Dr. Behr graduated from University of Nevada-Reno with honors and earned his Ph.D. with emphasis in Reproductive Physiology in 1991, board certified as High Complexity Laboratory Director in 1994, and board certified as Embryology Laboratory Director in 2003.

Dr. Behr is a nationally and internationally renowned clinical and scientific leader in the research and advances of human reproduction. As a world-renown scientist and lecturer, he is highly sought after to chair scientific programs and conferences, was the first non M.D. President of the Pacific Coast Reproductive Society, and has been appointed as program chair of several professional meetings. He is in the forefront of clinical and scientific advances in reproduction, nationally and internationally as an entrepreneur and has founded several fertility related businesses. Dr. Behr developed a culture medium for embryo culture to the blastocyst stage, which improved pregnancy rates, implantation rates and reduced the risks of multiple gestation in IVF. Dr. Behr has been widely recognized for his research. He has published over 90 peer-reviewed publications, and has authored over 170 abstracts and 14 book chapters.

Dr. Behr enjoys racing cars, golfing, barbequing, and spending time with his family.

Professional Education

  • H.C.L.D., Am. Bd. of Bioanalysists
  • Ph.D., Univ. of Nevada-Reno, Reno, NV, Biology (1991)

Research & Scholarship

Current Research and Scholarly Interests

Development of improved embryo culture conditions in vitro. Blastocyst cultures. Embryo metabolism in vitro. Embryo maternal dialogue. Clinical application and integration of extended embryo culture systems. Monozygotic twinning. Prevention of multiple pregnancy. Sperm motility enhancers. Fluorescent and non-fluorescent markers of sperm morphology and viablility.Oocyte cryopreservation. Fertility preservation. Improving IVF outcome.

Clinical Trials

  • Day of Embryo Transfer for Patients Undergoing In Vitro Fertilization Not Recruiting

    We are examining whether pregnancy rates differ based on day of embryo transfer in patients who replace all available embryos after an In vitro Fertilization (IVF) cycle. Patients must be undergoing IVF treatment at Stanford University and patients will not receive compensation for their participation (no medical costs covered or patient payment for participation).

    Stanford is currently not accepting patients for this trial. For more information, please contact Lora Shahine, (650) 498 - 7911.

    View full details


2017-18 Courses


All Publications

  • Paternal Age and Total Motile Sperm Parameters Are Not Correlated with Embryo Aneuploidy Rates. 62nd Annual Pacific Coast Reproductive Society Conference Kort, J., Zhao, Q., Behr, B.
  • A comparison of aneuploidy rates between Asian and Caucasian patients. 30th Annual European Society of Human Reproduction and Embryology Meeting Behr, B., Smotrich, D., Gaona, M., Hamic , A., Wang, X., Kort, J.
  • Assessing the Effects of Vitrification on Embryo Viability using Label-Free Imaging. 70th Annual American Society for Reproductive Medicine Conference Zarnescu, L., Abeyta, M., Baer, T., Behr, B., Ellerbee, A.
  • Mechanical Biomarkers of Oocyte Maturation. 70th Annual American Society for Reproductive Medicine Conference Zarnescu, L., Han, J., Behr, B., Reijo Pera, R., Camarillo, D.
  • Is there an Association between Gonadotropin Dosing and Aneuploidy Rates? 70th Annual American Society for Reproductive Medicine Conference Kort, J., Zolton, J., Behr, B., Lathi, R., Baker, V.
  • Shared Oocyte Donation Program Produces High Clinical Pregnancy Rate-A Three Year Follow Up Study. 63rd Annual Pacific Coast Reproductive Society Conference Khoury, C., Frederick, J., Coffler, M., Sills, E., Behr, B., Potter, D.
  • Non-invasive Technology Combining Time-lapse Imaging and Statistical Modeling: Bringing Automation into the Lab to Improve Blastocyst Selection. 71st Annual American Society for Reproductive Medicine Conference Behr, B., Tan, L., Conaghan , J., Liebermann, J., Bartolucci, A., Chen, A.
  • Computer-automated Time-lapse Test Results are Predictive of Pregnancy Following Blastocyst Transfer. 71st Annual American Society for Reproductive Medicine Conference Behr, B., Beltsos, A., Yee, B., Kingsland , C., Benadiva, C., Liebermann, J.
  • Blastocyst Implantation is Correlated with Outputs from Automated Time-lapse Analysis by the EEVA Test. 71st Annual American Society for Reproductive Medicine Conference Liebermann , J., Bartolucci, A., Troup , S., Wagner Coughlin, C., Yee, B., Behr, B.
  • The relationship between a man’s somatic health and ART outcomes. 71st Annual American Society for Reproductive Medicine Conference Eisenberg, M., Li, S., Behr, B., Nakajima, S., Baker, V.
  • Microfluidic sperm sorting device for selection of functional human sperm for IUI application. 64th Annual Pacific Coast Reproductive Society Conference Chinnasamy, T., Behr, B., Demirci, U.
  • Does euploid embryo ranking by trophectoderm cell mitochondrial DNA content correspond with ranking by blastocyst morphology within an individual patient’s cohort of blastocysts? 72nd Annual American Society for Reproductive Medicine Conference Kort, J., Lathi, R., Behr, B.
  • Pilot clinical study to predict IVF outcomes using embryo mechanical parameters. 72nd Annual American Society for Reproductive Medicine Conference Yanez, L., Sedan, O., Baker, V., Behr, B., Camarillo, D.
  • Concurrent genome and transcriptome analysis from a single trophectoderm biopsy. (Prize Paper Award.) 66th Annual Pacific Coast Reproductive Society Conference Chiang, H., Kort, J., Behr, B., Snyder, M.
  • Preliminary study to investigate telomerase reverse transcriptase expression among human cumulus cells. 66th Annual Pacific Coast Reproductive Society Conference Kort, J., Garbuzov, A., Artandi , S., Behr, B.
  • The effect of embryo biopsy on perinatal outcomes: An analysis of SART CORS. 73rd Annual American Society for Reproductive Medicine Conference Kort, J., Behr, B., Wantman, E., Baker, V.
  • Oocyte telomerase levels correlate with blastocyst development. 73rd Annual American Society for Reproductive Medicine Conference Kort, J., Garbuzov, A., Arand, A., Behr, B., Artand, S.
  • Do embryo time-lapse parameters predict euploid embryo transfer outcomes? 73rd Annual American Society for Reproductive Medicine Conference Drejza, M., Kort, J., Behr, B.
  • A magnetic levitation platform for the isolation of mature sperm from TESE/TESA samples. 66th Annual Pacific Coast Reproductive Society Conference Durmus, G., Gupta, R., Badamjav, O., Reddy, V., Eisenberg, M., Behr, B., Demirci, U.
  • Does euploid embryo ranking by trophectoderm cell mitochondrial DNA content correspond with ranking by blastocyst morphology within an individual patient’s cohort of blastocysts? 72nd Annual American Society for Reproductive Medicine Conference Kort, J., Lathi, R., Behr, B.
  • Racial Variation in Semen Quality at Fertility Evaluation. Urology Khandwala, Y. S., Zhang, C. A., Li, S., Behr, B., Guo, D., Eisenberg, M. L. 2017


    To identify racial differences in semen quality among men living in the same geographic area and seeking fertility evaluation.Men obtaining a semen analysis for infertility evaluation or treatment between 2012 and 2016 at a single center were identified, and demographic data including height, weight, body mass index (BMI), and age were described. Mean semen parameters and the proportions of men with suboptimal parameters based on the World Health Organization's fifth edition criteria were also compared based on race. Multivariable regression analysis was conducted incorporating age, BMI, and year of evaluation. Further subanalyses based on BMI were subsequently performed.White men produced greater volumes of semen on average; however, Asian men had higher sperm concentrations and total sperm count. A lower proportion of Asian men compared to white men had semen quality in the suboptimal range for most semen parameters, whereas a higher proportion of white men were found to have azoospermia. Stratification by BMI groups attenuated the observed differences between whites and Asians, yet Asian male semen quality remained higher.Among men evaluated for infertility at a single center, Asians had lower volume but higher sperm concentrations compared with whites, which was influenced by differences in azoospermia prevalence. Although anthropometric and demographic factors attenuated the differences, even after adjustment, the contrasts remained. Our study suggests that racial differences exist in semen quality at the time of infertility evaluation.

    View details for DOI 10.1016/j.urology.2017.03.064

    View details for PubMedID 28522219

  • Functional topography of the fully grown human oocyte EUROPEAN JOURNAL OF HISTOCHEMISTRY Monti, M., Calligaro, A., Behr, B., Pera, R. R., Redi, C. A., Wossidlo, M. 2017; 61 (1): 32-35
  • Hypertension and Male Fertility. The world journal of men's health Guo, D., Li, S., Behr, B., Eisenberg, M. L. 2017; 35 (2): 59–64


    As the age of paternity rises in the developed world, issues of chronic disease may affect prospective fathers. Given the high prevalence of hypertension, researchers have begun to explore the relationship between hypertensive disease and male fertility. The current literature suggests an association between hypertension and semen quality. The use of various antihypertensive medications has also been linked to impaired semen parameters, making it difficult to discern whether the association exists with hypertension or its treatment. Further investigation is warranted to determine whether the observed associations are causal.

    View details for DOI 10.5534/wjmh.2017.35.2.59

    View details for PubMedID 28868816

    View details for PubMedCentralID PMC5583372

  • Biomechanics and developmental potential of oocytes and embryos. Fertility and sterility Kort, J., Behr, B. 2017; 108 (5): 738–41


    The high incidence of multiple embryo transfers is evidence of the need for better methods of embryo selection. Additionally, methods to determine the reproductive competence of unfertilized oocytes are critically needed to inform the growing population of patients undergoing fertility preservation. The ideal method of oocyte and embryo selection would be noninvasive, inexpensive, and able to be incorporated into embryology workflow with minimal disruption. Methods to assess the biomechanical properties of cells offer many of these traits, and there is a growing body of evidence in multiple cell types demonstrating the biomechanical properties of cells are reflective of a cell's intrinsic health. The associations with these properties are not mere coincidence, as many of the biomechanical properties are critical to cellular function. The biomechanical properties of oocytes and embryos undergo a dynamic, characteristic transformation from oocyte maturation through blastocyst formation, lending itself to biomechanical assessment. Many of the assessments made by embryologists, from ease of microinjection during intracytoplasmic sperm injection to degree of blastocyst expansion, are direct proxies for cellular biomechanics. Newer, objective and quantitative methods of biomechanical assessment are being applied to oocyte and embryo selection, with early use supporting their application in assisted reproduction.

    View details for DOI 10.1016/j.fertnstert.2017.09.016

    View details for PubMedID 28987788

  • Guidance and Self-Sorting of Active Swimmers: 3D Periodic Arrays Increase Persistence Length of Human Sperm Selecting for the Fittest. Adv Sci. Chinnasamy, T., Kingsley, J., Inci, F., Turek, P., Rosen, M., Behr, B., Tüzel , E., Demirci, U. 2017: 1-13
  • Relationship between paternal somatic health and assisted reproductive technology outcomes. Fertility and sterility Eisenberg, M. L., Li, S., Wise, L. A., Lynch, C. D., Nakajima, S., Meyers, S. A., Behr, B., Baker, V. L. 2016; 106 (3): 559-565


    To study the association between paternal medical comorbidities and the outcomes of assisted reproductive technology (ART).Retrospective cohort study.Academic reproductive medicine center.We analyzed fresh ART cycles uszing freshly ejaculated sperm from the male partner of couples undergoing ART cycles from 2004 until 2014. We recorded patient and partner demographic characteristics. The cohort was linked to hospital billing data to obtain information on selected male partners' comorbidities identified using ICD-9-CM codes.None.Fertilization, clinical pregnancy, miscarriage, implantation, and live-birth rates as well as birth weights and gestational ages.In all, we identified 2,690 men who underwent 5,037 fresh ART cycles. Twenty-seven percent of men had at least one medical diagnosis. Men with nervous system diseases had on average lower pregnancy rates (23% vs. 30%) and live-birth rates (15% vs. 23%) than men without nervous system diseases. Lower fertilization rates were also observed among men with respiratory diseases (61% vs. 64%) and musculoskeletal diseases (61% vs. 64%) relative to those without these diseases. In addition, men with diseases of the endocrine system had smaller children (2,970 vs. 3,210 g) than men without such diseases. Finally, men with mental disorders had children born at an earlier gestational age (36.5 vs. 38.0 weeks).The current report identified a possible relationship between a man's health history and IVF outcomes. As these are potentially modifiable factors, further research should determine whether treatment for men's health conditions may improve or impair IVF outcomes.

    View details for DOI 10.1016/j.fertnstert.2016.04.037

    View details for PubMedID 27179785

  • Human Embryonic Stem Cell Lines with Lesions in FOXP3 and NF1 PLOS ONE Zhu, H., Behr, B., Reddy, V. V., Hughes, M., Pan, Y., Baker, J. 2016; 11 (3)
  • Human oocyte developmental potential is predicted by mechanical properties within hours after fertilization. Nature communications Yanez, L. Z., Han, J., Behr, B. B., Reijo Pera, R. A., Camarillo, D. B. 2016; 7: 10809-?


    The causes of embryonic arrest during pre-implantation development are poorly understood. Attempts to correlate patterns of oocyte gene expression with successful embryo development have been hampered by the lack of reliable and nondestructive predictors of viability at such an early stage. Here we report that zygote viscoelastic properties can predict blastocyst formation in humans and mice within hours after fertilization, with >90% precision, 95% specificity and 75% sensitivity. We demonstrate that there are significant differences between the transcriptomes of viable and non-viable zygotes, especially in expression of genes important for oocyte maturation. In addition, we show that low-quality oocytes may undergo insufficient cortical granule release and zona-hardening, causing altered mechanics after fertilization. Our results suggest that embryo potential is largely determined by the quality and maturation of the oocyte before fertilization, and can be predicted through a minimally invasive mechanical measurement at the zygote stage.

    View details for DOI 10.1038/ncomms10809

    View details for PubMedID 26904963

  • Label-free characterization of vitrification-induced morphology changes in single-cell embryos with full-field optical coherence tomography. Journal of biomedical optics Zarnescu, L., Leung, M. C., Abeyta, M., Sudkamp, H., Baer, T., Behr, B., Ellerbee, A. K. 2015; 20 (9): 96004-?

    View details for DOI 10.1117/1.JBO.20.9.096004

    View details for PubMedID 26334977

  • Increased body mass index negatively impacts blastocyst formation rate in normal responders undergoing in vitro fertilization JOURNAL OF ASSISTED REPRODUCTION AND GENETICS Comstock, I. A., Kim, S., Behr, B., Lathi, R. B. 2015; 32 (9): 1299-1304

    View details for DOI 10.1007/s10815-015-0515-1

    View details for Web of Science ID 000362519600002

    View details for PubMedID 26109331

  • Aneuploidy rates and blastocyst formation after biopsy of morulae and early blastocysts on day 5. Journal of assisted reproduction and genetics Kort, J. D., Lathi, R. B., Brookfield, K., Baker, V. L., Zhao, Q., Behr, B. R. 2015; 32 (6): 925-930


    Studies have demonstrated high implantation rates after trophectoderm biopsy of day 5 expanded blastocysts. However, biopsy of cleavage stage embryos may adversely affect embryo development and implantation. No studies have assessed the utility of day 5 morulae and early blastocyst biopsy. This study sought to better understand these slower embryos' aneuploidy rates and implantation potential.This was a retrospective review of all autologous IVF cycles utilizing PGS at a single academic infertility center.The biopsy of day 5 morulae and early blastocysts provided 22 % additional euploid blastocysts available for fresh day 6 transfer compared to day 5 biopsy of only expanded blastocysts. Aneuploidy did correlate with embryo stage on day 5, even after controlling for maternal age, with 16 % of morulae and 35 % of blastocysts being euploid. The majority (83 %) of euploid morulae progressed to the blastocyst stage by day 6. Experience transferring slower developing embryos is limited, but preliminary pregnancy and implantation rates appear similar to euploid embryos biopsied as expanded blastocysts.The biopsy of all non-arrested embryos on day 5 provides genetic information for all blastocysts on day 6, increasing the pool of euploid blastocysts available for fresh transfer and avoiding the need to cryopreserve developmentally competent embryos without genetic information.

    View details for DOI 10.1007/s10815-015-0475-5

    View details for PubMedID 25921084

  • Relationship between semen production and medical comorbidity FERTILITY AND STERILITY Eisenberg, M. L., Li, S., Behr, B., Pera, R. R., Cullen, M. R. 2015; 103 (1): 66-71


    To study the relationship between semen quality and current health status in a cohort of men evaluated for infertility.Cross-sectional study.Fertility clinic.Nine thousand three hundred eighty-seven men evaluated for infertility between 1994 and 2011.None.Charlson comorbidity index, medical diagnoses by organ system.At the time of evaluation, 9,387 men with a mean age of 38 years had semen data available. Of these men, 44% had at least one medical diagnosis unrelated to infertility. When stratifying the cohort by the Charlson comorbidity index (CCI), differences in all measured semen parameters were identified. Men with a higher CCI had lower semen volume, concentration, motility, total sperm count, and morphology scores. In addition, men with diseases of the endocrine, circulatory, genitourinary, and skin diseases all showed significantly higher rates of semen abnormalities. Upon closer examination of diseases of the circulatory system, men with hypertensive disease, peripheral vascular and cerebrovascular disease, and nonischemic heart disease all displayed higher rates of semen abnormalities.The current report identified a relationship between medical comorbidites and male semen production. Although genetics help guide a man's sperm production, his current condition and health play an important role.

    View details for DOI 10.1016/j.fertnstert.2014.10.017

    View details for Web of Science ID 000346911400015

    View details for PubMedID 25497466

  • Comparison of epigenetic mediator expression and function in mouse and human embryonic blastomeres HUMAN MOLECULAR GENETICS Chavez, S. L., McElroy, S. L., Bossert, N. L., De Jonge, C. J., Vera Rodriguez, M., Leong, D. E., Behr, B., Westphal, L. M., Pera, R. A. 2014; 23 (18): 4970-4984


    A map of human embryo development that combines imaging, molecular, genetic and epigenetic data for comparisons to other species and across pathologies would be greatly beneficial for basic science and clinical applications. Here, we compared mRNA and protein expression of key mediators of DNA methylation and histone modifications between mouse and human embryos, embryos from fertile/infertile couples, and following growth factor supplementation. We observed that individual mouse and human embryos are characterized by similarities and distinct differences in DNA methylation and histone modification patterns especially at the single-cell level. In particular, while mouse embryos first exhibited sub-compartmentalization of different histone modifications between blastomeres at the morula stage and cell sub-populations in blastocysts, differential histone modification expression was detected between blastomeres earlier in human embryos at the four- to eight-cell stage. Likewise, differences in epigenetic mediator expression were also observed between embryos from fertile and infertile couples, which were largely equalized in response to growth factor supplementation, suggesting that select growth factors might prevent alterations in epigenetic profiles during prolonged embryo culture. Finally, we determined that reduced expression via morpholino technologies of a single histone-modifying enzyme, Rps6ka4/Msk2, resulted in cleavage-stage arrest as assessed by time-lapse imaging and was associated with aneuploidy generation. Taken together, data document differences in epigenetic patterns between species with implications for fertility and suggest functional roles for individual epigenetic factors during pre-implantation development.

    View details for DOI 10.1093/hmg/ddu212

    View details for Web of Science ID 000343184400018

    View details for PubMedID 24821703

  • The role of serum testosterone in early pregnancy outcome: a comparison in women with and without polycystic ovary syndrome. Journal of obstetrics and gynaecology Canada : JOGC = Journal d'obstétrique et gynécologie du Canada : JOGC Lathi, R. B., Dahan, M. H., Reynolds-May, M. F., Milki, A. A., Behr, B., Westphal, L. M. 2014; 36 (9): 811-816


    Hyperandrogenic conditions in women are associated with increased rates of miscarriage. However, the specific role of maternal testosterone in early pregnancy and its association with pregnancy outcome is unknown. The purpose of this study was to compare serum testosterone levels during early pregnancy in women with and without polycystic ovary syndrome (PCOS) who either had successful pregnancies or miscarried.We collected serum samples from women attending a university-based fertility centre at the time of their first positive serum beta human chorionic gonadotropin pregnancy test. The samples were subsequently assayed for total testosterone level. We used logistical regression modelling to control for PCOS diagnosis, BMI, and age.Total testosterone levels were available for 346 pregnancies, including 286 successful pregnancies and 78 first trimester miscarriages. We found no difference in total testosterone levels between women who subsequently had an ongoing pregnancy (mean concentration 3.6 ± 2.6 nmol/L) and women with a miscarriage (mean 3.6 ± 2.4 nmol/L). Using the Rotterdam criteria to identify women with PCOS, we also found no differences in serum testosterone between women who had ongoing pregnancies or miscarriages, either with PCOS (P = 0.176) or without PCOS (P = 0.561).Our findings show that early pregnancy testosterone levels do not predict pregnancy outcome, and they call into question the role of testosterone in causing miscarriage in populations of women with PCOS. Further research is needed to elucidate the normal progression of testosterone levels during pregnancy and to investigate further the relationship between PCOS and miscarriage.

    View details for PubMedID 25222360

  • The evaluation of pre and post processing semen analysis parameters at the time of intrauterine insemination in couples diagnosed with male factor infertility and pregnancy rates based on stimulation agent. A retrospective cohort study EUROPEAN JOURNAL OF OBSTETRICS & GYNECOLOGY AND REPRODUCTIVE BIOLOGY Luco, S. M., Agbo, C., Behr, B., Dahan, M. H. 2014; 179: 159-162


    To identify pre or post processing semen analysis parameters that may be predictive of successful pregnancy in couples with male factor infertility undergoing intra uterine insemination (IUI). To evaluate the pregnancy rate based on ovulation inducing agent in couples with male factor infertility per the 2010 world health organization criteria treated with IUI.This retrospective study was performed at Stanford University medical center. All couples with male factor infertility fitting inclusion criteria were included over a 2 year period of time. 147 couples with male factor infertility were included and 356 IUIs were analyzed. All subjects in this study had Kruger strict analysis >4% normal forms. Logistic regression analysis was used to control for confounding effects and multiplicity.The overall pregnancy rate was 5.3%. No parameter in either the pre or post analysis predicted pregnancy. Furthermore, it was found that natural cycle and letrazole treatment had similar pregnancy rates (3% and 3%) p=ns. Similar outcomes were also observed between clomiphene citrate and gonadotropin stimulated cycles (7.5% and 6.0%) p=ns.Total motile sperm count which has been found to be a predictor of pregnancy when evaluated in isolation, may be due to a confounding effect. These low pregnancy rates should be considered when deciding whether to suggest IUI and when selecting a protocol for ovulation induction for couples with male factor infertility.

    View details for DOI 10.1016/j.ejogrb.2014.05.003

    View details for Web of Science ID 000340318200030

    View details for PubMedID 24965998

  • Semen quality, infertility and mortality in the USA HUMAN REPRODUCTION Eisenberg, M. L., Li, S., Behr, B., Cullen, M. R., Galusha, D., Lamb, D. J., Lipshultz, L. I. 2014; 29 (7): 1567-1574
  • Atypical embryo phenotypes identified by time-lapse microscopy: high prevalence and association with embryo development FERTILITY AND STERILITY Wirka, K. A., Chen, A. A., Conaghan, J., Ivani, K., Gvakharia, M., Behr, B., Suraj, V., Tan, L., Shen, S. 2014; 101 (6): 1637-U495


    To characterize atypical dynamic embryo phenotypes identified by time-lapse microscopy, evaluate their prevalence, and determine their association with embryo development.Retrospective multicenter cohort study.Five IVF clinics in the United States.Sixty-seven women undergoing IVF treatment with 651 embryos.Embryo videos were retrospectively analyzed for atypical phenotypes.Identification of four groups of atypical embryo phenotypes: abnormal syngamy (AS), abnormal first cytokinesis (A1(cyt)), abnormal cleavage (AC), and chaotic cleavage (CC). Prevalence and association with embryo morphology and development potential were evaluated.A high prevalence of atypical phenotypes was observed among embryos: AS 25.1% (163/649), A1(cyt) 31.0% (195/639), AC 18% (115/639) and CC 15% (96/639). A high percentage of embryos with atypical phenotype(s) had good quality on day 3 (overall grade good or fair): AS 78.6% (70/89); A1(cyt) 79.7% (94/119), AC 86.4% (70/81), and CC 35.2% (19/54), but the blastocyst formation rates for these embryos were significantly lower compared with their respective control groups: AS 21.5% vs. 44.9%, A1(cyt) 21.7% vs. 44.6%, AC 11.7% vs. 43.1%, and CC 14.0% vs. 42.3%.Embryos exhibiting atypical phenotypes are highly prevalent in human embryos and show significantly lower developmental potential than control embryos.NCT01369446.

    View details for DOI 10.1016/j.fertnstert.2014.02.050

    View details for Web of Science ID 000337364300033

    View details for PubMedID 24726214

  • Testosterone Changes Bladder and Kidney Structure in Juvenile Male Rats JOURNAL OF UROLOGY Shortliffe, L. M., Ye, Y., Behr, B., Wang, B. 2014; 191 (6): 1913-1919


    Testosterone affects male development, maturation and aging but limited data exist on testosterone effects on the juvenile genitourinary system. We hypothesized that testosterone has bladder and kidney developmental effects, and investigated this in juvenile male rats.To examine the testosterone effect 21-day-old prepubertal male Wistar rats were divided into 3 groups of 12 each, including sham orchiectomy as controls, and bilateral orchiectomy with vehicle and bilateral orchiectomy with testosterone. Starting at age 28 days (week 0) testosterone enanthate (5 mg/100 gm) or vehicle was injected weekly. Testosterone was measured at study week 0 before injection, and at weeks 1, 6 and 16. Whole bladders and kidneys were evaluated for androgen receptor, bladder collagen-to-smooth muscle ratio, and renal morphometry and immunohistochemistry.Testosterone was not detectable at week 0 in all groups. It remained undetectable at weeks 1, 6 and 16 in the orchiectomy plus vehicle group. Testosterone levels were physiological in controls and rats with orchiectomy plus testosterone but levels were higher in the latter than in the former group. Rats with orchiectomy plus testosterone had increased bladder-to-body and kidney-to-body weight ratios (p<0.01 and <0.05, respectively), and decreased collagen-to-smooth muscle ratio than the orchiectomy plus vehicle and control groups. Rats with orchiectomy plus testosterone had a lower renal total glomerular count (p<0.01) but increased androgen receptor density.In juvenile male rats testosterone was associated with increased bladder and renal mass, and increased bladder smooth muscle. Testosterone associated kidneys also appeared to have fewer but larger glomeruli. These data support an important role for sex hormones in structural and functional development of the bladder and kidney.

    View details for DOI 10.1016/j.juro.2014.01.012

    View details for Web of Science ID 000336531100104

    View details for PubMedID 24518779

  • Morphological Assessment of Embryo Viability SEMINARS IN REPRODUCTIVE MEDICINE Abeyta, M., Behr, B. 2014; 32 (2): 114-126


    Morphological assessment is discussed in the context of significant literature at all stages of in vitro development, beginning with the oocyte and culminating at the blastocyst stage. Current evidence is used to debate the inclusion of commonly observed morphological features in grading schemes. The biological rationale behind observed phenomena such as multinucleation and fragmentation are also explored. Current limitations as well as technological advancements that increase our ability to assess viability are highlighted. Particular attention is paid to the relationship between developmental timing and assessment schemes. Failure to standardize assessment timing and inclusion criteria is glaring weaknesses of the literature that currently make consensus unattainable. Mounting evidence suggests that the future of static assessment is very likely to be influenced by information gathered from preimplantation genetic screening and other invasive techniques as well as from continuous monitoring tools such as time lapse.

    View details for DOI 10.1055/s-0033-1363553

    View details for Web of Science ID 000331288800007

    View details for PubMedID 24515906

  • Assessment of imaging parameters correlated with the effects of cryopreservation on embryo development Conference on Optical Methods in Developmental Biology II Zarnescu, L., Abeyta, M., Baer, T. M., Behr, B., Ellerbee, A. K. SPIE-INT SOC OPTICAL ENGINEERING. 2014

    View details for DOI 10.1117/12.2040487

    View details for Web of Science ID 000336049000006

  • Embryo culture and selection: morphological criteria. Methods in molecular biology (Clifton, N.J.) Hegde, A., Behr, B. 2014; 1154: 501-532


    In this chapter, we have outlined the various morphological criteria for selection of the best embryo at each important milestone encountered in the progress from the oocyte to the blastocyst. As Gerris et al. stated, a combination of one, two, or even three selection points should lead to a more accurate selection of the best embryo, as no one criterion is better than the other. An embryo that fails to meet the entire set of selection criteria must be avoided as culture cannot correct an impaired embryo.

    View details for DOI 10.1007/978-1-4939-0659-8_23

    View details for PubMedID 24782025

  • EED and KDM6B Coordinate the First Mammalian Cell Lineage Commitment To Ensure Embryo Implantation MOLECULAR AND CELLULAR BIOLOGY Saha, B., Home, P., Ray, S., Larson, M., Paul, A., Rajendran, G., Behr, B., Paul, S. 2013; 33 (14): 2691-2705


    The first mammalian cell lineage commitment is the formation of the trophectoderm (TE) and the inner cell mass (ICM) lineages during preimplantation development. Proper development of the TE and ICM lineages is dependent upon establishment of specific transcriptional programs. However, the epigenetic mechanisms that functionally contribute to establish TE- and ICM-specific transcriptional programs are poorly understood. Here, we show that proper development of the TE and ICM lineages is coordinated via combinatorial regulation of embryonic ectoderm development (EED) and lysine-specific demethylase 6B (KDM6B). During blastocyst formation, the relative levels of EED and KDM6B expression determine altered polycomb repressor 2 (PRC2) complex recruitment and incorporation of the repressive histone H3 lysine 27 trimethylation (H3K27Me3) mark at the chromatin domains of TE-specific master regulators CDX2 and GATA3, leading to their activation in the TE lineage and repression in the ICM lineage. Furthermore, ectopic gain of EED along with depletion of KDM6B in preimplantation mouse embryos abrogates CDX2 and GATA3 expression in the nascent TE lineage. The loss of CDX2 and GATA3 in the nascent TE lineage results in improper TE development, leading to failure in embryo implantation to the uterus. Our study delineates a novel epigenetic mechanism that orchestrates proper development of the first mammalian cell lineages.

    View details for DOI 10.1128/MCB.00069-13

    View details for Web of Science ID 000320714400005

    View details for PubMedID 23671187

  • Time-lapse microscopy and image analysis in basic and clinical embryo development research REPRODUCTIVE BIOMEDICINE ONLINE Wong, C., Chen, A. A., Behr, B., Shen, S. 2013; 26 (2): 120-129


    Mammalian preimplantation embryo development is a complex process in which the exact timing and sequence of events are as essential as the accurate execution of the events themselves. Time-lapse microscopy (TLM) is an ideal tool to study this process since the ability to capture images over time provides a combination of morphological, dynamic and quantitative information about developmental events. Here, we systematically review the application of TLM in basic and clinical embryo research. We identified all relevant preimplantation embryo TLM studies published in English up to May 2012 using PubMed and Google Scholar. We then analysed the technical challenges involved in embryo TLM studies and how these challenges may be overcome with technological innovations. Finally, we reviewed the different types of TLM embryo studies, with a special focus on how TLM can benefit clinical assisted reproduction. Although new parameters predictive of embryo development potential may be discovered and used clinically to potentially increase the success rate of IVF, adopting TLM to routine clinical practice will require innovations in both optics and image analysis. Combined with such innovations, TLM may provide embryologists and clinicians with an important tool for making critical decisions in assisted reproduction. In this review, we perform a literature search of all published early embryo development studies that used time-lapse microscopy (TLM). From the literature, we discuss the benefits of TLM over traditional time-point analysis, as well as the technical difficulties and solutions involved in implementing TLM for embryo studies. We further discuss research that has successfully derived non-invasive markers that may increase the success rate of assisted reproductive technologies, primarily IVF. Most notably, we extend our discussion to highlight important considerations for the practical use of TLM in research and clinical settings.

    View details for DOI 10.1016/j.rbmo.2012.11.003

    View details for Web of Science ID 000314664400003

    View details for PubMedID 23273754

  • Dynamic blastomere behaviour reflects human embryo ploidy by the four-cell stage NATURE COMMUNICATIONS Chavez, S. L., Loewke, K. E., Han, J., Moussavi, F., Colls, P., Munne, S., Behr, B., Pera, R. A. 2012; 3


    Previous studies have demonstrated that aneuploidy in human embryos is surprisingly frequent with 50-80% of cleavage-stage human embryos carrying an abnormal chromosome number. Here we combine non-invasive time-lapse imaging with karyotypic reconstruction of all blastomeres in four-cell human embryos to address the hypothesis that blastomere behaviour may reflect ploidy during the first two cleavage divisions. We demonstrate that precise cell cycle parameter timing is observed in all euploid embryos to the four-cell stage, whereas only 30% of aneuploid embryos exhibit parameter values within normal timing windows. Further, we observe that the generation of human embryonic aneuploidy is complex with contribution from chromosome-containing fragments/micronuclei that frequently emerge and may persist or become reabsorbed during interphase. These findings suggest that cell cycle and fragmentation parameters of individual blastomeres are diagnostic of ploidy, amenable to automated tracking algorithms, and likely of clinical relevance in reducing transfer of embryos prone to miscarriage.

    View details for DOI 10.1038/ncomms2249

    View details for Web of Science ID 000316356700019

    View details for PubMedID 23212380

  • Promotion of Human Early Embryonic Development and Blastocyst Outgrowth In Vitro Using Autocrine/Paracrine Growth Factors PLOS ONE Kawamura, K., Chen, Y., Shu, Y., Cheng, Y., Qiao, J., Behr, B., Pera, R. A., Hsueh, A. J. 2012; 7 (11)


    Studies using animal models demonstrated the importance of autocrine/paracrine factors secreted by preimplantation embryos and reproductive tracts for embryonic development and implantation. Although in vitro fertilization-embryo transfer (IVF-ET) is an established procedure, there is no evidence that present culture conditions are optimal for human early embryonic development. In this study, key polypeptide ligands known to be important for early embryonic development in animal models were tested for their ability to improve human early embryo development and blastocyst outgrowth in vitro. We confirmed the expression of key ligand/receptor pairs in cleavage embryos derived from discarded human tri-pronuclear zygotes and in human endometrium. Combined treatment with key embryonic growth factors (brain-derived neurotrophic factor, colony-stimulating factor, epidermal growth factor, granulocyte macrophage colony-stimulating factor, insulin-like growth factor-1, glial cell-line derived neurotrophic factor, and artemin) in serum-free media promoted >2.5-fold the development of tri-pronuclear zygotes to blastocysts. For normally fertilized embryos, day 3 surplus embryos cultured individually with the key growth factors showed >3-fold increases in the development of 6-8 cell stage embryos to blastocysts and >7-fold increase in the proportion of high quality blastocysts based on Gardner's criteria. Growth factor treatment also led to a 2-fold promotion of blastocyst outgrowth in vitro when day 7 surplus hatching blastocysts were used. When failed-to-be-fertilized oocytes were used to perform somatic cell nuclear transfer (SCNT) using fibroblasts as donor karyoplasts, inclusion of growth factors increased the progression of reconstructed SCNT embryos to >4-cell stage embryos. Growth factor supplementation of serum-free cultures could promote optimal early embryonic development and implantation in IVF-ET and SCNT procedures. This approach is valuable for infertility treatment and future derivation of patient-specific embryonic stem cells.

    View details for DOI 10.1371/journal.pone.0049328

    View details for Web of Science ID 000311234000064

    View details for PubMedID 23152897

  • Outcomes of trophectoderm biopsy on cryopreserved blastocysts: a case series REPRODUCTIVE BIOMEDICINE ONLINE Lathi, R. B., Massie, J. A., Gilani, M., Milki, A. A., Westphal, L. M., Baker, V. L., Behr, B. 2012; 25 (5): 504-507


    Preimplantation genetic diagnosis (PGD) is an increasingly common adjunct to IVF. The information gained from PGD may be used to reduce the incidence of chromosomally abnormal pregnancies and augment the current selection process of embryos. As such, patients may choose to utilize PGD in either fresh or cryopreserved IVF cycles. It is a common practice to cryopreserve excess embryos at the blastocyst stage. In these cases, trophectoderm biopsy is the only technique available for PGD. This articles reports this study centre's experience with trophectoderm biopsies of cryopreserved blastocysts in 12 patients who underwent 13 cycles of PGD. The implantation rate per embryo transferred was 46% and the ongoing pregnancy rate per embryo transfer was 63%. The results from this case series demonstrate that trophectoderm biopsy on cryopreserved blastocysts to perform PGD is logistically feasible. In addition, the rate of implantation and ongoing pregnancy were maintained within a reasonable range to justify the procedure. Preimplantation genetic diagnosis (PGD) is an increasingly common adjunct to IVF and is used to evaluate the genetic makeup of the embryo prior to transfer of the embryo into the uterus. The information gained from PGD may be used to identify single-gene disorders that result in genetic disease, reduce the incidence of chromosomally abnormal pregnancies and/or augment the selection process of embryos to be transferred. In order to perform PGD, a biopsy of the embryo is the performed and cells are removed for testing. PGD may be performed in either fresh or frozen (cryopreserved) IVF cycles. Patients who have cryopreserved embryos remaining in storage from a previous fresh cycle may wish to have these embryos tested with PGD. Many embryos are frozen on day 5 of development, referred to as the blastocyst stage. At this stage of development, embryo biopsy is performed via a technique known as 'trophectoderm biopsy', in which 1-3 of the cells destined to become the placenta are removed from the embryo for chromosomal testing. We report our experience with trophectoderm biopsy of frozen blastocysts in 12 patients who underwent 13 cycles of PGD. The implantation rate per embryo transferred was 46% and the ongoing pregnancy rate per embryo transfer was 63%. The results from this case series demonstrate that trophectoderm biopsy on cryopreserved blastocysts to perform PGD is logistically feasible. In addition, the rate of implantation and ongoing pregnancy were maintained within a reasonable range to justify the procedure.

    View details for DOI 10.1016/j.rbmo.2012.06.021

    View details for Web of Science ID 000310639600010

    View details for PubMedID 22985500

  • Genome-wide Single-Cell Analysis of Recombination Activity and De Novo Mutation Rates in Human Sperm CELL Wang, J., Fan, H. C., Behr, B., Quake, S. R. 2012; 150 (2): 402-412


    Meiotic recombination and de novo mutation are the two main contributions toward gamete genome diversity, and many questions remain about how an individual human's genome is edited by these two processes. Here, we describe a high-throughput method for single-cell whole-genome analysis that was used to measure the genomic diversity in one individual's gamete genomes. A microfluidic system was used for highly parallel sample processing and to minimize nonspecific amplification. High-density genotyping results from 91 single cells were used to create a personal recombination map, which was consistent with population-wide data at low resolution but revealed significant differences from pedigree data at higher resolution. We used the data to test for meiotic drive and found evidence for gene conversion. High-throughput sequencing on 31 single cells was used to measure the frequency of large-scale genome instability, and deeper sequencing of eight single cells revealed de novo mutation rates with distinct characteristics.

    View details for DOI 10.1016/j.cell.2012.06.030

    View details for Web of Science ID 000306595700018

    View details for PubMedID 22817899

  • Altered subcellular localization of transcription factor TEAD4 regulates first mammalian cell lineage commitment PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Home, P., Saha, B., Ray, S., Dutta, D., Gunewardena, S., Yoo, B., Pal, A., Vivian, J. L., Larson, M., Petroff, M., Gallagher, P. G., Schulz, V. P., White, K. L., Golos, T. G., Behr, B., Paul, S. 2012; 109 (19): 7362-7367


    In the preimplantation mouse embryo, TEAD4 is critical to establishing the trophectoderm (TE)-specific transcriptional program and segregating TE from the inner cell mass (ICM). However, TEAD4 is expressed in the TE and the ICM. Thus, differential function of TEAD4 rather than expression itself regulates specification of the first two cell lineages. We used ChIP sequencing to define genomewide TEAD4 target genes and asked how transcription of TEAD4 target genes is specifically maintained in the TE. Our analyses revealed an evolutionarily conserved mechanism, in which lack of nuclear localization of TEAD4 impairs the TE-specific transcriptional program in inner blastomeres, thereby allowing their maturation toward the ICM lineage. Restoration of TEAD4 nuclear localization maintains the TE-specific transcriptional program in the inner blastomeres and prevents segregation of the TE and ICM lineages and blastocyst formation. We propose that altered subcellular localization of TEAD4 in blastomeres dictates first mammalian cell fate specification.

    View details for DOI 10.1073/pnas.1201595109

    View details for Web of Science ID 000304090600052

    View details for PubMedID 22529382

  • Day 2 Transfer in Clinical ART SEMINARS IN REPRODUCTIVE MEDICINE Behr, B., McElroy, S. 2012; 30 (3): 222-229


    Over the past 20 years, numerous techniques have enhanced assisted reproductive technology outcomes to help couples have >60,000 infants in the United States in 2008. Several different days for embryo transfers have been studied, but debate for the best timing of embryo transfer is still ongoing. With growing concern about multiple gestations and neonatal outcomes, early cleavage stage embryo transfer with novel embryo selection tools may be attractive to some patients and in vitro fertilization programs. In this review, we summarize clinical and basic studies relating to the timing of embryo transfer and highlight the possibilities of safe embryo transfer by combining advanced embryo screening tools with potentially high efficiency and low adverse effects on clinical outcome.

    View details for DOI 10.1055/s-0032-1311524

    View details for Web of Science ID 000303703400009

    View details for PubMedID 22585633

  • Testosterone concentrations in early pregnancy: relation to method of conception in an infertile population REPRODUCTIVE BIOMEDICINE ONLINE Lathi, R. B., Moayeri, S. E., Reddy, C. D., Gebhardt, J., Behr, B., Westphal, L. M. 2012; 24 (3): 360-363


    This prospective cohort study of infertility patients compared testosterone concentrations in early pregnancy in infertility patients who conceived naturally or after treatment. Although all groups demonstrated some increase in pregnancy testosterone from baseline concentrations, subjects who conceived following ovulation induction showed a significantly increased rise in testosterone as compared with controls (P<0.01).

    View details for DOI 10.1016/j.rbmo.2011.11.018

    View details for Web of Science ID 000303046700015

    View details for PubMedID 22285241

  • Skeletogenic phenotype of human Marfan embryonic stem cells faithfully phenocopied by patient-specific induced-pluripotent stem cells PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Quarto, N., Leonard, B., Li, S., Marchand, M., Anderson, E., Behr, B., Francke, U., Reijo-Pera, R., Chiao, E., Longaker, M. T. 2012; 109 (1): 215-220


    Marfan syndrome (MFS) is a heritable connective tissue disorder caused by mutations in the gene coding for FIBRILLIN-1 (FBN1), an extracellular matrix protein. MFS is inherited as an autosomal dominant trait and displays major manifestations in the ocular, skeletal, and cardiovascular systems. Here we report molecular and phenotypic profiles of skeletogenesis in tissues differentiated from human embryonic stem cells and induced pluripotent stem cells that carry a heritable mutation in FBN1. We demonstrate that, as a biological consequence of the activation of TGF-β signaling, osteogenic differentiation of embryonic stem cells with a FBN1 mutation is inhibited; osteogenesis is rescued by inhibition of TGF-β signaling. In contrast, chondrogenesis is not perturbated and occurs in a TGF-β cell-autonomous fashion. Importantly, skeletal phenotypes observed in human embryonic stem cells carrying the monogenic FBN1 mutation (MFS cells) are faithfully phenocopied by cells differentiated from induced pluripotent-stem cells derived independently from MFS patient fibroblasts. Results indicate a unique phenotype uncovered by examination of mutant pluripotent stem cells and further demonstrate the faithful alignment of phenotypes in differentiated cells obtained from both human embryonic stem cells and induced pluripotent-stem cells, providing complementary and powerful tools to gain further insights into human molecular pathogenesis, especially of MFS.

    View details for DOI 10.1073/pnas.1113442109

    View details for Web of Science ID 000298876500045

    View details for PubMedID 22178754

  • Media composition: growth factors. Methods in molecular biology (Clifton, N.J.) Hegde, A., Behr, B. 2012; 912: 177-198


    Despite the fact that the fundamental principle underlying the most common method of culture media constitution is that of mimicking the natural environment of the preimplantation embryo, one major difference that remains between current embryo culture media and in vivo conditions is the absence of growth factors in vitro. Numerous growth factors are known to be present in the in vivo environment of human and nonhuman preimplantation embryos, often with peak concentrations corresponding to when fertilization and preimplantation embryo growth would occur. Although these growth factors are found in very small concentrations, they have a profound effect on tissue growth and differentiation through attachment to factor-specific receptors on cell surfaces. Receptors for many different growth factors have also been detected in human preimplantation embryos. Preimplantation embryos themselves express many growth factors. The growth factors and receptors are metabolically costly to produce, and thus their presence in the environment of the preimplantation embryo and in the embryo respectively strongly implies that embryos are designed to encounter and respond to the corresponding factors. Studies of embryo coculture also indirectly suggest that growth factors can improve in vitro development. Several animal and human studies attest to a probable beneficial effect of addition of growth factors to culture media. However, there is still ambiguity regarding the exact role of growth factors in embryonic development, the optimal dose of growth factors to be added to culture media, the combinatorial effect and endocrine of growth factors in embryonic development.

    View details for DOI 10.1007/978-1-61779-971-6_11

    View details for PubMedID 22829375

  • National collection of embryo morphology data into Society for Assisted Reproductive Technology Clinic Outcomes Reporting System: associations among day 3 cell number, fragmentation and blastomere asymmetry, and live birth rate FERTILITY AND STERILITY Racowsky, C., Stern, J. E., Gibbons, W. E., Behr, B., Pomeroy, K. O., Biggers, J. D. 2011; 95 (6): 1985-1989


    To evaluate the validity of collecting day 3 embryo morphology variables into the Society for Assisted Reproductive Technology Clinic Outcomes Reporting System (SART CORS).Retrospective.National database-SART CORS.Fresh autologous assisted reproductive technology (ART) cycles from 2006-2007 in which embryos were transferred singly (n=1,020) or in pairs (n=6,508) and embryo morphology was collected.None.Relationship between live birth, maternal age, and morphology of transferred day 3 embryos as defined by cell number, fragmentation, and blastomere symmetry. Logistic multiple regressions and receiver operating characteristic curve analyses were applied to determine specificity and sensitivity for correctly classifying embryos as either failures or successes.Live birth rate was positively associated with increasing cell number up to eight cells (<6 cells: 2.9%; 6 cells: 9.6%; 7 cells: 15.5%; 8 cells: 24.3%; and >8 cells: 16.2%), but was negatively associated with maternal age, increasing fragmentation, and asymmetry scores. An area under the receiver operating curve of 0.753 (95% confidence interval 0.740-0.766) was derived, with a sensitivity of 45.0%, a specificity of 83.2%, and 76.4% of embryos being correctly classified with a cutoff probability of 0.3.This analysis provides support for the validity of collecting morphology fields for day 3 embryos into SART CORS. Standardization of morphology collections will assist in controlling for embryo quality in future database analyses.

    View details for DOI 10.1016/j.fertnstert.2011.02.009

    View details for Web of Science ID 000289620900028

    View details for PubMedID 21411078

  • Donation of Embryos for Human Development and Stem Cell Research CELL STEM CELL Kalista, T., Freeman, H. A., Behr, B., Pera, R. R., Scott, C. T. 2011; 8 (4): 360-362


    Using donated human embryos for scientific research raises ethical questions about the donation process. We describe a two-stage consent process designed to help couples make informed decisions about embryo disposition. This consent methodology minimizes conflict of interest, respects patient choice, and provides a much-needed resource to patients and the research community.

    View details for DOI 10.1016/j.stem.2011.02.018

    View details for Web of Science ID 000289707100007

    View details for PubMedID 21474099

  • IVF Outcomes: Effects of Blood or Mucus on the Tip of a Soft Embryo Transfer Catheter After Embryo Transfer 59th Annual Meeting of the Pacific-Coast-Reproductive-Society Dasig, J., Zhao, Q., Shu, Y., Reddy, V., Gebhardt, J., Behr, B. ELSEVIER SCIENCE INC. 2011: S26–S26
  • Day 2 versus day 3 embryo transfer in poor responders: a prospective randomized trial FERTILITY AND STERILITY Shahine, L. K., Milki, A. A., Westphal, L. M., Baker, V. L., Behr, B., Lathi, R. B. 2011; 95 (1): 330-332


    Day 2 embryo transfer has been suggested as a method to improve pregnancy rates in poor responders compared with day 3 transfer. Our prospective randomized controlled trial does not show a difference in outcomes based on day of embryo transfer.

    View details for DOI 10.1016/j.fertnstert.2010.06.093

    View details for Web of Science ID 000285411600086

    View details for PubMedID 20813357

  • Inter-laboratory validation of the measurement of follicle stimulating hormone (FSH) after various lengths of frozen storage REPRODUCTIVE BIOLOGY AND ENDOCRINOLOGY Scriver, J., Baker, V. L., Young, S. L., Behr, B., Pastore, L. M. 2010; 8


    Serum follicle stimulating hormone (FSH) levels are used clinically to evaluate infertility, pituitary and gonadal disorders. With increased frequency of research collaborations across institutions, it is essential that inter-laboratory validation is addressed.An inter-laboratory validation of three commercial FSH immunoassays was performed with human serum samples of varying frozen storage length (2 batches of 15 samples each) at -25 degree C. Percentage differences and Bland-Altman limits of agreement were calculated.The inter- and intra-laboratory consistency of FSH values with the same assay manufacturer was much higher after shorter-term storage (frozen for less than 11 months, mean percentage degradation less than 4%) than after long-term storage (2-3 years, mean percentage degradation = 23%). Comparing assay results from different manufacturers, there was similar overall long term degradation as seen with the same manufacturer (-25%), however the degradation was greater when the original FSH was greater than 20 mIU/mL relative to less than 10 mIU/mL (p < 0.001 trend test).The findings suggest that degradation of serum samples stored between 11 months and 2-3 years at -25 degrees C can lead to unstable FSH measurements. Inter-laboratory variability due to frozen storage time and manufacturer differences in assay results should be accounted for when designing and implementing research or clinical quality control activities involving serum FSH at multiple study sites.

    View details for DOI 10.1186/1477-7827-8-145

    View details for Web of Science ID 000285635900001

    View details for PubMedID 21114859

  • Non-invasive imaging of human embryos before embryonic genome activation predicts development to the blastocyst stage NATURE BIOTECHNOLOGY Wong, C. C., Loewke, K. E., Bossert, N. L., Behr, B., De Jonge, C. J., Baer, T. M., Pera, R. A. 2010; 28 (10): 1115-U199


    We report studies of preimplantation human embryo development that correlate time-lapse image analysis and gene expression profiling. By examining a large set of zygotes from in vitro fertilization (IVF), we find that success in progression to the blastocyst stage can be predicted with >93% sensitivity and specificity by measuring three dynamic, noninvasive imaging parameters by day 2 after fertilization, before embryonic genome activation (EGA). These parameters can be reliably monitored by automated image analysis, confirming that successful development follows a set of carefully orchestrated and predictable events. Moreover, we show that imaging phenotypes reflect molecular programs of the embryo and of individual blastomeres. Single-cell gene expression analysis reveals that blastomeres develop cell autonomously, with some cells advancing to EGA and others arresting. These studies indicate that success and failure in human embryo development is largely determined before EGA. Our methods and algorithms may provide an approach for early diagnosis of embryo potential in assisted reproduction.

    View details for DOI 10.1038/nbt.1686

    View details for Web of Science ID 000282870500034

    View details for PubMedID 20890283

  • Parthenogenic Blastocysts Derived from Cumulus-Free In Vitro Matured Human Oocytes PLOS ONE McElroy, S. L., Byrne, J. A., Chavez, S. L., Behr, B., Hsueh, A. J., Westphal, L. M., Pera, R. A. 2010; 5 (6)


    Approximately 20% of oocytes are classified as immature and discarded following intracytoplasmic sperm injection (ICSI) procedures. These oocytes are obtained from gonadotropin-stimulated patients, and are routinely removed from the cumulus cells which normally would mature the oocytes. Given the ready access to these human oocytes, they represent a potential resource for both clinical and basic science application. However culture conditions for the maturation of cumulus-free oocytes have not been optimized. We aimed to improve maturation conditions for cumulus-free oocytes via culture with ovarian paracrine/autocrine factors identified by single cell analysis.Immature human oocytes were matured in vitro via supplementation with ovarian paracrine/autocrine factors that were selected based on expression of ligands in the cumulus cells and their corresponding receptors in oocytes. Matured oocytes were artificially activated to assess developmental competence. Gene expression profiles of parthenotes were compared to IVF/ICSI embryos at morula and blastocyst stages. Following incubation in medium supplemented with ovarian factors (BDNF, IGF-I, estradiol, GDNF, FGF2 and leptin), a greater percentage of oocytes demonstrated nuclear maturation and subsequently, underwent parthenogenesis relative to control. Similarly, cytoplasmic maturation was also improved as indicated by development to blastocyst stage. Parthenogenic blastocysts exhibited mRNA expression profiles similar to those of blastocysts obtained after IVF/ICSI with the exception for MKLP2 and PEG1.Human cumulus-free oocytes from hormone-stimulated cycles are capable of developing to blastocysts when cultured with ovarian factor supplementation. Our improved IVM culture conditions may be used for obtaining mature oocytes for clinical purposes and/or for derivation of embryonic stem cells following parthenogenesis or nuclear transfer.

    View details for DOI 10.1371/journal.pone.0010979

    View details for Web of Science ID 000278465900004

    View details for PubMedID 20539753

  • Normal pregnancy after tetraploid karyotype on trophectoderm biopsy FERTILITY AND STERILITY Krieg, S. A., Lathi, R. B., Behr, B., Westphal, L. M. 2009; 92 (3)


    To report a case of successful pregnancy after trophectoderm biopsy and fluorescence in situ hybridization (FISH) revealed a tetraploid karyotype.Case report.A university medical center.An infertility patient desiring trophectoderm biopsy on frozen blastocysts to facilitate preimplantation genetic screening.Frozen blastocysts were thawed on the evening before transfer. Trophectoderm biopsy was performed the following morning. FISH results were available the same day, and two embryos with tetraploid results were transferred.Chorionic villus sample (CVS) and newborn exam.Normal diploid CVS result and a healthy male infant.Although multiple cells can be analyzed using trophectoderm biopsy, abnormalities in the trophectoderm may not be present in the inner cell mass.

    View details for DOI 10.1016/j.fertnstert.2009.06.007

    View details for Web of Science ID 000283282700007

    View details for PubMedID 19608167

  • The Role of Prolactin- and Endometriosis-Associated Infertility OBSTETRICAL & GYNECOLOGICAL SURVEY Wang, H., Gorpludolo, N., Behr, B. 2009; 64 (8): 542-547


    This review will address the current understanding of the relationship between prolactin (PRL) and endometriosis-associated infertility. Although the exact mechanisms of action of hyperprolactinemia in patients with endometriosis-associated infertility have not been clearly established, this report reviews results from relevant studies in the literature. These include serum PRL levels in endometriosis-associated infertility, PRL receptors in ectopic endometriotic tissues, basal PRL levels after TSH and Danazol (isoxazolic derivative of the synthetic steroid 5alpha-ethinyl-testosterone) therapy, peritoneal fluid and nocturnal serum PRL levels in endometriosis, infertility, and luteal phase PRL concentrations in patients with endometriosis.Obstetricians & Gynecologists, Family Physicians.After completion of this article, the reader should be able to explain the relationship between prolactin- and endometriosis-associated infertility, relate endometriosis with infertility, and summarize two ways in which prolactin and endometriosis may be linked in the pathophysiology of infertility.

    View details for Web of Science ID 000268610800017

    View details for PubMedID 19624865

  • Pregnancy after trophectoderm biopsy of frozen-thawed blastocyst FERTILITY AND STERILITY Lathi, R. B., Behr, B. 2009; 91 (5): 1938-1940


    To report a case of a successful pregnancy after trophectoderm biopsy and three-probe fluorescent in situ hybridization of a frozen blastocyst.Techniques and instrumentation.A University Medical Center.Infertility patient desiring trophectoderm biopsy on frozen blastocyst for preimplantation testing, from an IVF cycle at a referring IVF program.Frozen blastocysts were thawed the evening before the planned transfer. Trophectoderm biopsy was performed in the morning. The fluorescent in situ hybridization results were obtained the same day; embryo transfer was performed under ultrasound guidance.Serum betahCG and transvaginal ultrasound.Positive betahCG and ongoing pregnancy.Trophectoderm biopsy can be used as a means for testing frozen blastocysts in patients with excess embryos cryopreserved on day 5 or 6 from previously preformed IVF cycles.

    View details for DOI 10.1016/j.fertnstert.2008.02.132

    View details for Web of Science ID 000265969200055

    View details for PubMedID 18371958

  • Embryo quality before and after surgical treatment of endometriosis in infertile patients JOURNAL OF ASSISTED REPRODUCTION AND GENETICS Shahine, L. K., Burney, R. O., Behr, B., Milki, A. A., Westphal, L. M., Lathi, R. B. 2009; 26 (2-3): 69-73


    To investigate the hypothesis that surgical treatment of endometriosis in infertile patients may improve pregnancy rates by improving embryo quality.We conducted a retrospective evaluation of 30 infertile patients treated with in vitro fertilization (IVF) before and after surgery for endometriosis. Patients served as their own controls and only cycles with similar stimulation protocols were compared.Using standard visual evaluation, embryo quality on day 3 was similar before and after surgical treatment of endometriosis. Fifty seven percent of patients had stage I-II endometriosis and 43% had stage III-IV disease. No patients had a live birth after the first IVF cycle and 43% of patients had a live birth with the IVF cycle after surgery.Surgical treatment of endometriosis does not alter embryo quality in patients with infertility treated with IVF.

    View details for DOI 10.1007/s10815-008-9287-1

    View details for Web of Science ID 000264178200001

    View details for PubMedID 19214735

  • Non-Redundant Prognostic Factors in First-Cycles in In Vitro Fertilization 56th Annual Meeting of the Society-for-Gynecological-Investigation Shahine, L. K., Choi, B., O'Leary, K., Jun, S. H., Westphal, L. M., Behr, B., Wong, W. H., Yao, M. W. SAGE PUBLICATIONS INC. 2009: 279A–279A
  • The value of fast blastocoele re-expansion in the selection of a viable thawed blastocyst for transfer 61st Annual Meeting of the American-Society-for-Reproductive-Medicine/51st Annual Meeting of the Canadian-Fertility-and-Andrology-Society Shu, Y., Watt, J., Gebhardt, J., Dasig, J., Appling, J., Behr, B. ELSEVIER SCIENCE INC. 2009: 401–6


    To investigate the role of fast blastocoele re-expansion in the selection of viable thawed blastocysts for transfer.Retrospective study.Academic assisted reproductive program.Transfer cycles were divided into two groups according to the presence or absence of fast re-expanded blastocysts. In group I (124 cycles), all transferred blastocysts had fast re-expanding blastocoele. In group II (113 cycles), no fast re-expanded blastocysts were included in the transfer.Blastocyst survival was defined as >50% of cells remaining intact after thaw and re-expansion after culture in vitro for 2-4 hours before transfer. Blastocysts with >or=50% re-expansion were designated as fast re-expanded blastocysts.Percentage of blastomere loss immediately after thaw, degree of blastocoele re-expansion, and clinical outcomes (pregnancy and implantation rates).The rates of survival and fast blastocoele re-expansion of partially intact blastocysts were significantly reduced as compared with fully intact blastocysts. Significantly higher rates of clinical pregnancy (37.1% vs. 16.8%) and implantation (26.7% vs. 11.3%) were obtained when all transferred blastocysts had fast re-expanding blastocoele as compared with those transfers without fast re-expanded blastocysts included.Our results showed that blastomere loss of thawed blastocyst was associated with a reduced ability to re-expand. As a discriminative morphologic marker of superior embryo viability, a fast re-expanded blastocyst would be given priority for transfer to better utilize the cryopreserved blastocysts.

    View details for DOI 10.1016/j.fertnstert.2007.11.083

    View details for Web of Science ID 000263445300016

    View details for PubMedID 18304536

  • Metabolomic assessment of oocyte viability REPRODUCTIVE BIOMEDICINE ONLINE Nagy, Z. P., Jones-Colon, S., Roos, P., Botros, L., Greco, E., Dasig, J., Behr, B. 2009; 18 (2): 219-225


    The aim of the current study was to evaluate whether near-infrared (NIR) spectroscopy-generated metabolomic data obtained from oocyte culture samples would correlate with nuclear maturity status and derived embryo development. A total of 412 oocyte culture samples were collected from 43 patient cycles. Metabolomic profiles of metaphase I and II oocytes were obtained by NIR spectroscopy and were significantly different from each other and from profiles of prophase I (germinal vesicle) oocytes (P +/- 0.001 at the 95% confidence interval). Additionally, NIR spectroscopic analysis of culture medium of oocytes that developed to grade A embryos on day 3 demonstrated significantly higher viability indices (0.62 +/- 0.23) than those that developed to grades C/D (0.42 +/- 0.26; P < 0.006); and on day 5 grade A (0.37 +/- 0.20) was also higher than grades C/D (0.14 +/- 0.21; P < 0.02). Metabolomic profiles of oocytes that resulted in pregnancy had higher viability indices (0.87 +/- 0.27) than those that did not (0.44 +/- 0.17; P < 0.0001). The results of the current study demonstrate that metabolomic profiling from spent culture medium of the oocyte is related to nuclear maturity, is able to predict embryo development at day 3 and day 5 stages, and relates to embryo viability.

    View details for Web of Science ID 000263251700010

    View details for PubMedID 19192342

  • Non-invasive assessment of embryo viability by metabolomic profiling of culture media ('metabolomics') 22nd Annual Meeting of the European-Society-of-Human-Reproduction-and-Embryology Nagy, Z. P., Sakkas, D., Behr, B. ELSEVIER SCI LTD. 2008: 502–7
  • Defining Human Embryo Phenotypes by Cohort-Specific Prognostic Factors PLOS ONE Jun, S. H., Choi, B., Shahine, L., Westphal, L. M., Behr, B., Pera, R. A., Wong, W. H., Yao, M. W. 2008; 3 (7)


    Hundreds of thousands of human embryos are cultured yearly at in vitro fertilization (IVF) centers worldwide, yet the vast majority fail to develop in culture or following transfer to the uterus. However, human embryo phenotypes have not been formally defined, and current criteria for embryo transfer largely focus on characteristics of individual embryos. We hypothesized that embryo cohort-specific variables describing sibling embryos as a group may predict developmental competence as measured by IVF cycle outcomes and serve to define human embryo phenotypes.We retrieved data for all 1117 IVF cycles performed in 2005 at Stanford University Medical Center, and further analyzed clinical data from the 665 fresh IVF, non-donor cycles and their associated 4144 embryos. Thirty variables representing patient characteristics, clinical diagnoses, treatment protocol, and embryo parameters were analyzed in an unbiased manner by regression tree models, based on dichotomous pregnancy outcomes defined by positive serum beta-human chorionic gonadotropin (beta-hCG). IVF cycle outcomes were most accurately predicted at approximately 70% by four non-redundant, embryo cohort-specific variables that, remarkably, were more informative than any measures of individual, transferred embryos: Total number of embryos, number of 8-cell embryos, rate (percentage) of cleavage arrest in the cohort and day 3 follicle stimulating hormone (FSH) level. While three of these variables captured the effects of other significant variables, only the rate of cleavage arrest was independent of any known variables.Our findings support defining human embryo phenotypes by non-redundant, prognostic variables that are specific to sibling embryos in a cohort.

    View details for DOI 10.1371/journal.pone.0002562

    View details for Web of Science ID 000263288200029

    View details for PubMedID 18596962

  • A comparison of blastocyst formation from one-cell mouse zygotes following an aseptic vitrification system 56th Annual Meeting of the Pacific-Coast-Reproductive-Society Dasig, J., Bertocci, E., Zhao, Q., Shu, Y., Behr, B. ELSEVIER SCIENCE INC. 2008: S7–S7
  • Application of custom-made electrofusion pipettes in mouse somatic cell nuclear transfer 56th Annual Meeting of the Pacific-Coast-Reproductive-Society Shu, Y., Rodriguez, R., Kim, S., Behr, B. ELSEVIER SCIENCE INC. 2008: S28–S28
  • Derivation of human embryonic stem cells in standard and chemically defined conditions STEM CELL CULTURE Chiao, E., Kmet, M., Behr, B., Baker, J. 2008; 86: 1-?
  • Normal pregnancy resulting from a non-pronuclear oocyte at the time of examination for fertilization CLINICAL AND EXPERIMENTAL OBSTETRICS & GYNECOLOGY Burney, R. O., Gebhardt, J., Shu, Y., Behr, B., Westphal, L. M. 2008; 35 (3): 170-171


    To report the case of a patient undergoing in vitro fertilization (IVF) in which a non-pronuclear (0PN) oocyte resulted in a normal pregnancy.A 36-year-old woman underwent an IVF-embryo transfer treatment cycle.Four oocytes were retrieved for insemination by IVF. Examination for fertilization revealed two polypronuclearpolygynic and two non-pronuclear oocytes. The non-pronuclear oocytes were observed further for development. One embryo developed from the non-pronuclear cohort and was transferred at the 8-cell stage on day 3. Subsequently, a pregnancy developed, and resulted in the delivery of a healthy term infant.Non-pronuclear oocytes may represent a source of developmentally competent embryos, and further observation of this cohort should be considered, particularly in situations involving a low yield of oocytes at retrieval.

    View details for Web of Science ID 000257942800003

    View details for PubMedID 18754284

  • Unique biomarkers of human oocyte maturation assessed by non-invasive metabolomic profiling. 63rd Annual Meeting of the American-Society-for-Reproductive-Medicine Nagy, Z. P., Behr, B., Roos, P., Dasig, J., Burns, D. ELSEVIER SCIENCE INC. 2007: S4–S4
  • Use and outcomes of intracytoplasmic sperm injection for non-male factor infertility 101st Annual Meeting of the American-Urological-Association Kim, H. H., Bundorf, M. K., Behr, B., McCallum, S. W. ELSEVIER SCIENCE INC. 2007: 622–28


    To determine whether intracytoplasmic sperm injection (ICSI) is associated with improved outcomes for non-male factor infertility.We examined the patient characteristics associated with treatment choice-ICSI and conventional in vitro fertilization (IVF)-among patients without a diagnosis of male factor infertility and compared outcomes between the two groups, adjusting for patient characteristics using multivariate regression models.Academic fertility center.We evaluated 696 consecutive assisted reproductive technology (ART) cycles performed for couples with normal semen analysis at the Stanford Reproductive Endocrinology and Infertility Center between 2002 and 2003. We compared patient characteristics, cycle details, and outcomes for ICSI and IVF.Fertilization, pregnancy, and live birth rates.Patient characteristics were similar between the two groups, except the proportion of patients with unexplained infertility (IVF 15.1% vs. ICSI 23.5%), previous fertility (IVF 62.6% vs. ICSI 45.5%), and previous ART cycle (IVF 41.2% vs. ICSI 67.7%). More oocytes were fertilized per cycle for the IVF group (6.6 oocytes versus 5.1 oocytes). Fertilization failure, pregnancy, and live birth rates did not differ between IVF and ICSI. Using logistic regressions, having had previous ART was found to be positively associated with ICSI. Treatment choice of ICSI was not associated with fertilization, pregnancy, or live birth rates.No clear evidence of improved outcomes with ICSI was demonstrated for non-male factor infertility.

    View details for DOI 10.1016/j.fertnstert.2006.12.013

    View details for Web of Science ID 000249751900014

    View details for PubMedID 17445809

  • Effects of blastomere loss on developmental competency of frozen-thawed human blastocysts. 63rd Annual Meeting of the American-Society-for-Reproductive-Medicine Shu, Y., Watt, J., Janice, G., Dasig, J., Jensen, J., Behr, B. ELSEVIER SCIENCE INC. 2007: S356–S356
  • Importance of integrity of blastomere in the re-expansion of cryopreserved blastocysts. 63rd Annual Meeting of the American-Society-for-Reproductive-Medicine Shu, Y., Watt, J., Gebhardt, J., Zhao, Q., Behr, B. ELSEVIER SCIENCE INC. 2007: S314–S314
  • Expression of CD9 in frozen-thawed mouse oocytes: preliminary experience FERTILITY AND STERILITY Wen, Y., Quintero, R., Chen, B., Shu, Y., Polan, M. L., Behr, B. 2007; 88 (2): 526-529


    CD9 mRNA and protein expression levels in mouse slow frozen-rapid thawed oocytes were compared with those in fresh oocytes by using comparative quantitative real time reverse transcription-PCR and semiquantitative Western blot, respectively. The expression levels of both CD9 mRNA and protein in the frozen oocytes were significantly lower than those found in the fresh oocytes.

    View details for DOI 10.1016/j.fertnstert.2006.11.130

    View details for Web of Science ID 000248716000044

    View details for PubMedID 17307168

  • Identification of unique biomarkers of human oocyte maturation by non-invasive metabolomic profiling 23rd Annual Meeting of the European-Society-of-Human-Reproduction-and-Embryology Behr, B., Nagy, Z. P., Roos, R. P., Dasig, J., Kort, H. I., Burns, D. H. OXFORD UNIV PRESS. 2007: I166–I167
  • Fertilization, embryo development, and clinical outcome of immature oocytes from stimulated intracytoplasmic sperm injection cycles 60th Annual Meeting of the American-Society-for-Reproductive-Medicine Shu, Y., Gebhardt, J., Watt, J., Lyon, J., Dasig, D., Behr, B. ELSEVIER SCIENCE INC. 2007: 1022–27


    To evaluate the fertilization and developmental potential of immature oocytes obtained from controlled ovarian hyperstimulated cycles of patients undergoing intracytoplasmic sperm injection (ICSI).Retrospective study.Academic assisted reproductive technology program.Two hundred patients with at least one mature oocyte and one immature oocyte (study 1), and 44 patients with no mature oocytes (study 2) at time of oocyte denudation.Oocyte denudation was performed immediately after retrieval. Oocytes were cultured in vitro for 4-6 hours before ICSI and then categorized into four groups: group I, metaphase II (MII) oocytes at denudation; group II, in vitro matured MII oocytes; group III, metaphase I (MI) oocytes that did not progress to MII; and group 4, germinal-vesicle (GV) oocytes that converted to MI.Fertilization and embryo development were compared among groups in study 1. Pregnancy and implantation rates were evaluated in study 2.Although the fertilization rate in group III was significantly lower than in groups I and II, no significant difference was found between groups I and II. Day 3 embryos in group I had the highest mean number of blastomeres, proportions of good embryos, and blastocyst formation rate when compared with groups II and III. Two clinical pregnancies were achieved from 26 transfer cycles in study 2, resulting in pregnancy and implantation rates of 7.7% and 4% per transfer cycle, respectively.Although our results show that immature oocytes from stimulated cycles can be normally fertilized and used to increase the number of embryos available for transfer, the increase in number of embryos derived from immature oocytes cannot be efficiently translated into pregnancies and live births. The clinical significance of using immature oocytes in stimulated cycles needs further investigation.

    View details for DOI 10.1016/j.fertnstert.2006.08.110

    View details for Web of Science ID 000246583600005

    View details for PubMedID 17261289

  • Risk of monozygotic twinning with blastocyst transfer decreases over time: an 8-year experience FERTILITY AND STERILITY Moayeri, S. E., Behr, B., Lathi, R. B., Westphal, L. M., Milki, A. A. 2007; 87 (5): 1028-1032


    The purpose of our study is to compare the occurrence of monozygotic twinning (MZT) from blastocyst transfer (BT) in our program between an earlier and more recent time period.Retrospective.Academic IVF practice.All pregnancies conceived between March 2002 and December 2005 (N = 932) in our program were compared to pregnancies conceived before March 2002 (N = 554), which were the subject of a previous study.None.The incidence of MZT with day 3 embryo transfer and BT were compared between the study and control groups.During the study period, the rate of MZT was not significantly different for BT at 2.3% (9/385) compared to day 3 embryo transfer at 1.8% (10/547). This rate of 2.3% for BT was significantly lower than the rate of 5.6% (11/197) reported at our institution for BT before March 2002.Our study suggests that the risk of MZT with BT is significantly lower in the more recent time period and is in the range of what is seen with cleavage stage transfer. It is likely that improvements in culture systems as experience is gained with BT played a role.

    View details for DOI 10.1016/j.fertnstert.2006.09.013

    View details for Web of Science ID 000246583600006

    View details for PubMedID 17343858

  • Effect of reduced oxygen concentrations on the outcome of in vitro fertilization FERTILITY AND STERILITY Kea, B., Gebhardt, J., Watt, J., Westphal, L. M., Lathi, R. B., Milki, A. A., Behr, B. 2007; 87 (1): 213-216


    We compared the effects of two standard oxygen concentrations, physiological (5% O(2), 5% CO(2), and 90% N(2)) and atmospheric (5% CO(2) with the balance as air), on fertilization, embryo development, and pregnancy rate in 106 patients undergoing IVF, excluding donor oocyte cycles and preimplantation genetic diagnosis cycles. The differences in oxygen concentration did not significantly affect fertilization rate, blastocyst formation, or pregnancy rate, but there was a significant difference in mean embryo score between physiological and atmospheric groups on day 3.

    View details for DOI 10.1016/j.fertnstert.2006.05.066

    View details for Web of Science ID 000243436600035

    View details for PubMedID 17081523

  • Successful pregnancies after transplantation of frozen-thawed mouse ovaries into chimeric mice that received lethal-dose radiation FERTILITY AND STERILITY Migishima, F., Suzuki-Migishima, R., Quintero, R. B., Yokoyama, M., Behr, B. R. 2006; 86: 1080-1087


    To study whether fecundity was recovered in mice into which umbilical cord blood cells (UCBCs) were transfused after lethal-dose radiation, followed by transplantation of frozen-thawed ovaries.Prospective basic research study.Academic research laboratory.Female C57BL/6 mice as recipients of UCBCs and ovaries, male B6C3F1 mice for mating, and green fluorescent protein (GFP)-transgenic mice: 18.5-day-old fetuses (-/+) for UCBCs and adult GFP mice (+/+) for ovarian tissues.The UCBCs were transfused into each irradiated mouse, with GFP+ ovaries transplanted 4 weeks later. The chimeric mice were mated 3 weeks after ovarian transplantation and were examined 14 to 16 weeks after the transfusion of UCBCs.Percentage of chimerism, number of GFP+ pups.The percentage of chimerism in these mice tends to increase with the radiation dose. The recovery of fecundity was observed in the chimeric mice that were transplanted with fresh and previously vitrified ovaries after exposure to radiation.Even when the exposure dose of radiation administered as pretreatment is lethal, the fecundity of recipients can be maintained if their ovaries are cryopreserved before they are exposed to radiation.

    View details for DOI 10.1016/j.fertnstert.2006.03.023

    View details for Web of Science ID 000241289300007

    View details for PubMedID 16978625

  • Preliminary experience on the CD9 expression on frozen-thawed mouse oocytes. 62nd Annual Meeting of the American-Society-for-Reproductive-Medicine (ASRM) Behr, B., Wen, Y., Quintero, R., Chen, B., Polan, M. L. ELSEVIER SCIENCE INC. 2006: S196–S196
  • Non-invasive metabolomic profiling of human embryo culture media correlates with pregnancy outcome. Initial results of the metabolomics study group for ART. 62nd Annual Meeting of the American-Society-for-Reproductive-Medicine (ASRM) Seli, E., Sakkas, D., Behr, B., Nagy, P., Kwok, J. S., Burns, D. H. ELSEVIER SCIENCE INC. 2006: S115–S115
  • Trophectoderm: A possible indicator of blastocyst survival and re-expansion after cryopreservation. 62nd Annual Meeting of the American-Society-for-Reproductive-Medicine (ASRM) Shu, Y., Gebhardt, J., Watt, J., Lyon, J., Jensen, J., Behr, B. ELSEVIER SCIENCE INC. 2006: S146–S147
  • A novel microfluidic device for male subfertility screening JOURNAL OF UROLOGY McCormack, M. C., McCallum, S., Behr, B. 2006; 175 (6): 2223-2227


    To our knowledge no simple, disposable and accurate test of semen quality currently exists. A novel technique to assess the motile sperm concentration of the human ejaculate has been designed and its results are presented.In a micromachined device fluorescently labeled motile sperm traverse a hydrostatic microfluid line to a target detection cuvette. A microfluorometer assesses the fluorescence signal generated by sperm accumulating there throughout a 50-minute study period. A total of 21 semen specimens from men presenting to our university based reproductive endocrinology and infertility center were tested a total of 67 times. Semen parameters determined by computer assisted semen analysis were compared to the signal reported by the microfluidic device.The fluorescence signal increased throughout the data collection period for all samples. Pearson r values relating the device signal to total and progressive motile concentration were 0.79 and 0.80, respectively (each p <0.001). A signal threshold based on the aggregate data were established, correlating with the WHO standard of the normal total motile sperm concentration. As a screening test, the device was 94% sensitive and 97% specific for identifying samples with less than the WHO standard for the total motile concentration, and 96% sensitive and 90% specific when considering the progressive motile concentration.A novel microfluidic device is presented that enables accurate assessment of the motile sperm concentration in human ejaculate compared to computer assisted semen analysis. Its size and design demonstrate the feasibility of applying laboratory on chip technology to male infertility screening.

    View details for DOI 10.1016/S0022-5347(06)00276-X

    View details for Web of Science ID 000237585100067

    View details for PubMedID 16697844

  • Regulation of cyclooxygenase activity in cultured endometrial stromal cells by granulocyte-macrophage colony-stimulating factor FERTILITY AND STERILITY Wang, H. B., Wen, Y., Polan, M. L., Boostanfar, R., Feinman, M., Behr, B. 2006; 85: 1118-1124


    To assess the ability of granulocyte-macrophage colony-stimulating factor (GM-CSF) to regulate cyclooxygenase (COX) enzyme activity and prostaglandins (PGs) synthesis, specifically PGE2 production in stromal cells, neither of which have been addressed in the literature.Prospective study.Department of obstetrics and gynecology at a university hospital.Human luteal phase endometrium was obtained from surgical specimens (n = 6) for clinical indications.Confluent stromal cells were stimulated with GM-CSF.Expression of COX mRNA, COX enzyme activity, and PGE2 level in cultured stromal cells.Confluent stromal cell cultures treated with P and E2 for 9 days were stimulated with GM-CSF. After treatment of 12 hours, low-dose GM-CSF (0.001-0.1 ng/mL) increased COX-2 mRNA levels in stromal cell, whereas high dose GM-CSF (1-100 ng/mL) decreased COX-1 and COX-2 mRNA levels. After treatment of 48 hours, low concentrations of GM-CSF (0.001-0.1 ng/mL) increased total COX and COX-2 enzyme activity, whereas high concentrations of GM-CSF (1-100 ng/mL) inhibited COX and COX-2 activity; The PGE2 levels decreased by 31% to 393.3 pg/mL (P < .05) with concentrations of GM-CSF increasing from 1 ng/mL to 100 ng/mL.There appeared to be a biphasic pattern of COX-2 enzyme response to GM-CSF with low concentrations increasing activity and high concentrations inhibiting activity. It is possible that GM-CSF may provide critical regulation of PG production in the preimplantation period.

    View details for DOI 10.1016/j.fertnstert.2005.09.040

    View details for Web of Science ID 000236902300007

    View details for PubMedID 16616083

  • Exogenous granulocyte-macrophage colony-stimulating factor promotes follicular development in the newborn rat in vivo HUMAN REPRODUCTION Wang, H. B., Wen, Y., Polan, M. L., Boostanfar, R., Feinman, M., Behr, B. 2005; 20 (10): 2749-2756


    Expression and selective cellular localization of granulocyte-macrophage colony-stimulating factor (GM-CSF) and its receptor in ovarian tissue imply an autocrine/paracrine role in ovarian function. Evidence indicating a functional role for GM-CSF in ovarian follicular cell function has been provided by studies with GM-CSF knockout (GM-/-) mice, which suggest that GM-CSF influences events associated with murine follicular maturation.Immature female rats were treated with GM-CSF, FSH or saline for 5 or 10 days. Ovaries were collected for histologic examination and immunostaining determination of CYP17, a theca cell marker. In addition, ovarian section slides were evaluated by immunofluorescence for CD45, an ovarian leukocyte marker. To investigate the possible mechanism of GM-CSF action on follicular development, theca-interstitial cells (T-I) were separated and cultured. Cells were treated with increasing concentrations of GM-CSF, then evaluated for CYP17 mRNA and protein expression assays.After 10 days of treatment with GM-CSF, the number of small preantral and large preantral follicles was significantly increased compared with the control group (P < 0.05). Similarly, treatment with FSH increased the number of small preantral and large preantral follicles (P < 0.05). CD45 expression measured by immunofluorescence was not different in the three groups, indicating that the distribution of leukocytes was unchanged. In addition, CYP17 was increased in the T-I cells both in vivo and in vitro after GM-CSF treatment.The present results suggest that GM-CSF may play a significant role in follicular development.

    View details for DOI 10.1093/humrep/dei123

    View details for Web of Science ID 000232427600013

    View details for PubMedID 15958400

  • Effects of repetitive vitrification on the survival of mouse oocytes. 61st Annual Meeting of the American-Society-for-Reproductive-Medicine/51st Annual Meeting of the Canadian-Fertility-and-Andrology-Society Migishima, F., Shu, Y., Chen, B., Zhao, Y., Polan, M. L., Behr, B. ELSEVIER SCIENCE INC. 2005: S186–S186
  • IVF results in de novo DNA methylation and histone methylation at an Igf2-H19 imprinting epigenetic switch MOLECULAR HUMAN REPRODUCTION Li, T., Vu, T. H., Ulaner, G. A., Littman, E., Ling, J. Q., Chen, H. L., Hu, J. F., Behr, B., Giudice, L., Hoffman, A. R. 2005; 11 (9): 631-640


    Recent studies suggest that IVF and assisted reproduction technologies (ART) may result in abnormal genomic imprinting, leading to an increased frequency of Angelman syndrome (AS) and Beckwith-Weidemann syndrome (BWS) in IVF children. To learn how ART might alter the epigenome, we examined morulas and blastocysts derived from C57BL/6J X M. spretus F1 mice conceived in vivo and in vitro and determined the allelic expression of four imprinted genes: Igf2, H19, Cdkn1c and Slc221L. IVF-derived mouse embryos that were cultured in human tubal fluid (HTF) (Quinn's advantage) media displayed a high frequency of aberrant H19 imprinting, whereas in vivo and IVF embryos showed normal maternal expression of Cdkn1c and normal biallelic expression of Igf2 and Slc221L. Embryonic stem (ES) cells derived from IVF blastocysts also showed abnormal Igf2/H19 imprinting. Allele-specific bisulphite PCR reveals abnormal DNA methylation at a CCCTC-binding factor (CTCF) site in the imprinting control region (ICR), as the normally unmethylated maternal allele acquired a paternal methylation pattern. Chromatin immunoprecipitation (ChIP) assays indicate an increase of lysine 4 methylation (dimethyl Lys4-H3) on the paternal chromatin and a gain in lysine 9 methylation (trimethyl Lys9-H3) on the maternal chromatin at the same CTCF-binding site. Our results indicate that de novo DNA methylation on the maternal allele and allele-specific acquisition of histone methylation lead to aberrant Igf2/H19 imprinting in IVF-derived ES cells. We suggest that ART, which includes IVF and various culture media, might cause imprinting errors that involve both aberrant DNA methylation and histone methylation at an epigenetic switch of the Igf2-H19 gene region.

    View details for DOI 10.1093/molehr/gah230

    View details for Web of Science ID 000233361600003

    View details for PubMedID 16219628

  • Osmotic behavior of human blastocysts as a potential predictor for survival. 61st Annual Meeting of the American-Society-for-Reproductive-Medicine/51st Annual Meeting of the Canadian-Fertility-and-Andrology-Society Shu, Y., Gebhardt, J., Watt, J., Lyon, J., Jensen, J., Behr, B. ELSEVIER SCIENCE INC. 2005: S186–S187
  • Efficacy of recombinant human serum albumin as a protein source for the development of in vitro cultured human embryos: a pilot study. 53rd Annual Meeting of the Pacific-Coast-Reproductive-Society Lyon, J., QUINN, P., Carter, D. C., Ye, P., Dasig, D., Behr, B. ELSEVIER SCIENCE INC. 2005: S25–S25
  • The use of recombinant human serum albumin (rHSA) for mouse IVF and embryo culture: Effect of diluent and protein stabilizers. 53rd Annual Meeting of the Pacific-Coast-Reproductive-Society QUINN, P., Carter, D. C., Ye, P., Behr, B. ELSEVIER SCIENCE INC. 2005: S24–S24
  • Effect of GMCSF on development and gene imprinting in preimplantation mouse embryos. 53rd Annual Meeting of the Pacific-Coast-Reproductive-Society Littman, E. D., Ling, J. Q., Ulaner, G. A., Vu, T. H., Dasig, D., Lyon, J., Giudic, L. C., Hoffman, A. R., Behr, B. ELSEVIER SCIENCE INC. 2005: S8–S8
  • Pregnancies and live births after transfer of cryopreserved hatching or hatched blastocysts. 53rd Annual Meeting of the Pacific-Coast-Reproductive-Society Shu, Y., Gebhardt, J., Watt, J., Lyon, J., Jensen, J., Behr, B. ELSEVIER SCIENCE INC. 2005: S20–S20
  • Preliminary experience with low concentration of granulocyte-macrophage colony-stimulating factor: A potential regulator in preimplantation mouse embryo development and apoptosis JOURNAL OF ASSISTED REPRODUCTION AND GENETICS Behr, B., Mooney, S., Wen, Y., Pollan, M. L., Wang, H. B. 2005; 22 (1): 25-32


    To investigate the effects of granulocyte-macrophage colony-stimulating factor (GM-CSF) on the development of preimplantation mouse embryos.Mouse 2-cell embryos were collected and cultured in P-1 medium supplemented with GM-CSF at different concentrations. Using reverse transcription-polymerase chain reaction, expression Bcl-2 and Bax mRNA in blastocyst were evaluated in the GM-CSF group and control group. Apoptosis detection was performed using the in situ apoptosis detection kit in mouse blastocysts. The statistical significance of the data was analyzed using t-test and chi-square test.The development of blastocyst increased to 89% in the addition of GM-CSF (0.125 ng/mL), compared to controlled group (80%). The number of cells staining for apoptosis was lower in GM-CSF group than that in the control group. Bcl-2 expression was found to be upregulated in blastocysts in the GM-CSF supplemented group compared to the control group.These results suggest that GM-CSF might be an important regulator in embryo development.

    View details for DOI 10.1007/s10815-005-0817-9

    View details for Web of Science ID 000227837900006

    View details for PubMedID 15807219

  • Monozygotic twin birth after the transfer of a cleavage stage embryo resulting from a single pronucleated oocyte JOURNAL OF ASSISTED REPRODUCTION AND GENETICS Dasig, D., Lyon, J., Behr, B., Milki, A. A. 2004; 21 (12): 427-429


    To present a case involving the transfer of a single pronucleated oocyte resulting in a monozygotic twin pregnancy.A descriptive case report of a single patient.The patient conceived and was found to have a monochorionic diamnionic pregnancy which resulted in the birth of normal identical twin boys at 32 weeks of gestation.The case report addresses an issue that has not received proper attention in the literature. It illustrates that observing a single PN in an oocyte at fertilization check should not be an absolute deterrent to transferring the resulting embryo even in an older patient with a high FSH level. This report also suggests that single observations, especially at the assessment of fertilization, in the IVF laboratory are limited when evaluating embryo potential and normalcy.

    View details for Web of Science ID 000226417000002

    View details for PubMedID 15704517

  • Maturation, embryo development and clinical outcomes in ICSI treatment cycles with completely immature oocytes at the removal of cumulus cells. 60th Annual Meeting of the American-Society-for-Reproductive-Medicine Shu, Y., Gebhardt, J., Watt, J., Lyon, J., Dasig, D., Behr, B. ELSEVIER SCIENCE INC. 2004: S331–S331
  • Correlation between normal sperm head morphology (NSHM) and sperm binding potential to the sperm hyaluronan-binding assay (HBA (TM)). 60th Annual Meeting of the American-Society-for-Reproductive-Medicine Worrilow, K. C., Lyon, J., Belcak, N., Peters, A. J., Johnston, J. B., Behr, B. ELSEVIER SCIENCE INC. 2004: S95–S95
  • Significance of immature oocytes in intracytoplasmic sperm injection treatment cycles 60th Annual Meeting of the American-Society-for-Reproductive-Medicine Behr, B., Gebhardt, J., Watt, J., Lyon, J., Dasig, D., Shu, Y. ELSEVIER SCIENCE INC. 2004: S293–S294
  • Effects of culture conditions on IVF outcome Serono Symposium on Genesis and Fate of the Preimplantation Empryo Behr, B., Wang, H. ELSEVIER SCIENCE BV. 2004: S72–S76


    Although in vitro fertilization (IVF) success rates have improved over the past decade, multiple pregnancies have become a formidable problem. The solution to this problem seems simple by mandating the reduction in numbers of embryos transferred. However, this is typically not accomplished without a compromise in the pregnancy rate. There have been a number of approaches designed to address high order multiple pregnancies from multi factorial analysis of early cleavage stage embryos to the development of extended culture systems, both of which require manipulations in the culture environment. Manipulations in embryo culture environment may not be benign. Several studies have demonstrated that adverse culture conditions have effects on gene expression and imprinting. Studies have also demonstrated that singleton human IVF babies have lower birth weight and higher incidence of congenital anomalies than natural conception babies. All of these factors need to be considered in relation to long term viability of IVF babies and the Barker hypothesis.

    View details for DOI 10.1016/j.ejogrb.2004.01.016

    View details for Web of Science ID 000222748600015

    View details for PubMedID 15196720

  • The effect of a two-hour, room temperature incubation of human spermatozoa in TEST-yolk buffer on the rate of fertilization in vitro JOURNAL OF ASSISTED REPRODUCTION AND GENETICS Jensen, J. R., Walker, J. H., Milki, A. A., Westphal, L., Behr, B. 2004; 21 (5): 169-173


    To reassess the use of TEST-yolk buffer (TYB) in an in vitro fertilization (IVF) program by comparing fertilization rates achieved in a glucose-free cleavage medium by the standard IVF preparation of sperm versus a 2-h, room temperature incubation of sperm in TYB.Oocytes collected for IVF were randomly split into two groups and inseminated with either TYB-treated sperm or IVF-prepared sperm.Stanford Reproductive Endocrinology and Infertility Center.Fifty couples undergoing IVF with at least 10 mature oocytes.Fertilization rates in vitro.Fertilization rates were significantly higher (p = 0.015) with TYB treatment. The average 2PN fertilization rate was 49.6% (188/379) for the IVF group and 57.4% (221/385) in the IVF with TYB group.A 2-h, room temperature incubation of sperm in TYB produces significantly higher 2PN fertilization rates as compared to standard IVF preparation of sperm in a current generation cleavage medium.

    View details for Web of Science ID 000221941300007

    View details for PubMedID 15279324

  • Igf2 gene imprinting in preimplantation mouse embryos 52nd Annual Meeting of the Pacific-Coast-Reproductive-Society Littman, E. D., Ulaner, G. A., Vu, T. H., Otero, J., Dasig, D., Lyon, J., Gebhardt, J., Giudice, L. C., Behr, B., Hoffman, A. R. ELSEVIER SCIENCE INC. 2004: S14–S15
  • Visualization of atypical hatching of a human blastocyst in vitro forming two identical embryos FERTILITY AND STERILITY Behr, B., Milki, A. A. 2003; 80 (6): 1502-1503
  • Matrix metalloproteinase and tissue inhibitor of matrix metal loproteinase expression in human preimplantation embryos FERTILITY AND STERILITY Wang, H. B., Wen, Y., Mooney, S., Li, H., Behr, B., Polan, M. L. 2003; 80: 736-742


    To examine human embryos at various stages of preimplantation development for simultaneous expression of matrix metalloproteinases (MMPs) and tissue inhibitor of matrix metalloproteinases (TIMPs).mRNAs of specific MMPs and TIMPs were examined in single human embryos, at different stages of preimplantation development, by reverse transcription-polymerase chain reaction (RT-PCR). Single embryo immunohistochemistry was applied to examine the protein expression.University-affiliated IVF-ET program.Couples, attending the university-affiliated IVF-ET program, electing to donate poor prognosis embryos with anomalous morphology.None.The expression of MMP-1, MMP-2, MMP-9, TIMP-1, TIMP-2, and TIMP-3 in preimplantation embryos.The MMP-2 mRNA was expressed consistently during development from one-cell to blastocyst stage. The TIMP-1 and TIMP-3 mRNAs were detected in embryos at all stages; however, in the later preimplantation developmental stages, an increasing proportion of embryos expressing TIMP-1 and TIMP-3 mRNA were noted. The MMP-1, MMP-9, and TIMP-2 mRNAs were detected in only a minority of human embryos studied. Immunohistochemistry showed MMP-1 and TIMP-1 protein expression in preimplantation embryos.The existence of MMP and TIMP mRNA expression in human preimplantation embryos argues for a role for these metalloproteinases and their inhibitors during the process of implantation in humans.

    View details for DOI 10.1016/S0015-0282(03)00782-9

    View details for Web of Science ID 000185603100008

    View details for PubMedID 14505747

  • Comparison of the sex ratio with blastocyst transfer and cleavage stage transfer 58th Annual Meeting of the American-Society-for-Reproductive-Medicine Milki, A. A., Jun, S. H., Hinckley, M. D., Westphal, L. W., Giudice, L. C., Behr, B. SPRINGER/PLENUM PUBLISHERS. 2003: 323–26


    To evaluate the sex ratio in births conceived with blastocyst transfer compared to day 3-ET.A retrospective analysis of IVF patients who became pregnant after blastocyst or cleavage stage transfer at Stanford University Hospital and a literature review were performed.In the day 3-ET group, the male-to-female (M/F) ratio was 157/139 (53%/47%) compared to 97/66 (59.5%/40.5%) in the blastocyst group (P = 0.18). Similar trends have been found in individual studies in the literature but reached statistical significance in only one out of six reports reviewed. The combined data from our study and the literature show a male-to-female ratio of 797/594 (57.3%/42.7%) in blastocyst transfer compared to 977/932 (51.2%/48.8%) in day 3-ET (P = 0.001).Although individual studies may lack power to show an altered sex ratio with blastocyst transfer, the combined data presented in this report do suggest that the M/F ratio is higher with blastocyst transfer compared to cleavage stage transfer.

    View details for Web of Science ID 000184279700007

    View details for PubMedID 12948095

  • Significance of one pronucleus before fertilization FERTILITY AND STERILITY Westphal, L. M., Rosencrantz, M., Behr, B., Milki, A. A. 2003; 79 (4): 1031-1033


    To describe a case of primary infertility associated with oocytes having one pronucleus before fertilization on repeated IVF attempts.Case report.A university-based assisted reproduction unit.A 30-year-old woman with primary infertility and oocytes containing one pronucleus before fertilization.Oocyte donation.Pregnancy.Conceived triplets after transfer of three embryos using donor oocytes.This patient's infertility was likely associated with an oocyte abnormality, as evidenced by the premature formation of one pronucleus before fertilization. In the future, more studies on the appearance of a single pronucleus before fertilization will be needed to determine its overall significance on fertility.

    View details for DOI 10.1016/S0015-0282(02)04852-5

    View details for Web of Science ID 000182045400034

    View details for PubMedID 12749450

  • Granulocyte-macrophage colony-stimulating factor enhances human embryo development to the blastocyst stage: a randomized study. 51st Annual Meeting of the Pacific-Coast-Reproductive-Society Shapiro, B. S., Richter, K. S., Daneshmand, S. T., QUINN, P., Behr, B. ELSEVIER SCIENCE INC. 2003: S15–S16
  • The effect of a two hour, room temperature incubation of human spermatozoa in test-yolk buffer on the rate of fertilization in vitro. 51st Annual Meeting of the Pacific-Coast-Reproductive-Society Behr, B., Lyon, J., Watt, J., Dasig, D., Gebhardt, J., Jensen, J. ELSEVIER SCIENCE INC. 2003: S10–S10
  • Effect of ICSI on subsequent blastocyst development and pregnancy rates 50th Annual Meeting of the Pacific-Coast-Reproductive-Society Westphal, L. M., Hinckley, M. D., Behr, B., Milki, A. A. SPRINGER/PLENUM PUBLISHERS. 2003: 113–16


    To investigate whether ICSI (intracytoplasmic sperm injection) results in decreased blastocyst formation and pregnancy compared to IVF (in vitro fertilization).We performed a retrospective analysis of blastocyst transfer (BT) offered routinely to patients under age 40 with > or = three 8-cell embryos on day 3 and compared IVF to ICSI cycles. Sequential media were used with P1 until day 3, then Blastocyst Medium until day 5/6.There were 131 IVF and 75 ICSI cycles. There was no difference in age, number of oocytes, zygotes, 8-cell embryos, blastocysts on days 5 and 6, or embryos transferred. Progression to blastocyst was similar (78% for IVF and 73% for ICSI) as was the viable pregnancy rate (51.4% for IVF and 55% for ICSI). No cycles failed to form blastocysts.The progression to blastocyst and the likelihood of conceiving a viable pregnancy were unaltered by ICSI. Thus it seems appropriate for programs to offer BT to patients undergoing ICSI using the same inclusion criteria applied to their IVF patients.

    View details for Web of Science ID 000181337000002

    View details for PubMedID 12735386

  • Incidence of monozygotic twinning with blastocyst transfer compared to cleavage-stage transfer FERTILITY AND STERILITY Milki, A. A., Jun, S. H., Hinckley, M. D., Behr, B., Giudice, L. C., Westphal, L. M. 2003; 79 (3): 503-506


    To evaluate the incidence of monozygotic twinning (MZT) in pregnancies conceived after blastocyst transfer compared to cleavage-stage transfer.Retrospective study.University IVF program.All IVF patients with viable pregnancies conceived during a 4-year period.Blastocyst transfer or day 3 ET.Incidence of MZT assessed by transvaginal ultrasound.There were 11 incidences of MZT in 197 viable pregnancies (5.6%) with blastocyst transfer compared to 7 of 357 viable pregnancies (2%) with day 3 ET. In 10 of 18 pregnancies, MZT was observed in the setting of a higher order multiple gestation (6 of 11 for blastocyst transfer and 4 of 7 for day 3 ET). In the day 3 ET group, assisted hatching or intracytoplasmic sperm injection (ICSI) did not increase MZT (4 of 213, 1.9%) compared to cycles without zona breaching (3 of 144, 2.1%). Similarly, in the blastocyst-transfer group, ICSI did not increase the incidence of MZT (4 of 74, 5.5% for ICSI and 7 of 123, 5.7% for non-ICSI IVF).Compared to day 3 ET, blastocyst transfer appears to significantly increase the incidence of gestations with MZT. This information should be taken into account when counseling patients about the pros and cons of extended culture.

    View details for DOI 10.1016/S0015-0282(02)04754-4

    View details for Web of Science ID 000181605000007

    View details for PubMedID 12620430

  • Comparison of blastocyst transfer to day 3 transfer with assisted hatching in the older patient 56th Annual Meeting of the American-Society-for-Reproductive-Medicine Milki, A. A., Hinckley, M. D., Behr, B. ELSEVIER SCIENCE INC. 2002: 1244–47


    To compare cycle outcomes in similar populations of women over 40 who underwent blastocyst transfer compared with women who had day 3 embryo transfer with assisted hatching (ET/AH).Retrospective study. STTING: University hospital-based program.Eighty-six IVF cycles in women ages 40 to 43 years who had more than three eight-cell embryos on day 3. On day 3 of embryo culture, patients chose either to undergo blastocyst transfer or day 3 ET/AH.Pregnancy and cryopreservation rates were recorded.In 48 cycles, blastocyst transfer was performed, and in 38 cycles day 3 ET/AH was performed. There was no statistically significant difference between the blastocyst transfer group and the day 3 ET/AH group with respect to age (41.1 +/- 0.9 years vs. 41.6 +/- 0.8 years), percentage of intracytoplasmic sperm injection cycles (29.2% vs. and 27.6%), number of oocytes (14.9 +/- 5.6 vs. 12.8 +/- 4.0), or number of eight-cell embryos (6.1 +/- 2.2 vs. 5.4 +/- 1.5). Significantly fewers embryos were transferred per cycle with blastocyst transfer (2.6 +/- 1.0) compared with day 3 ET/AH (5.9 +/- 2.0). The viable pregnancy rate was similar in the blastocyst transfer group (29.2%) and in the day 3 ET/AH group (26.3%). Embryos for cryopreservation were available in significantly more cycles in the blastocyst transfer group (52.1%) than in the day 3 ET/AH group (21.1%). Cleavage stage arrest occurred only in one cycle.Blastocyst transfer appears to be as effective as day 3 ET/AH in older patients with good embryos. Higher cryopreservation rate in the blastocyst transfer group may represent an advantage over day 3 ET/AH. Older women may also benefit from the information that extended culture provides them regarding their oocyte quality.

    View details for Web of Science ID 000179837200018

    View details for PubMedID 12477519

  • Non-invasive embryo viability assessments: The influence of media constituents during the first culture interval. 58th Annual Meeting of the American-Society-for-Reproductive-Medicine Dasig, D., Gebhardt, J., Lyon, J., Milki, A. A., Watt, J., Behr, B. ELSEVIER SCIENCE INC. 2002: S285–S285
  • Regulation of cyclooxygenase activity in human endometrial stromal cells by granulocyte-macrophage colony-stimulating factor in vitro. 58th Annual Meeting of the American-Society-for-Reproductive-Medicine Wang, H. B., Wen, Y., Chen, B., Mooney, S., Behr, B., Polan, M. L. ELSEVIER SCIENCE INC. 2002: S220–S221
  • Effect of osteopontin on mouse embryo development in vitro. 58th Annual Meeting of the American-Society-for-Reproductive-Medicine Behr, B., Ben Shlomo, I., Dasig, D., Lyon, J., Polan, M. L., Wang, H. ELSEVIER SCIENCE INC. 2002: S40–S41
  • Follicle development in grafted mouse ovaries after cryopreservation and subcutaneous transplantation AMERICAN JOURNAL OF OBSTETRICS AND GYNECOLOGY Wang, H. B., Mooney, S., Wen, Y., Behr, B., Polan, M. L. 2002; 187 (2): 370-374


    Our aim was to determine the impact of freezing, thawing, and subcutaneous transplantation on follicular development in grafted mouse ovaries.The mice were divided into 3 groups: control (group 1), frozen-thawed grafting (group 2), and frozen-thawed grafting with human menopausal gonadotropin injection (group 3). After freezing and thawing, the ovaries were transplanted into the subcutaneous tissue. Two weeks after transplantation, grafted ovaries and blood samples were collected.Ovaries from group 3 contained significantly more follicles (246 +/- 43 follicles) than group 2(P <.05). The pattern and intensity of Cx37 immunohistochemical staining was similar in all groups. Follicle-stimulating hormone concentrations were significantly decreased in group 2 after ovarian grafting.In mice, gonadotropin treatment before subcutaneous grafting improved the survival of growing follicles. Subcutaneous ovarian transplantation may restore ovarian function and could obviate many of the problems that are related to ovarian banking for humans.

    View details for DOI 10.1067/mob.2002.123606

    View details for Web of Science ID 000177661200029

    View details for PubMedID 12193927

  • Phospholipase A(2) and cyclooxygenase gene expression in human preimplantation embryos JOURNAL OF CLINICAL ENDOCRINOLOGY & METABOLISM Wang, H. B., Wen, Y., Mooney, S., Behr, B., Polan, M. L. 2002; 87 (6): 2629-2634


    Phospholipase A(2) (PLA(2)) and cyclooxygenase (COX) are two key enzymes in PG synthesis; the latter has two forms, COX-1 and COX-2. mRNA was extracted from single preimplantation embryos and examined for PLA(2), COX-1, and COX-2 gene expression by RT-PCR to investigate whether PLA(2) and COX genes are expressed in human preimplantation conceptuses from zygote to blastocyst stage and to compare COX-1 and COX-2 gene expression within the same stage of embryonic development. Expression of PLA(2), COX-1, and COX-2 was detected in 48, 37, and 45%, respectively, of total embryos examined. COX-1 was expressed in approximately 66% of early human preimplantation embryos from zygote to two-cell stage, whereas COX-2 was expressed in about 58% of later stage embryos from eight-cell to blastocyst stage (P < 0.05). Furthermore, COX-2 mRNA and protein were localized to trophectoderm in blastocyst stage embryos. In conclusion, PLA(2), COX-1, and COX-2 are expressed during early human embryonic development and may contribute to the production of PGs such as PGE(2) in human embryogenesis. COX-1 and COX-2 are differentially expressed, with COX-2 being primarily expressed by trophectoderm in late-stage human preimplantation embryos, which may promote embryonic differentiation and implantation.

    View details for Web of Science ID 000176241000033

    View details for PubMedID 12050227

  • Accuracy of day 3 criteria for selecting the best embryos FERTILITY AND STERILITY Milki, A. A., Hinckley, M. D., Gebhardt, J., Dasig, D., Westphal, L. M., Behr, B. 2002; 77 (6): 1191-1195


    To assess the accuracy of day 3 morphologic criteria in identifying the best embryos.Prospective observational study.University IVF program.One hundred cycles in women desiring blastocyst transfer who had > or =3 eight-cell embryos on day 3.On day 3, the embryologist chose the two embryos that would have been transferred that day. On day 5, embryos were examined to determine the best and second-best blastocysts.Accuracy of day 3 picks as measured in culture on day 5, outcome of nontransferred picks, and cryopreservation rate.All cycles reached the blastocyst stage and 73% had cryopreservation. The mean number of blastocysts was 4.8 (3.2 on day 5 and 1.6 on day 6). Neither pick was chosen in 39% of cycles; one pick was transferred in 38%; and both picks were transferred in 23%. Of 116 nontransferred picks, 51 were frozen and 65 arrested, with both picks arresting in 9 cycles. The single best blastocyst was chosen from the picks in 39% of cycles.Morphologic criteria for cleavage-stage embryo selection may fall short when the transfer is limited to two embryos. Culture to blastocyst is warranted in this population to avoid high-order multiples and still be able to choose the two embryos with the highest implantation potential.

    View details for Web of Science ID 000176176000016

    View details for PubMedID 12057727

  • Factors relating to a successful cryopreserved blastocyst transfer program 56th Annual Meeting of the American-Society-for-Reproductive-Medicine Behr, B., Gebhardt, L., Lyon, J., Milki, A. A. ELSEVIER SCIENCE INC. 2002: 697–99


    To review the authors' experience in a successful frozen blastocyst program.Retrospective study.University IVF program.Women of all ages undergoing 64 frozen blastocyst nondonor thaw cycles.Thaw cycles with day 5 or day 6 frozen blastocysts replaced into luteal day 5 endometrium in natural or programmed cycles; cryopreservation of blastocysts by Menezo two-step protocol and Testart slow cool program; thawing by two step thaw protocol.Implantation, clinical pregnancy, and delivery rates.The implantation rate was 16% and was similar with day 5 frozen blastocysts and day 6 frozen blastocysts cycles. The clinical pregnancy rate was 36% and the delivery rate was 27%, with no significant difference between day 5 and 6 blastocysts.Blastocyst cryopreservation is a viable option for patients of all ages and complements fresh blastocyst culture and transfer. The presence of good quality blastocysts for freezing on day 5 and day 6 yields comparable results and is critical for the success of thaw cycles.

    View details for Web of Science ID 000175066400010

    View details for PubMedID 11937118

  • CD146 expression in human preimplantation blastocyst. Wang, H., Wen, Y., Mooney, S., Behr, B., Polan, M. L. ELSEVIER SCIENCE INC. 2001: S203–S203
  • Vascular endothelial growth factor (VEGF) mRNA splice variants are differentially expressed in human blastocysts 16th Annual Meeting of the European-Society-of-Human-Reproduction-and-Embryology (ESHRE) Krussell, J. S., Behr, B., Milki, A. A., Hirchenhain, J., Wen, Y., Bielfeld, P., Polan, M. L. OXFORD UNIV PRESS. 2001: 57–63


    The aim of our study was to detect and characterize mRNA expression of VEGF isoforms VEGF(121), VEGF(145), VEGF(165), VEGF(189), and VEGF(206) in human blastocysts. We recently demonstrated VEGF mRNA expression during human preimplantation embryo development, and further information regarding the alternatively spliced mRNAs resulting in freely secreted proteins or proteins bound to cell surface heparan-sulphate proteoglycans is needed to better understand the process of angiogenesis during implantation. Human blastocysts unsuitable for transfer obtained from the IVF programme at Stanford University were examined by reverse transcription/hemi-nested polymerase chain reaction for their expression of VEGF mRNA splice variants. VEGF mRNA was expressed in 17 out of 19 (89%) blastocysts. Of the 17 blastocysts, VEGF(121) mRNA was detected in 88%, VEGF(145) mRNA in 100%, VEGF(165) mRNA in 71%, and VEGF(189) mRNA in 24% of blastocysts. There was co-expression of mRNA for VEGF(121) and VEGF(145) only in 29% blastocysts, of mRNA for VEGF(165) and VEGF(145) only in 12%, and of mRNA for VEGF(121), VEGF(145) and VEGF(165) in 59% blastocysts. VEGF(206) mRNA could not be detected. In conclusion, we demonstrated that blastocysts express the mRNAs encoding for the free VEGF proteins, enabling the implanting embryo to immediately induce angiogenesis at the implantation site.

    View details for Web of Science ID 000166460800009

    View details for PubMedID 11134361

  • Expression of vascular endothelial growth factor mRNA in human preimplantation embryos derived from tripronuclear zygotes 15th Annual Conference of the European-Society-of-Human-Reproduction-and-Embryology Krussel, J. S., Behr, B., Hirchenhain, J., Wen, Y., Milki, A. A., Cupisti, S., Bielfeld, P., Polan, M. L. ELSEVIER SCIENCE INC. 2000: 1220–26


    To detect the expression of vascular endothelial growth factor (VEGF) mRNA and/or secretion of VEGF protein by human preimplantation embryos.Human preimplantation embryos not suitable for uterine transfer were examined for beta-actin and VEGF mRNA expression. Culture media from normally fertilized and developing preimplantation embryos were assessed for VEGF protein secretion.Clinics and academic research laboratories at the Departments of Obstetrics and Gynecology at the Stanford University, Palo Alto, California and the Heinrich-Heine-University, Düsseldorf, Germany.Couples undergoing IVF by intracytoplasmic sperm injection for various reasons.Six unfertilized oocytes and 33 pathologically fertilized (tripronucleic, 3PN) preimplantation embryos were examined for VEGF mRNA expression, and 16 embryos were examined for VEGF protein secretion.Embryonic expression of VEGF mRNA and VEGF protein as determined by reverse transcription (RT)/nested polymerase chain reaction (PCR) and ELISA.VEGF mRNA and protein could not be detected in unfertilized oocytes. However, 30/33 preimplantation embryos did express VEGF mRNA (11/12 10-to-16-cell embryos, 3/4 morulae, 11/12 early blastocysts, 5/5 hatched blastocysts). The VEGF protein level was below the sensitivity of the ELISA.Production of VEGF may give the embryo the ability to induce neoangiogenesis at the implantation site, thus creating an environment necessary for its survival.

    View details for Web of Science ID 000165897300027

    View details for PubMedID 11119754

  • Blastocyst-ET and monozygotic twinning 15th Annual Conference of the European-Society-of-Human-Reproduction-and-Embryology Behr, B., Fisch, J. D., Racowsky, C., Miller, K., Pool, T. B., Milki, A. A. SPRINGER/PLENUM PUBLISHERS. 2000: 349–51


    To examine the rate of monozygotic twinning associated with blastocyst transfer using commercially available, cell-free culture systems with unmanipulated blastocysts.A retrospective analysis was conducted in multiple private and academic infertility centers throughout the United States, of 199 pregnant patients following in vitro fertilization (IVF) blastocyst embryo transfer (ET). Human embryos obtained through standard IVF stimulation protocols were cultured in commercially available, cell-free media systems and transferred as blastocysts. The main outcome measure was the rate of monozygotic twinning.A total of 199 blastocyst-ET pregnancies were achieved during the study period at the fertility centers examined. Monozygotic twinning was noted in 10/199 (5%) of these pregnancies. All were monochorionic diamnionic.Monozygotic twinning previously has been reported following IVF, especially in relation to assisted hatching. While blastocyst transfer has been available for many years using coculture, there have been no published multicenter reports of monozygotic twinning associated with unmanipulated blastocysts. In a multicenter analysis, a definite increase in monozygotic twinning was seen following blastocyst-ET. We believe this phenomenon is real and that this information should be considered when counseling patients for treatment.

    View details for Web of Science ID 000089429800010

    View details for PubMedID 11042833

  • Vascular endothelial growth factor (VEGF) mRNA isoforms are differentially expressed in human blastocysts Krussel, J. S., Behr, B., Milki, A. A., Wen, Y., Hirchenhain, J., Biefeld, P., Polan, M. L. OXFORD UNIV PRESS. 2000: 93–93
  • Comparison of blastocyst transfer with day 3 embryo transfer in similar patient populations FERTILITY AND STERILITY Milki, A. A., Hinckley, M. D., Fisch, J. D., Dasig, D., Behr, B. 2000; 73 (1): 126-129


    To compare implantation and pregnancy rates (PRs) achieved with blastocyst transfer (BT) and day 3 ET in similar patient populations.Retrospective analysis.Academic infertility center.One hundred consecutive patients <40 years undergoing IVF, each with more than three eight-cell embryos on day 3.Patients used their own eggs for IVF or IVF and intracytoplasmic sperm injection. Embryos were cultured in P1 medium (Irvine Scientific, Santa Ana, CA) until day 3, when they were either transferred or, in the case of embryos for BT, incubated in Blastocyst Medium (Irvine Scientific), followed by transferring on day 5.Implantation and PRs.There were no statistically significant differences in patient age, FSH level, or number of oocytes or zygotes. The BT group had fewer embryos transferred (mean, 2.4) compared with the day 3-ET group (mean, 4.6). The viable PR (cardiac activity at 6-7 weeks was considered indicative of a viable pregnancy) was higher with BT (68%, 34/50) than with day 3 ET (46%, 23/50). The implantation rate was increased with BT (47%, 56 sacs/120 embryos) compared with day 3 ET (20%, 46 sacs/231 embryos).The BT group in our study had higher implantation and PRs compared with the day 3-ET group. Better embryo selection, improved embryo-uterine synchrony, and decreased cervical mucus on day 5 may have accounted for the enhanced outcome. Our data support the use of BT to limit the number of embryos transferred while improving PRs.

    View details for Web of Science ID 000084537500024

    View details for PubMedID 10632426

  • Two-blastocyst transfer has similar pregnancy rates and a decreased multiple gestation rate compared with three-blastocyst transfer FERTILITY AND STERILITY Milki, A. A., Fisch, J. D., Behr, B. 1999; 72 (2): 225-228


    To examine the effect of the number of blastocysts transferred on pregnancy and multiple gestation rates.Retrospective study.Academic infertility center.Patients < 40 years undergoing IVF, with FSH levels of < 15 mIU/mL and more than three eight-cell embryos.Embryos were cultured in P1 until day 3 and then transferred to blastocyst medium. A maximum of three blastocysts were transferred.Pregnancy, multiple gestation, and implantation rates.All 55 patients developed blastocysts and underwent ET. Twenty-four patients had three embryos transferred and 29 patients had two embryos transferred. Two patients had only one embryo each for transfer. There was no difference in the viable pregnancy rate between the two-blastocyst transfer and three-blastocyst transfer groups (62% vs. 58%). In the two-blastocyst transfer group, 39% of pregnancies were multiple gestations (all twin gestations), compared with 79% of pregnancies in the three-blastocyst transfer group (50% twin gestations, 29% triplet gestations). The implantation rate was 47% in both groups.A commercially available, sequential culture system is highly effective for producing viable blastocysts. Two-blastocyst transfer eliminated the risk of triplets while maintaining the same high success rates seen with three-blastocyst ET.

    View details for Web of Science ID 000081761300005

    View details for PubMedID 10438984

  • Human preimplantation embryos express vascular endothelial growth factor mRNA Kruessel, J. S., Wen, Y., Behr, B., Milki, A. A., Hirchenhain, J., Cupisti, S., Bielfield, P., Polan, M. L. OXFORD UNIV PRESS. 1999: 39–39
  • Sibling embryo blastocyst development correlates with the in vitro fertilization day 3 embryo transfer pregnancy rate in patients under age 40 XVI World Congress on Fertility and Sterility/54th Annual Meeting of the American-Society-for-Reproductive-Medicine Fisch, J. D., Milki, A. A., Behr, B. ELSEVIER SCIENCE INC. 1999: 750–52


    To examine the IVF day 3-ET pregnancy rate in patients under 40 with sibling embryo blastocyst development, compared with similar patients without blastocyst formation.Retrospective analysis.Academic infertility center.One hundred twenty-five IVF day 3-ET patients under 40 with sibling embryos for extended culture.Extended culture of nontransferred sibling embryos for blastocyst development.Pregnancy and multiple gestation rates, number of oocytes, embryos formed, and embryos transferred.Thirty-eight percent of patients became pregnant. Forty-eight percent of patients had sibling embryos develop to blastocyst. The blastocyst group had more oocytes retrieved (17.4+/-6.6 versus 14.4+/-5.6), more embryos formed (11.2+/-4.2 versus 8.8+/-3.2), and a higher clinical pregnancy rate (60% versus 18%) than the group without blastocyst development.Blastocyst transfer has been shown to improve implantation rates and reduce the risk of multiple gestations from assisted reproductive technology. Sibling embryo blastocyst development may reflect superior embryo quality, as manifested by increased IVF-ET pregnancy rates. In addition to predicting pregnancy in the current cycle, sibling embryo blastocyst development may provide information about the potential for fresh blastocyst transfer in subsequent cycles and help to identify patients at risk for multiple gestations.

    View details for Web of Science ID 000079355700028

    View details for PubMedID 10202891

  • Preliminary clinical experience with human blastocyst development in vitro without co-culture HUMAN REPRODUCTION Behr, B., Pool, T. B., Milki, A. A., Moore, D., Gebhardt, J., Dasig, D. 1999; 14 (2): 454-457


    This preliminary analysis was designed to quantify blastocyst development of supernumerary embryos without the use of feeder cells, conditioned medium or whole serum. Embryos derived from in-vitro fertilization (IVF) that were not transferred or cryopreserved were included in this study. Ova were harvested for IVF after a standard ovarian stimulation with gonadotrophin-releasing hormone agonist/ human menopausal gonadotrophin (GnRHa/HMG) or follicle-stimulating hormone (FSH). Ova were collected and culture in 150 microliters droplets of P1 medium under mineral oil, in groups at 37 degrees C under 5% CO2, 5% O2, 90% N2 (group A) or under 5% CO2 in air (group B) environment. Embryo transfer was performed 72 h post-harvest. Viable embryos not transferred or cryopreserved were placed in blastocyst medium and cultured for an additional 48 h in 5% CO2 in air. Embryos that exhibited an expanded blastocoelic cavity and well-defined inner cell mass at 120 h were counted. Of 838 supernumerary embryos cultured, 448 (53.5%) reached the expanded blastocyst stage by 120 h of culture. Patients were given the option of cryopreservation at that time. The embryos were cryopreserved using a standard protocol with serial addition of glycerol. Embryos reaching the blastocyst stage after more than 120 h of culture were not included. There was no difference in the proportions of blastocyst development between group A, 217/410 (53.5%) and group B, 231/428 (54%). To date, 16 patients have each had up to three thawed blastocysts transferred, out of whom seven became pregnant. This report demonstrates that a simple system of sequential culture generated acceptable, viable blastocyst development (54%) with supernumerary embryos, without the use of feeder cells, conditioned medium or whole serum. Recognizing the differential metabolic requirements of early and late cleavage stage embryos has enabled the application of a glucose/phosphate-free simple culture medium (P1) for up to 72 h of culture and a complex, glucose-containing medium (blastocyst medium) for subsequent blastocyst development.

    View details for Web of Science ID 000079002900034

    View details for PubMedID 10099993

  • Blastocyst culture and transfer Human Reproduction Behr B. 1999
  • Blastocyst culture and transfer HUMAN REPRODUCTION Behr, B. 1999; 14 (1): 5-6

    View details for Web of Science ID 000078341300003

    View details for PubMedID 10374084

  • Enhancement of motility and acrosome reaction in human spermatozoa: differential activation by type-specific phosphodiesterase inhibitors HUMAN REPRODUCTION Fisch, J. D., Behr, B., Conti, M. 1998; 13 (5): 1248-1254


    Inhibition of sperm phosphodiesterase (PDE) has been shown to increase cAMP concentrations and stimulate motility and the acrosome reaction. While several PDE genes exist in mammals, little is known about the physiological role of PDE forms expressed in human spermatozoa. Using type-selective inhibitors, we identified two of the PDE forms expressed in human spermatozoa and studied their involvement in sperm function. Selective inhibitors of calcium-calmodulin-regulated PDE1 (8-methoxy-isobutyl-methylxanthine) and cAMP-specific PDE4 (RS-25344, Rolipram) were used to study PDE forms in human sperm extracts. 8-MeIBMX and Rolipram/RS-25344 inhibited sperm PDE activity by 35-40 and 25-30% respectively. Subcellular fractionation of the sperm homogenate suggests these pharmacologically distinct forms may be located in separate cellular regions. To evaluate the functional significance of different PDE forms, the effect of type-specific PDE inhibition on sperm motility and the acrosome reaction was examined. PDE4 inhibitors enhanced sperm motility over controls without affecting the acrosome reaction, while PDE1 inhibitors selectively stimulated the acrosome reaction. These data indicate at least two distinct PDE types exist in human spermatozoa. Our findings also support the hypothesis that PDE subtypes affect sperm function by regulating separate pools of cAMP and may ultimately offer novel treatments to infertile couples with abnormal semen parameters.

    View details for Web of Science ID 000074105500034

    View details for PubMedID 9647555

  • Single blastomeres within human preimplantation embryos express different amounts of messenger ribonucleic acid for beta-actin and interleukin-1 receptor type I 13th Annual Meeting of the European-Society-of-Human-Reproduction-and-Embryology Krussel, J. S., Huang, H. Y., Simon, C., Behr, B., Pape, A. R., Wen, Y., Bielfeld, P., Polan, M. L. ENDOCRINE SOC. 1998: 953–59


    Gaining knowledge about the physiological timetable of gene expression during preimplantation embryo development is crucial, and a better understanding of cytokine and growth factor expression in early embryonic development could lead to improved in vitro culture conditions and enhance in vitro fertilization implantation rates. Our aim was to detect the patterns and levels of two messenger ribonucleic acids [mRNAs; beta-actin and interleukin-1 receptor type I (IL-1R tI)] in single human blastomeres by RT-nested PCR and to compare possible variations in the gene expression both between different embryos and in multiple blastomeres within the same embryo. Single blastomeres from nine human tripronucleic preimplantation embryos were examined by one round of RT and two rounds of nested competitive PCR. Beta-actin mRNA was detected in each blastomere, and IL-1R tI mRNA was found in 72% of the blastomeres examined. Beta-actin was expressed at a level of 511-12185 molecules of complementary DNA/blastomere, and IL-1R tI was expressed at a level of 2-290 molecules of complementary DNA/blastomere. Our results suggest that the mRNA pattern of an embryo cannot be reliably quantitated from the mRNA pattern of a single blastomere and therefore imply limitations for the use of this method for preimplantation diagnosis.

    View details for Web of Science ID 000072403500044

    View details for PubMedID 9506755

  • Single blastomeres obtained from human preimplantation embryos express different amounts of mRNA for various genes: implications for future applications of this method for preimplantation diagnosis Kruessel, J. S., Huang, H. Y., Simon, C., Behr, B., Kloodt, A. R., Wen, Y., Bielfeld, P., Polan, M. L. OXFORD UNIV PRESS. 1997: O218–O218
  • Quantification of hexokinase mRNA in mouse blastocysts by competitive reverse transcriptase polymerase chain reaction 44th Annual Meeting of the Pacific-Coast-Fertility-Society Johnson, M. D., BATEY, D. W., Behr, B. OXFORD UNIV PRESS. 1997: 359–65


    Hexokinase (HX), the enzyme that catalyses the initial reaction in glycolysis, is an important enzyme in glucose metabolism during human and mouse embryonic development. In our previous investigations of the genetic activities of HX, we observed an increased incidence of HX gene expression in blastocysts in comparison with morulae, and variability in the incidence of HX gene expression between embryos at the same developmental stages. These observations prompted us to quantify HX mRNA in mouse blastocysts to define the biological significance of the variable gene transcription. We modified our qualitative reverse transcription-nested polymerase chain reaction (RT-nPCR) assay for HX mRNA in single or groups of embryos to quantify HX mRNA by competitive RT-nPCR. HX mRNA was quantified in cohorts of mouse blastocysts cultured in glucose/phosphate-containing human tubal fluid (HTF) media. These blastocysts expressed HX in minute amounts, averaging 1.95 x 10(-18) g of mRNA. This is the first attempt at quantification of single gene mRNA in preimplantation embryos. Further investigations using similar techniques will enable comparative analyses between embryos to be performed to determine the correlation between specific levels of HX mRNA transcripts in individual embryos and embryonic viability and competence for further development and implantation.

    View details for Web of Science ID A1997XT38600011

    View details for PubMedID 9237264

  • Genetic expression of hexokinase and glucose phosphate isomerase in late-stage mouse preimplantation embryos: Transcription activities in glucose/phosphate-containing HTF and glucose/phosphate-free P1 media 42nd Annual Meeting of the Society-for-Gynecologic-Investigation Johnson, M. D., BATEY, D. W., Barro, J. OXFORD UNIV PRESS. 1997: 351–57


    In mouse and human preimplantation development, pyruvate is consumed preferentially during early embryogenesis; however, during the morula and blastocyst stages, glucose is the preferred energy substrate. Studies have suggested that the glycolytic enzymes, hexokinase and glucose phosphate isomerase, are important enzymes in glucose metabolism during these later stages of human and mouse preimplantation development. In order to investigate the genetic activities of these enzymes in late-stage mouse embryos developing in vitro, we analysed hexokinase and glucose phosphate isomerase transcription activities by qualitative RNA assays using reverse transcriptase-nested polymerase chain reaction amplification of individual mouse morulae and early blastocysts incubated in glucose/phosphate-free preimplantation stage one (P1) medium and glucose/phosphate-containing human tubal fluid (HTF) medium. We observed an increased incidence of hexokinase transcripts in the population of blastocysts compared with morulae, and differences in transcript incidence between early blastocysts developing in HTF medium and in P1 medium. In contrast, glucose phosphate isomerase transcripts were consistantly present in all embryos analysed, and appear to be constitutively expressed during late-stage mouse embryogenesis. The different activity patterns of the two glycolytic genes may reflect different mechanisms of gene regulation or differential transcript stability during the later stages of mouse preimplantation development.

    View details for Web of Science ID A1997XT38600010

    View details for PubMedID 9237263

  • Blastocyst culture without co-culture: role of embryo metabolism 10th World Congress of In Vitro Fertilization and Assisted Reproduction Behr, B. MONDUZZI EDITORE. 1997: 145–146


    While the precise timing of the maternal recognition of pregnancy is not known, it is known that the prevention of return to ovarian cyclicity relies on a conceptus-derived signal.In an attempt to identify the first luteotropic signals detectable in the maternal circulation, a sensitive Leydig cell luteotropin bioassay was employed, and data were compared for nine clinically pregnant and nine nonpregnant patients in an in vitro fertilization program. Blood samples were drawn every other day for 10 days after embryo transfer (ET).The first detectable rise in bioactive luteotropin levels occurred between 6 and 8 days post ET. Serum E2 levels increased on the same days. Differences in luteotropin levels between pregnant and nonpregnant patients were significant between days 6 and 8 (P < 0.0001) and between days 8 and 10 (P < 0.002).Based on morphological studies reported by others, bioactive luteotropic signals identified in this study were detectable in the maternal circulation at about the time of trophoblast lacunae coalescing with maternal uterine blood vessels.

    View details for Web of Science ID A1995RK91900013

    View details for PubMedID 7580026