibility complex (MHC) class II antigen HLA-DR2 and is the strongest HLA-disease association described to date. Canine narcolepsy resembles the human disease in its behavioral manifestations and responses to therapeutic drugs. Therefore, mixed leukocyte culture (MLC) was used to study differences in the canine MHC class II (DLA-D) antigens present in narcoleptic dogs to determine whether an analogous, unique DLA-D antigen could be identified in canine narcolepsy. Results show at least five different DLA-D antigens appear in potential narcoleptic haplotypes among the 29 dogs studied. The data demonstrate that, unlike man, in dogs there is no unique D locus antigen associated with narcolepsy and further suggest that linkage disequilibrium with a specific MHC antigen is unlikely to be essential for the manifestation of canine narcolepsy. Because human narcolepsy is thought to be multigenic, the canine narcolepsy-MHC dissociation suggests that the dog model may help elucidate the non-MHC narcolepsy gene(s).
View details for Web of Science ID A1989U131500003
View details for PubMedID 2523880
In light of increasing public and employee concern over potential infectious hazards associated with blood and other body fluids, several government agencies (the Food and Drug Administration, the Centers for Disease Control, the Occupational Safety and Health Administration, the Environmental Protection Agency, the Health Care Financing Administration and the National Heart, Lung and Blood Institute) cosponsored a Biosafety Workshop in April 1988. The objective of the workshop was to identify appropriate biosafety practices and standard control procedures to protect workers involved in the collection, storage, and transportation of human blood donations with the least possible disruption of the nation's blood supply. Speakers focused on human immunodeficiency virus (HIV) and hepatitis B virus (HBV); however, the safety principles discussed were considered equally applicable to other known (e.g., non-A, non-B hepatitis and human T-lymphotropic virus type I (HTLV-1) blood-transmitted infections. The resulting consensus included the need for blood establishments to develop and apply thoughtful biosafety programs to address staff training, accident prevention, HBV vaccination, handling spills, managing contaminated waste and transporting blood specimens. There was lack of agreement, however, on the usefulness of gloves during the phlebotomy of healthy blood donors.
View details for PubMedID 3420681
RecA protein-coated probe has been utilized to enrich genomic digests for desired genes in order to facilitate cloning from genomic libraries. Using a previously cloned HLA-B27 gene as the recA-coated enrichment probe, we obtained a mean 108x increase in the ratio of specific to nonspecific plaques in lambda libraries screened for B27 variant alleles of estimated 99% homology to the probe. Class I genes of lesser homology were less enriched: 6.7x for non-B27 genes of estimated greater than 95% homology and 3.7x for other-Class I genes of greater than 80% homology. Loss of genomic DNA during the enrichment procedure can, however, restrict application of this technique whenever starting genomic DNA is very limited. Nevertheless, the impressive reduction in cloning effort and material makes recA enrichment a useful new tool for cloning homologous genes from genomic DNA.
View details for Web of Science ID A1988P964900029
View details for PubMedID 2901713
Paroxysmal cold hemoglobinuria is a rare and potentially life-threatening acquired hemolytic anemia occurring either as an acute transient anemia following several different viral syndromes, or in a chronic idiopathic form. Episodic hemolysis in paroxysmal cold hemoglobinuria is usually associated with a biphasic (Donath-Landsteiner) IgG cold-reactive complement-fixing autohemolysin with anti-P specificity. Paroxysmal cold hemoglobinuria has not previously been associated with malignancy nor has it been clearly shown to be steroid-responsive. This report describes a patient with steroid-responsive autoimmune hemolytic anemia and immune thrombocytopenia (Evans' syndrome) associated with oat cell carcinoma of the lung and a unique biphasic anti-IgM autohemolysin. This case extends the spectrum of biphasic antibody-mediated immune cytopenias and widens both the clinical and the serologic definition of paroxysmal cold hemoglobinuria.
View details for Web of Science ID A1987H438200030
View details for PubMedID 3578344
B lymphocytes from Rh negative donors with serum anti-D antibodies were isolated and fused with the mouse-human heteromyeloma, SBC-H20, to produce hybridomas secreting IgM or IgG1 human monoclonal antibodies to D antigen. The IgM antibody in hybridoma supernatant agglutinates all normal D positive cells at the immediate spin phase of reactivity. Using concentrated IgM hybridoma supernatant of approximately 50 micrograms/ml, Du cells were also agglutinated. The IgG1 antibody reacts by indirect hemagglutination with all D and Du cells. Against Rh mosaics, different reactivity was noted for each antibody. Furthermore, D positive cells precoated with the IgG1 antibody inhibit the IgM direct hemagglutination, suggesting that the antibodies identify closely associated epitopes. These human monoclonal antibodies will be useful diagnostic reagents and, ultimately, should be useful in the prevention of Rh hemolytic disease of the newborn.
View details for Web of Science ID A1987J436100009
View details for PubMedID 3116771
Genomic DNA from 46 B27+ ankylosing spondylitis, Reiter's syndrome, or normal individuals was digested with Taq I and probed, in Southern blots, with the HLA-B locus specific probe, EI7. Four restriction fragment length polymorphisms (RFLP), 2.5, 3.4, 3.8 and 4.0 or 8.0 kb, were observed for the B27 gene. In Caucasians, one of the B27 variants (2.5 kb) was more frequent in normals and almost never appeared in patients, suggesting a trend that is not yet statistically significant. In the course of defining the B27 polymorphisms, three and two RFLP, respectively, were also found for the B18 and B44 genes.
View details for Web of Science ID A1987F630600004
View details for PubMedID 2879815
Post-dysenteric or reactive arthritis (ReA) is closely associated with HLA-B27. This histocompatibility antigen is heterogeneous and consists of 2 serologically defined variants: B27M1+M2+ and B27M1+M2-. This paper gives a qualitative evaluation of the antibodies present in the sera of 62 patients with dysentery due to Shigella flexneri 2a, a known arthritogenic bacterium. The patients were classified in 4 groups: B27M1+M2+ReA+ (n = 5), B27M1+M2+ReA- (n = 7); B27M1+M2-ReA- (n = 1); B27-ReA- (n = 49). The isolated infectant possessed cell envelope antigens with B27M2-like epitopes (Mr 20,000). Analysis of the spectrum of antibodies directed against the separated cell envelope antigens of S. flexneri in the sera of these patients revealed 7 main patterns of reactivity. The detectable immunogens encompassed protein stainable antigens (Mr 98, 78, 68, 54, 50, 44, 41, 35, 14 and 13 kDa), lipopolysaccharides and peptidoglycan. None of the sera possessed detectable antibodies to the B27M2-like antigen. Consequently, this antigen is unlikely to be associated with ReA, and this applies equally to other antigens or patterns of antigens. The arthritogenicity of S. flexneri may therefore not be determined by the presence or absence of detectable antibody titers to certain cell envelope antigens. We hypothesize that other properties of these antigens could be of significance.
View details for Web of Science ID A1986E077900006
View details for PubMedID 3533766
Events in pathogenesis and immunity during primary varicella-zoster virus (VZV) infection were examined in 64 healthy subjects and 21 immunocompromised patients. Activation of the interferon system and activation of circulating T lymphocytes were early immune responses that occurred during the incubation period in some healthy subjects. Elevated levels of 2-5A synthetase in peripheral blood mononuclear cells and detection of serum alpha interferon (IFN-alpha) and gamma interferon (IFN-gamma) were present in the majority of healthy subjects who had acute primary VZV infection. Expression of HLA-DR antigen occurred on circulating T lymphocytes from subjects with acute VZV infection. The early production of VZV-specific IgG or IgM antibodies did not correlate with the severity of the clinical infection, but the detection of T lymphocyte proliferation to VZV antigen within three days after the appearance of the varicella exanthem was associated with milder illness. The mean VZV-specific lymphocyte transformation for subjects with less than 100 lesions/m2 was 7.5 +/- 10.43 SD compared with 1.4 +/- 1.85 SD for those with greater than 400 lesions/m2 (P less than .05). Only one (7.7%) of 13 immunocompromised patients had early VZV-specific lymphocyte transformation compared with 19 (42%) of 45 healthy subjects (P less than .05). The rapid host response to primary VZV infection was associated with rapid termination of viremia in healthy subjects; VZV was isolated from only 11% of peripheral blood mononuclear cell samples cultured within 48 hr after the appearance of the exanthem.
View details for Web of Science ID A1986D760000006
View details for PubMedID 3016110
The heterogeneous HLA-B27 antigen is closely associated with post-infectious or reactive arthritis (ReA) and is comprised of two serologically defined variants: B27M1+M2+ and B27M1+M2-. An outbreak of dysentery (n = 120) caused by a Shigella flexneri 2a strain, which possessed cell envelope antigens with epitopes resembling B27M2, resulted in five B27M1+M2+ patients with ReA. The remaining seven B27M1+M2+, one B27M1+M2- and all but three B27-negative patients remained free of joint symptoms; the latter three displayed arthralgia. IgM, IgG and IgA serum titers were statistically raised in all patient groups, but were exceptionally and persistently high in the B27M1+M2+ patients with ReA, especially IgA, as determined in acute-phase sera and sera sampled 1 year after dysentery. B27M1+M2+ thus appears to be a marker for a subset of disease, characterized by a high immune response. It is concluded that the B27M2 epitope is not unequivocally disease-related to Shigella ReA, that B27M1+M2+ is not likely to be the only immune-response-regulating gene involved in this form of ReA and that cross-reactivity between bacterial antigenic epitopes and B27 can only be part of a multifactorial process leading to ReA and in itself not sufficient to produce ReA. The intensity of the immune response appears to be another important factor.
View details for Web of Science ID A1986D405300013
View details for PubMedID 2428743
A patient with common variable immunodeficiency syndrome tolerated intramuscular IgG (which contains IgA) and an initial infusion with intravenous (IV) IgG, but developed reactions to subsequent IV IgG. High-titre, class-specific anti-IgA antibodies were detected suggesting immunization by the IgA-contaminated IV immunoglobulin. Subsequent IgG replacement was achieved with IgA-deficient plasma infusions. Patients who tolerate intramuscular IgG may not tolerate the IV preparations.
View details for Web of Science ID A1986C336200010
View details for PubMedID 2422990
A monoclonal IgG antibody was produced from a mouse immunized with an A11, A24; B27, B44 Epstein-Barr virus transformed B lymphoblastoid cell line. The antibody, A11.1M, by standard lymphocytotoxicity assay, reacts with all cells expressing HLA-A11 and -A24. Absorption studies with both A11+, A24- and A11-, A24+ platelets removed antibody reactivity against A11 and A24 lymphocytes. The shared antigenic determinant between A11 and A24, as defined by this antibody, A11.1M, represents a new "supertypic" determinant.
View details for Web of Science ID A1986A381800006
View details for PubMedID 2420768
To investigate the possible coinheritance of autoimmune diseases that are associated with the same HLA antigen, we studied 70 families in which at least two siblings had either type I diabetes mellitus (IDDM), autoimmune thyroid disease (ATD), rheumatoid arthritis (RA), or a combination of these diseases. HLA-A, B, and C typing was performed on all affected sibs in one generation or more. First, we estimated by sib-pair analysis the disease allele frequency (pD) and the mode of inheritance for each disease. According to the method of ascertainment entered into the analysis, the pD for ATD ranged from .120 to .180, for an additive (dominant) mode of inheritance. For RA, the pD ranged from .254 to .341, also for additive inheritance, although recessive inheritance could not be excluded. For IDDM, the pD ranged from .336 to .337 for recessive inheritance; additive inheritance was rejected. Second, we examined the distribution of shared parental haplotypes in pairs of siblings that were discordant for their autoimmune diseases. The results suggested that the same haplotype may predispose to both IDDM and ATD, or IDDM and RA, but not to both RA and ATD. Analysis of pedigrees supported this hypothesis. In 16 families typed for HLA-DR also, the haplotype predisposing to both IDDM and ATD was assigned from pedigree information to DR3 (44%), DR4 (39%), or DR5, DR6, or DR7 (5.5% each). In some families, these haplotypes segregated over several generations with ATD only (either clinical or subclinical), suggesting that in such families, ATD was a marker for a susceptibility to IDDM. In several families, an IDDM haplotype segregated with RA but not with ATD. This suggests that ATD- and RA-associated susceptibilities to IDDM may be biologically different and thus independently increase the risk of IDDM.
View details for Web of Science ID A1986A311100006
View details for PubMedID 3456197
With the use of monoclonal antibodies in an indirect immunofluorescence technique we studied the distribution of Class I (HLA-ABC and B27) and Class II (HLA-DR) antigens in the human uvea. W6/32, directed against the core of HLA-ABC antigens, was used to study the distribution of Class I antigens. The anterior border layer of the iris, the non-pigmented and pigmented epithelium and the external basement membrane of the ciliary body and the vascular endothelium in the uvea showed a positive staining for Class I antigens. B27/M1, directed against an epitope of the HLA-B27 antigen, and the control antibody A11/Aw24, which was directed against an epitope of HLA-A11, revealed the same distribution pattern in respectively HLA-B27 and HLA-A11 positive donor eyes. The intensity of their staining was much weaker than the staining with W6/32. Class II antigens were studied with OkIa1, an antibody directed against the core of HLA-DR antigens. HLA-DR antigens were detectable on single cells scattered throughout the entire uvea. These cells did not seem to relate to any anatomical entity. No staining for Class II antigens was seen in the uveal blood vessel endothelium. The expression of HLA-antigens in the uvea is compatible with the distribution in other tissues. These findings suggest that the expression of HLA-B27 in the human uvea does not explain why the eye is one of the target tissues in HLA-B27 associated disease.
View details for Web of Science ID A1986A022400012
View details for PubMedID 3512217
Hybridomas secreting a human monoclonal IgG1 antibody against a variant of the rhG antigen were produced from B lymphocytes of an Rh-negative donor with serum antibodies to D and G [correction of C] antigens. The antibody reacts by indirect hemagglutination with nearly all C- or D-positive cells, confirming the strong association in the expression of G with D or C antigens. The lack of antibody reactivity to C-negative Du cells suggests a particular epitope on the D complex associated with the G antigen.
View details for Web of Science ID A1986A969300008
View details for PubMedID 2424176
The JY328 clone was identified in a human genomic library using cDNA corresponding to mRNA for HLA-B7 as a probe. The L/328 cell line was established by cotransformation of mouse Ltk- cells with the herpes thymidine kinase gene and clone JY328. On Northern blots, RNA from L/328 strongly hybridized to an HLA class I probe, and an antigen was recognized by an anti-HLA class I framework antibody on the cell surface. A DNA probe corresponding to a segment of intron 7 was developed by comparing the nucleotide sequence of clone JY328 with that of other HLA class I-type genes. Using the radiolabeled probe to screen Southern blots of DNA from families with siblings exhibiting intra-HLA recombinations, a restriction fragment length polymorphism was revealed--a 1.4 kb BstE II band not present in all individuals. A corresponding fragment was apparent in the base sequence of clone JY328. The occurrence of this band on Southern blots established that JY328 maps distinct from and centromeric to the HLA-C locus and near to the HLA-B locus. Antibody absorption studies and cytotoxicity tests indicated that the JY328 gene product was not an HLA-B antigen but that it did specifically absorb CW7-specific antibody. In sum, these results suggest a novel, polymorphic HLA class I gene which expresses a product serologically similar to HLA-Cw7 but which does not map within the corresponding locus.
View details for Web of Science ID A1986A627600004
View details for PubMedID 3007345
The incidence of transfusion-acquired primary cytomegalovirus (CMV) infection was studied in 483 cardiac surgery patients. Ninety-six patients (20%) were found to lack antibody to CMV [CMV Ab(-)] as measured by radioimmunoassay. Sixty-eight CMV Ab(-) were followed by viral culture and/or serology from eight weeks to one year after transfusion. Transfusion requirements in CMV Ab(-) patients were as follows: whole blood/packed red blood cells, mean 4.7 +/- 2.6 units; platelets (20 patients), 6.9 +/- 3.8 units; fresh frozen plasma (25 patients), mean 3.3 +/- 1.6 units. Forty-nine percent of 235 donor units tested had antibody to CMV. One donor unit (0.4%) had CMV-specific IgM. This was not associated with CMV infection in the recipient. One patient (1.5%) demonstrated evidence of seroconversion to CMV during the follow-up period. This is significantly less than reported in previously published studies (P less than .01). Serological methods used, the age of the transfused blood, the immune status of the transfusion recipient, and the administration of passive antibody in fresh frozen plasma are factors that may be responsible for the low incidence observed.
View details for Web of Science ID A1985ACT8400008
View details for PubMedID 2984327
A 12-year-old boy with severe combined immunodeficiency who had been kept in a gnotobiotic environment since birth received bone marrow from a histoincompatible sibling in an attempt to reconstitute immunologic function. To prevent graft versus host disease, the donor's marrow was treated in vitro with monoclonal antibody and complement to remove alloreactive T cells. Eighty days after transplantation, the patient had a systemic illness characterized by fever, thrombocytopenia, gastrointestinal pain, and bleeding; he died on the 124th post-transplantation day. Postmortem examination revealed multiple tumor-like B-cell proliferations, recipient in origin, in numerous organs. Epstein-Barr virus (EBV) was isolated from the patient's pharyngeal secretions; EBV nuclear antigen was found in spontaneously transformed peripheral-blood lymphocytes, inflammatory cells from peritoneal fluid, and bone marrow cells; and EBV genomes were discovered in all tumor tissues. The donor's serum showed evidence of past EBV infection. Analysis of cellular immunoglobulin and immunoglobulin gene DNA from the tumors indicated both monoclonal and oligoclonal B-cell proliferations. These findings provide evidence for the evolution of EBV-induced polyclonal activation of B cells to oligoclonal B-cell proliferation and finally to monoclonal B-cell lymphoma.
View details for Web of Science ID A1985AGA9500004
View details for PubMedID 2984567
Polymorphic restriction endonuclease sites within the HLA-DR alpha gene have been defined, localized, and used as genetic markers in the analysis of susceptibility to insulin-dependent diabetes mellitus (IDDM). Hybridization of Bgl II-digested human genomic DNA with a cDNA clone for the HLA-DR alpha chain (pDR alpha-1) has revealed three allelic restriction fragment lengths: 3.8 kilobase pairs (kb), 4.2 kb, and 4.5 kb. Hybridization of EcoRV-digested human genomic DNA with the same probe has revealed two allelic polymorphic restriction fragment lengths: 9.2 kb and 13.0 kb. By analysis of double digests of genomic DNA from individuals homozygous for each of the allelic variants, the polymorphic restriction sites were found to be clustered near the 3' end of the HLA-DR alpha gene. The observed correlations of DR alpha Bgl II restriction site variants with serologically determined DR specificities suggest linkage disequilibrium between the DR alpha and DR beta loci. The 3.8-kb fragment is correlated with the DR1 type (Pc = 4.4 X 10(-4)); and the 4.2-kb fragment, with a subset (B8,DR3) of the DR3 type (Pc = 5.1 X 10(-4)) and with the DR6 type. The segregation pattern of HLA-DR alpha polymorphic Bgl II restriction fragments was analyzed in six IDDM families. The observed association of IDDM with the Bgl II 4.2-kb DR alpha restriction variant is higher than with existing serological markers and supports the utility of this approach in elucidating IDDM inheritance.
View details for Web of Science ID A1985AVT4000062
View details for PubMedID 2999792
Despite major advances in genetic and structural studies of the HLA-B27 antigen, the underlying mechanism responsible for the remarkable association between this antigen and spondylarthropathies remains unknown. At a molecular level, the use of B27M1 and B27M2 monoclonal antibodies has permitted the identification of distinct allospecific epitopes on the B27 molecule. One of these epitopes, B27M2, is polymorphic and has allowed us to define B27 variants: B27M2[+], B27M2[-], and B27M2[int]. The heterogeneity of the B27 antigen correlates well with biochemical and cytotoxic evidence of genetic heterogeneity. These variants exhibit ethnic variation and also appear to correlate, in preliminary studies, with disease susceptibility, especially among Orientals. HLA gene probing is potentially an even more precise tool than monoclonal antibodies for the study of MHC-related disease susceptibilities. Initial work in our laboratory has resulted in the production of probes with specificity for HLA-B locus genes and current efforts are directed toward the derivation of B27 allele-specific probes. It seems likely that, when such probes are applied to B27-positive individuals, complexity in addition to the B27M2 variants will be revealed. Yet to be defined is the mechanism behind the association between B27 and AS. Is the association causal for disease, or is B27 indeed just a marker for other pathogenic factors somehow linked to it? Available evidence points to both causal and linked roles for B27 in ankylosing spondylitis. Products of both HLA and non-HLA gene families may interact with infectious disease pathogens in susceptible individuals to produce a disorder which may not be specific in its association with any one pathogenic factor.
View details for Web of Science ID A1985AQR7200004
View details for PubMedID 2412952
View details for Web of Science ID A1985ABP6600252
An assay has been developed to distinguish active from passive Rh0(D) immunization in a patient who had recently received hyperimmune anti-Rh0(D) immunoglobulin therapy. Isolated peripheral B lymphocytes from a pregnant woman at 32 weeks gestation were co-cultured with Epstein-Barr virus in a liquid growth medium. After 7 days, anti-Rh0(D) antibodies produced in vitro by the transformed lymphocytes were detected in culture supernatants, thereby proving active immunization and indicating the potential of hemolytic disease of the newborn in the current pregnancy. This assay was also performed with peripheral B lymphocytes from three groups of individuals: mothers known to be Rh0(D) immunized and who recently delivered Rh-positive infants, women with longstanding Rh0(D) immunization, and women who were treated with anti-Rh0(D) globulin. In the first group, anti-Rh0(D) antibodies were again detected after in vitro viral stimulation. In the latter two groups, essentially no anti-Rh0(D) activity was detected.
View details for Web of Science ID A1985APS1800005
View details for PubMedID 2996227
Hybridomas secreting human monoclonal antibodies to varicella-zoster virus were produced by fusing B cells of a patient recovering from acute varicella infection with a human-mouse cell line. Two hybrid lines have continued to secrete IgG1, one with kappa and the other with lambda chains, for at least 12 months. Each antibody neutralizes virus infectivity between 1-5 micrograms of partially purified immunoglobulin/ml, each shows a different pattern of immunofluorescent staining of virus-infected cells, and one identifies three viral proteins with molecular weights of 60,000, 95,000, and 97,000.
View details for Web of Science ID A1985AQZ7900006
View details for PubMedID 2993433
A human-mouse cell line that is hypoxanthine-aminopterin-thymidine sensitive and ouabain resistant was derived from a fusion between human B lymphocytes and a mouse myeloma line. This new mutant, when fused to a relatively unstable EBV-transformed B cell secreting a human monoclonal anti-A (red blood cell antigen) antibody, resulted in stable hybridomas capable of long term production of the specific human monoclonal antibody. Furthermore, some of the hybrid clones secreted antibody in far greater titer than the original EBV cell line. We conclude that fusion to this human-mouse line is an efficient approach to the production of human monoclonal alloantibodies and an effective method of 'rescuing' secretion of desired antibody from EBV cell lines.
View details for Web of Science ID A1984SR32200010
View details for PubMedID 6201562
Variants of a lymphoblastoid cell line, LCL 526 (SB3 MB1 DR1 B44 C5 A2/SB4 MT4 DR4 B27 C2 A24), which lost various HLA specificities were selected with monoclonal antibodies and complement using a method developed by Kavathas et al. (PNAS 77:4251, 1980). Using alpha B27 monoclonals, 8 B27 only loss mutants and 4 B27 haplotype multiple loss mutants were generated. The parental LCL 526 and two of the B27- mutants were used to select alpha B27 CTLs. The selection of six A2 loss, one A2-C5 loss, and 14 A2 haplotype multiple loss variants as well as secondary selection on haplotype loss variants to obtain A null, B null, DR null, and total A, B, C, null variants is also described. The usefulness of these mutants for the study of the relationship between B27 and disease and as two new haplotypes for immunologic, genetic, and molecular research is discussed. These mutants are available to other researchers.
View details for Web of Science ID A1984TG90400003
View details for PubMedID 6090351
Anti-Epstein-Barr virus and antiinfluenza A cytotoxic T lymphocytes (CTL) have been used to study the restriction of human antiviral responses by HLA-B27 antigens. Three functional subgroups of HLA-B27 have been clearly distinguished by this "restriction-typing assay". No cross-reaction could be detected between the three subgroups either at the CTL level or at the level of antigen-presenting cells. The cells of subgroup 1 are always positive [M2(+)] when tested in immunofluorescence with a monoclonal B27-specific antibody which divides HLA-B27 into a major M2(+) and a minor M2(-) subgroup. These M2(+) group 1 cells are apparently also HLA-B27W as previously shown by Ivanyi and co-workers using anti-HLA-CTL. Subgroup 2 includes only M2(-) cells. A comparison between this group and the previously described HLA-B27K is not fully conclusive, since two typing cells which were clearly HLA-B27K apparently did not belong to group 2. Only two donors, both of Oriental origin, have been included in subgroup 3. Both of them were "M2 intermediate". These results demonstrate (1) the existence of several functional subgroups of HLA-B27 with an interesting correlation with the M2(+), M2(-), or M2 intermediate phenotypes, and (2) the possibility of using the restriction-typing assay to define such functional subgroups not detected by classical allosera.
View details for Web of Science ID A1984TT41400005
View details for PubMedID 6094341
A trial of the killed Coccidioides immitis spherule vaccine was undertaken with 151 healthy skin test negative adult volunteers and controls to evaluate the safety of selected regimens, the induction of humoral and cell-mediated immune responses, and to determine if there were immunogenetic differences in these responses. The vaccine was given as three intra-deltoid doses over 8 weeks. No severe systemic symptoms were noted, although 3% of 3.5 mg doses (but no 1.75 mg doses) were associated with severe local reactions. Half the vaccinees had skin test conversions, which generally persisted greater than or equal to 6 months, two-thirds showed boosting of lymphocyte transformation in vitro, and 16% given three 3.5 mg doses developed antibody. There was an association between degree of local adverse vaccine reaction and immunostimulation, and a trend to immune response in persons of O blood type and with some HLA phenotypes. There was no evidence of deficient response to vaccination in subpopulations known to respond to coccidioidal infection poorly. A regimen of three 1.75 mg doses appears to be safe and without reduced immunogenicity, and there is no evidence dosage modification for certain subpopulations would be necessary in efficacy studies.
View details for Web of Science ID A1984SM10600013
View details for PubMedID 6424435
View details for Web of Science ID A1984TB41900004
Sensitivity of the carboxyfluorescein diacetate (C-FDA) thrombocytotoxicity technique for the detection of antiplatelet antibodies has been enhanced by the addition of an anti-Kappa light chain antibody facilitation step. This new technique, KC-FDA, was compared with the platelet suspension immunofluorescence test (PSIFT) by titering platelet-reactive allo-anti-PlAl and anti-HLA antibodies. The results show that compared to PSIFT, KC-FDA is more sensitive for detecting platelet specific antibodies (PlAl), is more or equally sensitive for detecting other antibodies (HLA), and is significantly faster and easier to perform.
View details for Web of Science ID A1983RV47400004
View details for PubMedID 6418696
Detailed study of HLA-B27 was prompted by the extremely strong associations between this antigen and spondyloarthropathies. Despite the relative homogeneity of this antigen when defined by alloantisera, B27 reactivity with the monoclonal antibody B27M2 suggests previously unrecognized heterogeneity. To define and confirm this heterogeneity on a molecular level, detergent extracts were prepared from B cell lines derived from individuals reactive (+) or unreactive (-) with the B27M2 antibody. Extracts were immunoprecipitated by specific allogeneic or monoclonal antibodies and analyzed by two-dimensional polyacrylamide gel electrophoresis. By this method the B27M2+ and B27M2- variants of HLA-B27 had different isoelectric points (pl) and could be distinguished from each other and from a different (Bw44) control alloantigen. Blockade of glycosylation by pretreatment of cells with tunicamycin did not alter pl but did reduce HLA antigens by approximately 3000 daltons. These data demonstrate that B27 antigens can be subdivided into subsets with different molecular composition. The effects of this heterogeneity upon the associations of B27 and disease are not yet known.
View details for Web of Science ID A1983QC72100051
View details for PubMedID 6600481
To explore the possibility that the HLA-B27 antigen may exist in more than one form, murine monoclonal antibodies were produced to B27 molecules. The first such antibody, anti-B27M1, reacts with 100% of B27 + cells and, therefore, does not distinguish B27 + healthy individuals from those with spondyloarthropathies. A second antibody, anti-B27M2, recognizes a variant of the B27 molecule that is present in most but not all Caucasian B27 + individuals. The frequency of B27M2(+) variants also appears to differ among different ethnic groups. Preliminary studies suggest that B27 + patients with ankylosing spondylitis are less likely than B27 + healthy individuals to express the B27M2 variant.
View details for Web of Science ID A1983RX57500015
View details for PubMedID 6420568
Because HLA-B27 is such an important genetic marker for susceptibility to spondyloarthropathies and other related diseases, study of this alloantigen was undertaken using murine monoclonal antibodies. The first monoclonal anti-B27 antibody, B27M1, is an IgG2a lymphocytotoxic antibody that reacts with all B27 antigens and cross-reacts to a lesser degree with the B7 antigen. Cross-reactivity with B7 is attributed to a B27-like epitope on the B7 antigen but which, as determined by cytotoxicity blocking studies, is distinct from the B7 allospecific epitope itself. The second B27 monoclonal antibody, B27M2, is a lymphocytotoxic IgM antibody reacting with most (greater than 85%) but not all of B27 antigens tested and having no cross-reactivity with B7. As determined by cytotoxicity blocking studies, the B27M2 epitope appears near the allospecific B27 epitope and the B27M1 epitope on the B27 antigen molecule. Immunochemical studies reveal isoelectric point and size differences between the B27M2+ and the B27M2- variants of HLA-B27. Based on these data, a model is proposed for the B27 antigen in which a B27M1 epitope is constant but the B27M2 epitope is present on the B27 antigen of some individuals and absent from that of others. B27M1 antibody reactivity does not appear to be associated with unusual disease susceptibility but preliminary data suggest the B27M2- variant of B27 may be more strongly associated with ankylosing spondylitis.
View details for Web of Science ID A1983RX69800021
View details for PubMedID 6360285
The role of genetic susceptibility in the pathogenesis of Hodgkin's Disease has been considered relatively minor because of the rarity of familial disease, the absence of an identified pattern of inheritance, and the weakness of HLA associations in population studies. The availability of four prospectively ascertained HD families permitted reappraisal of the cosegregation of HLA and HD susceptibility by a new extended concordance analysis method. HLA haplotype concordance among patients was greater than that expected by chance alone for our four families (P less than 0.022) and also for these in combination with twelve informative families in the literature (P less than 0.0015). This study thus provides a new method, based on genotype concordance of affected relatives, for assessing linkage of HLA and disease susceptibility, and new evidence for the genetic control of susceptibility to HD. The model presented, as well as alternative and more complex models, points to the existence of an HD susceptibility gene in or near the HLA region, which, in the presence of a suitable etiologic agent or additional genetic susceptibility, leads to the induction of HD.
View details for Web of Science ID A1983QJ57600005
View details for PubMedID 6221004
A new modification of an HLA-DR typing technique is described which makes DR typing as rapid and simple as routine HLA-A,B,C typing. In this new method, designated the TM1 technique, carboxyfluoresceindiacetate labeled peripheral blood lymphocytes are added directly to DR typing trays. The T cells are then lysed by addition of TM1, a pan-T cytotoxic IgM monoclonal antibody, and residual B-cell reactivity with cytotoxic DR alloantibodies is read as in routine fluorochromasia microlymphocytotoxicity. HLA-DR typing by the TM1 technique compares favorably to typing by methods using B cells enriched by sheep red blood cell rosetting or by Degalan bead columns. The TM1 technique also works well with cells that have been cryopreserved as well as with cells that have been separated from whole blood drawn as much as 3 days earlier. Finally, because TM1 is so effective in lysing normal T lymphocytes, this antibody may prove useful in functional in vitro and in vivo studies requiring T-cell depletion.
View details for Web of Science ID A1983QF45400001
View details for PubMedID 6601099
The production of hybridomas between immunologically activated T cells and malignant T-cell lines offers a potentially unlimited source of soluble T-cell-derived products. Recently, human T-T hybrids have been described; however, their use has been hampered by slow growth and chromosomal instability due at least in part to the presence of thymidine in the traditional hypoxanthine/aminopterin/thymidine (HAT) selection medium. In this report, we describe the development of a rapidly growing hypoxanthine phosphoribosyltransferase-deficient human T-cell line designated J3R7, the use of azaserine/hypoxanthine (AH) medium as an alternative selection medium to HAT medium, and the production of functional T-T hybrids by using the J3R7 line and the AH selection technique. Hybrids selected in AH medium were 4-fold greater in number and 3-fold faster in growth rate than hybrids grown in HAT medium. No stable clones were obtained from HAT cultures whereas AH-derived hybrids could be readily cloned by the method of limiting dilution. Evidence for hybridization included (i) the presence of approximately twice the number of chromosomes in hybrids than in J3R7 cells; (ii) the presence on hybrid cells of the Leu-3a surface antigen, present on normal helper T cells but not on J3R7 cells; (iii) the expression of HLA antigens of both the normal T-cell partner and the J3R7 line; and (iv) the constitutive secretion of interleukin 2 from multiple hybrid clones but not from the J3R7 cell line. Thus far, these clones have maintained their rapid growth, chromosome number, surface phenotype, and constitutive secretion of interleukin 2 for 4 months.
View details for Web of Science ID A1982PT40700084
View details for PubMedID 6984190
The need for anti-complement (anti-C') activity in antiglobulin antisera (AHG) for the detection of clinically significant antibodies was evaluated during a three-year period. While performing routine compatibility testing using standard blood banking procedures, eight patients were found whose antibodies were detectable primarily or only by AHG containing anti-C' activity; monospecific anti-igG AHG gave weak or negative reactions. Seven of the antibodies were anti-jka or jkb. Two of the anti-jka antibodies were responsible for clinically unsuspected delayed hemolytic transfusion reactions. The anti-jkb antibody resulted in a shortened survival of incompatible 51Cr-labelled red blood cells. The incidence of such "complement-only" Kidd antibodies was 23 percent of all Kidd antibodies found. These data suggest that the omission of anti-C' in AHG in routine compatability testing could result in substantial risk of failure to detect clinically significant antibodies.
View details for Web of Science ID A1982NZ87800003
View details for PubMedID 6980506
Using a large battery of Bw16, w38, and w39 antisera, a new variant of Bw16 has been identified in four unrelated Mexican-American families. The serologic pattern obtained is distinct from that for Bw38, Bw39, and 8W57 antigens. Absorption studies confirm the existence of this new Bw16 subtype which we term ST-16. ST-16 is Bw6-associated, with antigen frequency estimated to be 2.5% in Mexican-Americans.
View details for Web of Science ID A1982NB05700006
View details for PubMedID 7061240
Using a recently developed enzyme-linked immunosorbent assay, gene dosage effects were demonstrated for the Duffy, Ss, and Rh red blood cell antigen systems. We report a comparison by an antibody-binding assay of the relative amounts of Fya and Fyb antigens on FyaFya, FyaFyb, FybFyb, FyaFyx, and FyxFyx erthrocytes, and of s antigen on cells homozygous and heterozygous for s. The immunosorbent assay also was used to study Rh antigens, and data which had been obtained using radiolabelled antibodies were confirmed.
View details for Web of Science ID A1982PT76100003
View details for PubMedID 6815840
Splenocytes from mice immunized with purified, papain-solubilized HLA B27 antigen and/or human lymphocytes bearing the B27 specificity were fused with myeloma cell lines NSI or Sp2. The screening strategy employed a protein A binding assay in which various target cells were used. First, the hybrid cell supernatants were screened against B lymphocyte cell lines of known HLA specificities and the Daudi cell line, which does not express HLA-A, B, or C antigens. Second, a panel of PBLs were used as target cells. It was necessary to refine the protein A binding assay by preabsorbing the radiolabeled protein A with PBLs and by precoating the test wells with ovalbumin. Clones selected by these criteria were further tested by indirect immunoprecipitation and by inhibition of binding or microcytotoxicty to target call lines with purified HLA antigens or beta 2m. Forty-four clones were selected which showed varying degrees of specificity for allo- and nonallo-specific determinants and one clone was selected which was specific for beta 2m. Clone 27M1 which was previously shown to be specific or HLA-B27 as judged by conventional microcytotoxicity testing (Grumet et al., Lancet No. 8239, II:174, 1981) was compared with other clones using the above parameters for evaluation. Antibody from clone 27M1 showed preferential binding to B27 positive cell lines and PBLs, lesser binding to B7 positive target cells, and no binding to B40 positive target cells. Purified B27 antigen (papain) from two sources including the B27 target cell line, was able to inhibit the binding of antibodies from 27M1 to target cells. The extension of the protein A binding assay to PBLs has made it possible to more accurately quantitate the binding or inhibition of binding of antibodies to panels of PBLs.
View details for Web of Science ID A1982PA05300002
View details for PubMedID 6208127
The mechanism of the cytotoxic-negative, absorption-positive (CYNAP) phenomenon was studied using the model of the Bw49/Bw50 split of the BW21 antigen. Anti-Bw49 antibody bound 60% as well to Bw 50 as to Bw49 cells; however, even at a cytotoxic titer of 64 against Bw49 cells, the antibody was not cytotoxic to Bw50 Cells. At equal numbers of antibody molecules bound, the anti-Bw49 antibody activated C4 and C3, and induced lysis for Bw49 but not for the Bw50 cells. These data are consistent with a model in which different spatial orientations for shared epitopes can account for CYNAP reactivity for at least some selected Bw4/Bw6-associated splits of B locus antigens.
View details for Web of Science ID A1982PF84200004
View details for PubMedID 6176572
The mechanisms of enhanced antibody-mediated, complement-dependent cytotoxicity caused by proteolytic treatment of cells was studied in a model system based on HLA microlymphocytotoxicity methods. In this model, papain treatment of lymphocytes resulted in a) no change in antibody binding, b) a slight decrease in initial binding of C4, c) a marked increase in stability of cell-bound C4b, resulting in d) an increase of cell-bound C3, and e) no increase in lytic efficiency of the C5b-9 membrane attack complex. We conclude that the most important step in papain enhancement of lysis of lymphocytes is an increased stability of cell-bound C4b, possibly through decreased surface binding of C4-bp. This mechanism may be relevant to the pathologically increased lysis of cells occurring in patients with hereditary erythroblastic multinuclearity with a positive acidified-serum test (HEM-PAS).
View details for Web of Science ID A1982MZ49700062
View details for PubMedID 6915075
Human thymuses were examined by tissue section staining with antibodies specific for monomorphic and polymorphic HLA-A, B, C, and DR determinants. The principal cell type expressing high levels of HLA antigens has the distribution of epithelial cells. Immunoelectron microscopy confirmed their epithelial nature. As in the mouse, both medullary and cortical epithelial cells express high levels of class II (DR) antigens, a finding that is remarkable in that these antigens were originally thought to be restricted to lymphoid and accessory cells. Class I (A, B, and C) antigens are also present on thymic epithelial cells. They are easily detectable on medullary epithelial cells, but two distinct patterns of cortical staining were observed. One group of antibodies produced intense dendritic staining throughout the cortex; the other group produced only faint or no cortical dendritic staining at all. These different staining patterns do not correlate with known properties of the antibodies and thus appear to be due to intrinsic properties of the different A, B, and C antigens.
View details for Web of Science ID A1982PH53300002
View details for PubMedID 6956563
This study was directed at determining the major histocompatibility complex (MHC) antigens recognized by helper (Leu-3) and suppressor-cytotoxic (Leu-2) T lymphocyte subsets in man. These 2 subsets were isolated from peripheral blood with monoclonal antibodies and challenged in vitro with various stimuli. Only Leu-3 cells proliferated in response to autologous nonrosetting cells and soluble antigens, suggesting that helper but not suppressor-cytotoxic T cells recognize autologous HLA-DR antigen. Furthermore, only Leu-3 T cells responded to allogeneic DR antigen; this was shown in reactions between 2 siblings who were HLA-identical except for a single disparity at HLA-DR caused by a crossover event. Leu-3 cells activated in primary allogeneic mixed leukocyte reactions (MLR) responded equally in secondary allogeneic MLR to the priming cells and to cells that were identical at HLA-DR but discordant at HLA-A, B, and C to the priming cell. The antigens responsible for stimulating Leu-2 cells in allogeneic MLR were not identified, although the results are compatible with a role for HLA-A and B antigens and exclude a dominant role for HLA-DR. These data indicate that the helper and suppressor-cytotoxic T cell subsets in man respond differentially to MHC antigens in a manner analogous to the murine Lyt-2- 3- and Lyt-2+ 3+ populations.
View details for Web of Science ID A1981MP76800077
View details for PubMedID 6457863
Transfusion-acquired cytomegalovirus infections occurred in 13.5% of 74 infants of seronegative mothers who were exposed to one or more blood donors who had a CMV indirect hemagglutination titer of 1:8 or higher. None of 90 infants of seronegative mothers exposed only to donors with CMV IHA titers of less than 1:8 became infected. Ten of 41 (24%) infants of seronegative mothers who received more than 50 ml of packed red blood cells and who were exposed to at least one seropositive donor became infected. None of 23 infants of seronegative mothers who received this amount of blood but who were exposed only to seronegative donors became infected. Fatal or serious symptoms developed in 50% of the infected infants of seronegative mothers and in none of the 32 infected infants of seropositive mothers. Acquired CMV infections occurred in 15% of infants of seropositive mothers who were exposed to the red blood cells of seropositive donors and in 17.6% of infants of seropositive mothers exposed only to seronegative donors. Use of seronegative donors reduced the prevalence of excretion of CMV among hospitalized infants who were 4 weeks of age or older from 12.5 to 1.8% and eliminated acquired CMV infections in infants of seronegative mothers.
View details for Web of Science ID A1981LD52500029
View details for PubMedID 6257877
A new antibody, anti-Rh41, is described. This antibody was found prepartum in a multiparous, Caucasian female with no prior history of transfusions. Anti-Rh41 reacts with red blood cells from individuals with cis oriented C and e genes, but is nonreactive with cells possessing the products derived from individuals with cis-oriented Cw and e genes, a characteristic which distinguishes this antibody from anti-rhi. Furthermore, in contrast to anti-rhi, anti-Rh41 consistently reacts with Negro rh's (VS-positive) bloods. Also noteworthy in this study is the presence of an abnormal expression of chromosome 1 genetic markers in an offspring of the propositus.
View details for Web of Science ID A1981LK81400003
View details for PubMedID 6261428
Based on genotypic and phenotypic studies we have found strong linkage disequilibria in Caucasians among the genes HLA-Bw50, BfS1, and HLA-DR3 and/or -DR7. The relative disequilibria, which are among the highest described in man, are delta r (BfS1, DR7) = 0.51, delta r (Bw50, BfS1, DR7) = 0.36, delta r (Bw50, DR3 or 7) = 0.72, delta r (BfS1, DR3 or 7) = 0.91, delta r (Bw50, BfS1, DR3 or 7) = 0.73. The previously described high delta r (Bw50, BfS1) and delta r (Bw50, DR7) have also been confirmed. A B parallel DR crossover family is also presented that, together with previously reported recombinant families, confirms that the Bf locus resides between HLA-B and HLA-DR. These data suggest the existence of a supergene complex of Bw50, BfS1, DR3/7 (or MB2), and hypotheses to account for the observed disequilibria are discussed.
View details for Web of Science ID A1981MF43100005
View details for PubMedID 7263317
Phenotypic association and highly significant linkage disequilibria have been demonstrated for HLA-B18 and BfF1 and HLA-Bw50 and BfS1 alleles among Caucasians from Australia and the United States (San Francisco Bay area). The HLA-B18, BfF1 association appears to be associated with HLA-Aw30. It is possible that BfS1 arose as a mutation, after the evolutionary splitting of HLA-Bw21, on an HLA-Bw50 haplotype, and that BfF1 arose on an HLA-Aw30, B18 haplotype.
View details for Web of Science ID A1980KK94900004
View details for PubMedID 6911137
We have established a new transfusion program for an intensive care nursery which is based on crossmatching several infants to the same unit of type O Rh0(D) negative packed red blood cells, dividing the unit into quadpacks, and allowing multiple entry into each quadpack over a 24-hour period in the nursery. With this procedure, each donor unit can be used to provide multiple transfusions to four infants over a four-day period. Follow-up of transfusion recipients revealed that 20% had evidence of previous or ongoing CMB infection at 10 months of age, a prevalence comparable to that for transfused infants in other studies. We found no evidence for transmission of HB infection and a low risk of allosensitization to red cell and lymphocyte antigens.
View details for Web of Science ID A1980KQ18700025
View details for PubMedID 6253615
Definition of HLA-DR epitopes has been attempted by utilizing monoclonal antibody probes. Hybridoma antibodies L203 and L227, known to bind different epitopes on human Ia-like molecules, were tested for their ability to block cytotoxicity of monoclonal and allogeneic anti-DR antibodies. Monoclonal cytotoxic antibodies segregated into two groups: those more effectively blocked by L203, and those more effectively blocked by L227. Alloantisera also segregated into two groups, but according to their DR specificity. Anti-DR1, -2, and -3 alloantisera were effectively blocked by both L203 and L227, whereas anti-DR7, -w9, -w10, and MT1 alloantisera were not blocked by either. Blocking was not correlated with immunoglobulin class of the alloantibody and further definition of the mechanism of cytotoxicity blocking remains to be elucidated. Based on these data and prior binding and immunochemical studies with L203 and L227, a model is proposed in which the tertiary structure of each DR molecule, or complex of associated molecules on the cell surface, has two reference epitopes, one defined by L203, and another defined by L227. HLA-DR epitopes defined by the cytotoxic monoclonal or alloantibodies to the L203 or L227 epitope in order to begin epitope mapping or grouping.
View details for Web of Science ID A1980KS01300068
View details for PubMedID 6159421
A new microtechnique, C-FDA, for the in vitro detection of antiplatelet antibodies, is described. This technique is faster and simpler than either 51Cr thrombocytotoxicity or immunofluorescence (IF). C-FDA is more sensitive than 51Cr for all (anti-HLA, --P1A1, ABO, drug-related, and ITP-related) antibodies tested. Although IF was more sensitive for many types of antibodies, C-FDA was as good or better a clinical test method for all drug-related and isoimmune neonatal thrombocytopenia patient sera tested. Preliminary data also suggest that this method detects possible new non-HLA, non-ABO, nonP1A1 platelet antigens.
View details for Web of Science ID A1980KK94900009
View details for PubMedID 6167542
A new low incidence red cell antigen (Pe), and its identifying IgG antibody are described. The antigen is destroyed by enzymes and is absent from serum, urine, saliva, and platelets. The Pe gene frequency is estimated to be less than 0.0003, and the Pe locus does not appear to be linked to Rh or Fy, nor carried by the X or Y chromosomes. The clinical significance of this antibody remains to be determined.
View details for Web of Science ID A1979GX69700011
View details for PubMedID 462915
A nontoxic murine model has been developed in which sensitization to allogeneic platelet transfusions in adults can be prevented. This model is based upon allogeneic liver membrane (LM) induction of specific humoral tolerance to major histocompatibility alloantigens. In normal mice of the C3H background, syngeneic and H-2-incompatible congeneic platelets had a t1/2 = 15 +/- 3 hr; for mice of the C57BL background, the comparable t1/2 was 33 +/- 6 hr. Platelets of C3H background transfused into C57BL background recipients, and vice versa, had t1/22 midway between, 24 +/- 3 and 24 +/- 2 hr, respectively. These data suggest genetically determined variation in normal platelet survival. In passively immunized C3H background mice, the t1/2 was decreased to 10 +/- 2 hr. In C57BL background mice actively immunized i.p. with allogeneic lymphoid cells, t1/2 was decreased to 18 +/- 4 hr. When allogeneic LM was given concomitantly with the allogeneic cells, however, sensitization to foreign H-2 antigens was blocked, and survival of both C3H and C57BL background allogeneic platelets remained normal. These data demonstrate that in this free cell allograft system LM treatment is a safe and effective method for preventing adult sensitization to allogeneic platelets.
View details for Web of Science ID A1979HF66900004
View details for PubMedID 483363
A simple technique for red blood cell typing is described utilizing V-bottom microtiter trays with cell suspensions of 0.2%. This technique is many times more sensitive than tube or slide methods and can be used for direct agglutination, antiglobulin tests and mixed field agglutination.
View details for Web of Science ID A1978FP15500003
View details for PubMedID 684791
The genetics of murine susceptibility to Toxoplasma gondii was investigated in inbred mice and their F1 and F2 offspring. Among four strains of congenic mice of the B10 background, those with H-2a/a and H-2b/b genotypes were more susceptible than were those with H-2d/d and H-2k/k genotypes. Breeding studies utilizing three of these strains demonstrated linkage between the H-2a allele and greater susceptibility. These data suggest the existence of an H-2-linked gene affecting susceptibility to T. gondii. In challenge of recombinant inbred mice derived from C57Bl/6J (high susceptibility) and BALB/c (low susceptibility) strains, lines BE, BJ, and BK were more susceptible than lines BD, BG, BH, and BI. These data are consistent with the existence of a second disease susceptibility gene linked to the H-13 locus. F1 offspring of the C57B1/6J X B10.D2 mice were significantly less susceptible than either parent. This phenotypic complementary suggests the presence of more than one genetic mechanism of resistance to T. gondii. From these combined data, we conclude that (i) susceptibility to T. gondii in mice is affected by at least two genes, (ii) one of the genes is linked to the H-2 and one to the H-13 locus, and (iii) more than a single mechanism of resistance must be considered to explain the observed genetic controls of susceptibility.
View details for Web of Science ID A1978EN44600012
View details for PubMedID 631881
Available lymphocytotoxic antisera permitted the clear partition of the Bw21 antigen into two distinct components, Bw21.1 and Bw21.2. Bw21.1 is associated with W4 and is approximately twofold more frequent than the W6-associated Bw21.2. Cells of either Bw21 subtype were capable of absorbing specific anti-Bw21.1, anti-Bw21.2, and anti Bw21 (21.1 + 21.2) antibodies. Further, an F(ab')2 fragment prepared from an anti-Bw21.1 serum blocked cytotoxicity of anti-Bw21.1, anti-Bw21.2 and anti-Bw21 (21.1 + 21.2) sera. Based on the crossreactivity (by absorption) and blocking data, a model is proposed relating the Bw21 subtypes and the W4 and W6 antigens.
View details for Web of Science ID A1978EL67100004
View details for PubMedID 75587
View details for Web of Science ID A1977CX61600003
View details for Web of Science ID A1977EF33900036
An examination of HLA antigens in 72 unrelated Caucasian subjects with pernicious anaemia (PA) has revealed no significant association of any HLA-A or B genes with the disease. These data do not confirm the previous reports in the literature which had suggested an increased frequency of the B7 and/or A3 antigen among patients. In addition, the study of four families, each with two or more PA patients, does not support close linkage between disease susceptibility or autoantibody formation and the HLA locus. These data suggest that genes in or near the HLA region may not significantly affect susceptibility to PA.
View details for Web of Science ID A1977DH98800003
View details for PubMedID 871420
In patients with severe allergic reactions to plasma proteins it is possible to observe such reactions to even the small quantity of plasma contained in platelet concentrates. A platelet washing solution was designed, and platelet concentrates for four such patients were washed before infusion. Transfusion reactions were completely eliminated by the washing procedure. Platelet recovery was equivalent to that of unwashed platelets, and hemostatic effectiveness of the infused platelet concentrates was evidenced by abrupt cessation of bleeding episodes, including purpura and hematuria. Platelet washing represents a valuable, rapid and simple approach to the problem patient with thrombocytopenia and severe reactions to plasma proteins.
View details for Web of Science ID A1977CY47900006
View details for PubMedID 841669
The in vivo adjuvant effect of lipopolysaccharide (LPS) in mice was investigated with the soluble synthetic polypeptide antigen (T, G)-A--L, the antibody response to which is determined by the Ir-1A gene. With this specific antigen it can be demonstrated that the LPS adjuvant effect has the following modes of action: a) a T cell-dependent enhancement of primary and secondary IgM antibody response; b) a T cell-dependent enhancement of IgG secondary andibody response; and c) a T cell-dependent induction of switchover from IgM to IgG andibody in some strains of Ir-1A low responders. Although T cells are necessary for some aspects of the adjuvant effect, these data do not distinguish between a mechanism involving a direct interaction between LPS and T cells or a direct interaction of LPS and B cells with a general requirement for T cells for expression of IgG antibody.
View details for Web of Science ID A1976CH98200010
View details for PubMedID 1087242
Three high frequency reactive antisera (Kir, Oca, Mil) are described which, based on serologic and genetic characteristics, identify a set of apparently related antigens. The antibodies react only by indirect antiglobulin technique against both adult and cord red blood cells, are primarily IgG, are not complement dependent nor enhanced by papain pretreatment of red blood cells, are high titered but of low avidity, and are not neutralized by serum nor absorbed by platelets. The antisera are not identical with, but may be related to, the Kna antibody. Population data show reactivity frequencies of 99.8 per cent for Kir, 98.7 per cent for Oca, and 96.4 per cent for Mil. The four phenotypes found are Kir+, Oca+, Mil+; Kir+, Oca+, Mil- Kir+, Oca-, Mil+ and Kir-, Oca-, Mil-. The occurrence of five unrelated triple negative individuals is greater than would be expected by chance alone for three independent antigens. Family studies demonstrate that the triple negative phenotype appears to be a recessive trait not linked to the Fy or MNS loci, and the Mil-trait is not linked to ABO, Jk, or HLA. Clinical observations following infusion of incompatible blood and in vivo survival studies of 51Cr tagged red blood cells indicate that the antigens, though potent immunogens, are not clinically significant.
View details for Web of Science ID A1976CJ98700006
View details for PubMedID 982534
The immunological unreactive state occurring in (T,G)-A-L nonresponder mice after secondary antigen challenge was investigated. Syngeneic IgM anti-(T,G)-A-L antibody-containing plasma, transferred at the time of the time of primary challenge, induced persistent suppression of autologous specific antibody production. Removal of plasma IgM with goat anti-mu antisera removed the ability of the plasma to supress. The induction and maintenance of the suppressed state were not different in thymectomized or sham-thymectomized animals. Primed animals subjected to graft-vs-host reaction (GVHR) at the time of secondary challenge switched over to IgG production. Animals suppressed by passive antibody transfer reacted to GVHR, at the time of secondary challenge, with specific IgM but not IgG antibody production. Transfused normal spleen cells partially abrogated suppression only when (suppressed) hosts had been lethally irradiated. Spleen cells from antigen-plus-antibody suppressed donors, upon transfer to previously normal, syngeneic hosts, were less immunocompetent than spleen cells from untreated donors. These data are consistent with a model of IgM mediated, T cell-independent persistent suppression of humoral immunity.
View details for Web of Science ID A1976BM38300050
View details for PubMedID 3607
A group of Japanese were investigated for evidence of an association between Graves' disease and HL-A. Forty-four patients with the disease and 83 normal, unrelated random Japanese and Japanese-American controls were selected for study. The frequency of the W5 antigen among patients (57%) was significantly (P less than .0001) greater than among controls (20%). Of the 34 patients with abnormally elevated serum levels of anti-(thyroid) microsomal (anti-M) auto-antibodies, 56% had the W5 antigen. In contrast, of 48 control individuals tested for anti-M, only seven were seropositive and none (0%) of the seven had the W5 antigen. As expected the HL-A8 antigen was absent from this non-Caucasian population. These data demonstrate that the W5 antigen in Japanese, analogous to the HL-A8 antigen in Caucasians, is associated with Graves' disease but not with anti-M seropositivity in controls. The occurrence of different HL-A antigens in association with the same disease in different ethnic groups requires that the use of a major histocompatibility system antigen as a disease susceptibility marker must be confirmed for each ethnic group under study.
View details for Web of Science ID A1975BA78300008
View details for PubMedID 1243595
The purpose of our study was to determine why several of the glycolytic enzymes of the erythrocyte have the propensity for adhering to erythrocyte membranes while others do not. Two of these membrane-associated enzymes, glyceraldehyde phosphate dehydrogenase (GAPD) and phosphoglyceric kinase (PGK), have been shown to have controlling functions on intra-erythrocytic glycolysis and sodium-potassium transport. In order to test the hypothesis that the membrane-associated fraction of these enzymes consisted of isozymes with increased capacity to become membrane associated, the enzymes from cytosol or membrane sources were partially purified and characterized. Determination of molecular weight by gel filtration chromatography, measurement of certain kinetic parameters, and the curves relating to pH optimal activity indicated that there were no measurable differences between membrane and cytosol PGK and membrane and cytosol GAPD. It was possible to raise inhibitory antibodies to GAPD in certain species of mice, and these antibodies did not distinguish between GAPD isolated from either membrane or cytosol sources. GAPD was firmly bound to membranes and the membrane-associated fraction counted for approximately 60 per cent of total erythrocytic enzyme. Membrane-associated PGK was only loosely adherent, and accounted for only 1 per cent of the total erythrocytic enzyme. The reasons for membrane association have yet to be determined, but these inward facing membrane-associated enzymes appear to function by carrying the organization of the membrane into the cell interior.
View details for Web of Science ID A1975AB27400009
View details for PubMedID 235594
View details for Web of Science ID A1975BD73600002
In 101 white psoriatic patients, two histocompatibility (HL-A) specificities were significantly altered from expected values. The levels of W16 and W17 were found to be substantially increased, suggesting that persons with these antigens are at increased risk of having psoriasis. Clinically distinct patient groups were also observed. Antigens W16 or W17 or both were more prevalent in psoriatic patients who had extensive disease involvement, and patients with W17 antigen had an earlier age of onset as compared to patients with W16 antigen. In one family in this study, a linkage between psoriasis and a specific HL-A haplotype was also observed, further supporting the concept that the HL-A system may serve as a marker for genes affecting specific disease susceptibility.
View details for Web of Science ID A1975AL26300003
View details for PubMedID 1147630
View details for Web of Science ID A1974T658000008
View details for Web of Science ID 000247425800395
View details for Web of Science ID 000241751400121
View details for Web of Science ID 000227329000032
View details for Web of Science ID 000225127500186
View details for Web of Science ID 000174593901680
View details for Web of Science ID 000071717200022
View details for Web of Science ID A1996UM45500009
Canine narcolepsy is an animal model of the human disorder that is transmitted as a single autosomal recessive gene with full penetrance (canarc-1) in Dobermans and Labradors. In previous experiments, we have identified a very tight linkage marker for canarc-1. This marker, a 0.85-kb band cross reacting with a human mu-switch Heavy-Chain Immunoglobulin probe (maximum logarithm of odds [LOD] score Zmax = 10.8 at 0% recombination), has now been cloned and sequenced. The gene, composed of GC rich repeats, is 75% homologous to the human mu-switch gene and is similar in organization to immunoglobulin switch genes. Curiously, however, this mu-switchlike segment appears to be unlinked with other switchlike polymorphisms detected at high stringency with the human mu-switch probe. Because in most animal species all switch genes are located within 300-500 kb and show tight linkage in families, this result suggests two possible hypotheses: 1) Our 0.85 kb is a true immunoglobulin switch segment, but the map of the canine Variable Heavy-Chain loci is organized in unlinked clusters, or 2) our 0.85-kb segment is not an immunoglobulin switch segment and is located elsewhere in the genome in all species. We are now using chromosome walking and Yeast Artificial Chromosome Cloning techniques, together with corresponding studies in humans to identify the pathological gene.
View details for Web of Science ID A1994QD08100014
View details for PubMedID 7701203
In the present study, we tested 19 Caucasian and 28 Black American narcoleptics for the presence of the human leucocyte antigen (HLA) DQB1*0602 and DQA1*0102 (DQ1) genes using a specific polymerase chain reaction (PCR)-oligotyping technique. A similar technique was also used to identify DRB1*1501 and DRB1*1503 (DR2). Results indicate that all but one Caucasian patient (previously identified) were DRB1*1501 (DR2) and DQB1*0602/DQA1*102 (DQ1) positive. In Black Americans, however, DRB1*1501 (DR2) was a poor marker for narcolepsy. Only 75% of patients were DR2 positive, most of them being DRB1*1503, but not DRB1*1501 positive. DQB1*0602 was found in all but one Black narcoleptic patient. The clinical and polygraphic results for this patient were typical, thus confirming the existence of a rare, but genuine form of DQB1*0602 negative narcolepsy. These results demonstrate that DQB1*0602/DQA1*0102 is the best marker for narcolepsy across all ethnic groups.
View details for Web of Science ID A1994QD08100013
View details for PubMedID 7701202
A soluble HLA-B7 molecule, designated sB7 and generated by genetically engineering the B7 gene to remove the transmembrane and cytoplasmic domains, was tested as a tolerogen. Supernatants from cultures of C1R cells transfected with the gene for sB7 were harvested and concentrated, as were control supernatants. From days -17 to -1, C57Bl/6 mice were pretreated with a total of 11 intraperitoneal doses of 1.0 microgram each of sB7 or appropriate control supernatant, and then were challenged intraperitoneally on each of days 0, 7, and 14 with 10(6) C1R-B7 cells (expressing surface HLA-B7). Antibody kinetics revealed (1) anti-B7 was not induced after sB7 pretreatment; (2) the anti-B7 response of sB7-pretreated mice was marginal and of apparent low avidity compared with the brisk anti-B7 response of control mice; (3) none of the mice made antibody to a control HLA antigen, A24; (4) all mice made strong antibody responses to the non-B7 surface antigens of C1R; (5) free sB7 did not appear in the blood of the treated mice; and (6) all mice appeared to be generally healthy. These data show soluble B7 antigen is not immunogenic and appears to specifically block humoral immune response to cell membrane-bound HLA-B7 in a nontoxic manner.
View details for Web of Science ID A1994PA72000012
View details for PubMedID 7960967
A soluble, secreted form of HLA-B7 was engineered by replacing the exons encoding the transmembrane and cytoplasmic domains of the B7 gene with a CI. The modified gene, gsB7, transfected into J27.2 or C1R cell lines, produced a secreted protein, sB7, serologically recognized as B7. Size fractionation showed one species of sB7 at the approximately 55 kD expected for an sB7 alpha-chain-beta 2m heteroduplex, and another at approximately 120 kD which had the same constituent chains and was a dimer of the 55-kD species. Dimer formation appeared to be related to protein concentration but not to disulfide bridging. The sB7 heavy chain on SDS-PAGE showed a doublet at approximately 39 and approximately 42 kD; enzyme analysis indicated that the two bands differed only by a carboxyl terminal polypeptide. Analysis of gsB7 transfectants' mRNA by Northern blots and PCR revealed message fully spliced or with retained CI, accounting for the 39- and 42-kD bands, respectively, and apparently untranslated message with I3 retained. sB7 was not detectable on the surface of gsB7 transfectants by CTLs, nor did it inhibit those CTLs. Production of the sB7 protein provides a ready, consistent source of soluble class I antigen for further study, including test materials for tolerogenicity studies in animal models.
View details for Web of Science ID A1994PA72000013
View details for PubMedID 7960968
View details for Web of Science ID A1984SJ72502205
View details for Web of Science ID A1980KN83000085
View details for Web of Science ID A1978FU23000094
View details for Web of Science ID A1977DA32900989