Gene expression microarray profiles of cumulus cells in lean and overweight-obese polycystic ovary syndrome patients.

The aim of this work was to study gene expression patterns of cultured cumulus cells from lean and overweight-obese polycystic ovary syndrome (PCOS) patients using genome-wide oligonucleotide microarray. The study included 25 patients undergoing in vitro fertilization and intra-cytoplasmic sperm injection: 12 diagnosed with PCOS and 13 matching controls. Each of the groups was subdivided into lean (body mass index (BMI) < 24) and overweight (BMI > 27) subgroups. The following comparisons of gene expression data were made: lean PCOS versus lean controls, lean PCOS versus overweight PCOS, all PCOS versus all controls, overweight PCOS versus overweight controls, overweight controls versus lean controls and all overweight versus all lean. The largest number of differentially expressed genes (DEGs), with fold change (FC) |FC| >or= 1.5 and P-value < 0.01, was found in the lean PCOS versus lean controls comparison (487) with most of these genes being down-regulated in PCOS. The second largest group of DEGs originated from the comparison of lean PCOS versus overweight PCOS (305). The other comparisons resulted in a much smaller number of DEGs (174, 109, 125 and 12, respectively). In the comparison of lean PCOS with lean controls, most DEGs were transcription factors and components of the extracellular matrix and two pathways, Wnt/beta-catenin and mitogen-activated protein kinase. When comparing overweight PCOS with overweight controls, most DEGs were of pathways related to insulin signaling, metabolism and energy production. The finding of unique gene expression patterns in cumulus cells from the two PCOS subtypes is in agreement with other studies that have found the two to be separate entities with potentially different pathophysiologies.

EGF increases expression and activity of PAs in preimplantation rat embryos and their implantation rate.

BACKGROUND: Embryo implantation plays a major role in embryogenesis and the outcome of pregnancy. Plasminogen activators (PAs) have been implicated in mammalian fertilization, early stages of development and embryo implantation. As in-vitro developing embryos resulted in lower implantation rate than those developed in-vivo we assume that a reduced PAs activity may be involved. In the present work we studied the effect of EGF on PAs activity, quantity and embryo implantation. METHODS: Zygotes were flushed from rat oviducts on day one of pregnancy and grown in-vitro in R1ECM supplemented with EGF (10 ng/ml) and were grown up to the blastocyst stage. The control groups were grown in the same medium without EGF. The distribution and quantity of the PAs were examined using fluorescence immunohistochemistry followed by measurement of PAs activity using the chromogenic assay. Implantation rate was studied using the embryo donation model. RESULTS: PAs distribution in the embryos was the same in EGF treated and untreated embryos. Both PAs were localized in the blastocysts' trophectoderm, supporting the assumption that PAs play a role in the implantation process in rats.EGF increased the quantity of uPA at all stages studied but the 8-cell stage as compared with controls. The tissue type PA (tPA) content was unaffected except the 8-cell stage, which was increased. The activity of uPA increased gradually towards the blastocyst stage and more so due to the presence of EGF. The activity of tPA did not vary with the advancing developmental stages although it was also increased by EGF. The presence of EGF during the preimplantation development doubled the rate of implantation of the treated group as compared with controls.

Impact of hypnosis during embryo transfer on the outcome of in vitro fertilization-embryo transfer: a case-control study.

OBJECTIVE: To investigate whether hypnosis during ET contributes to successful IVF/ET outcome. DESIGN: Case-control clinical study. SETTING: Academic Fertility and IVF Unit, Soroka Medical Center, Beer-Sheva, Israel. PATIENT(S): Infertile couples undergoing IVF. INTERVENTION(S): Ninety-eight IVF/ET cycles with hypnosis during the ET procedure were matched with 96 regular IVF/ET cycles. MAIN OUTCOME MEASURES: Comparison of clinical pregnancy and implantation rates between the two groups. RESULT(S): We obtained 52 clinical pregnancies out of 98 cycles (53.1%) with an implantation rate of 28% among hypnosis IVF/ET cycles, and 29 out of 96 (30.2%) clinical pregnancies and an implantation rate of 14.4% in the control cycles. Our overall IVF program pregnancy rate for the same period was 32.1%. Logistic regression analysis was performed emphasizing the positive contribution of hypnosis to the IVF/ET conception rates. CONCLUSION(S): This study suggests that the use of hypnosis during ET may significantly improve the IVF/ET cycle outcome in terms of increased implantation and clinical pregnancy rates. Furthermore, it seems that the patients' attitude to the treatment was more favorable.

The effect of women's occupational psychologic stress on outcome of fertility treatments.

OBJECTIVE: The objective of this study was to examine the possible association between women's occupational stress and outcome of fertility treatments. METHODS: A prospective cohort study was performed, including a consecutive group of 75 working women with a female fertility problem attending fertility clinics between the years 1999 and 2000. A structured questionnaire measuring burnout, job strain, and job satisfaction was used. Workload was assessed by number of working hours and shift work. RESULTS: Women who perceived their job as more demanding were less likely to conceive (relative risk [RR], 0.6; 95% confidence interval [CI] = 0.42-0.96). Actual workload, measured by full-time versus part-time job, was found among women who conceived to be significantly associated with less likelihood to successfully complete a pregnancy (RR, 0.3; 95% CI = 0.11-0.96). CONCLUSIONS: An inverse association was found between perceived higher workload and conceiving. The likelihood to deliver after fertility treatment was associated with less working hours.

Cullen's sign following ultrasound-guided transvaginal oocyte retrieval.

OBJECTIVE: To describe two cases of periumbilical hematoma (Cullen's sign) following ultrasound-guided transvaginal oocyte retrieval. DESIGN: Case report. SETTING: University-affiliated teaching hospital. PATIENT(S): Two patients, 34 and 32 years old, who developed Cullen's sign following ultrasound-guided transvaginal oocyte retrieval. INTERVENTION(S): Workup for pancreatitis and coagulopathies. Laparoscopy was preformed in one patient. MAIN OUTCOME MEASURE(S): Resolution of the symptoms. RESULT(S): Pancreatitis, coagulopathy, and massive hemoperitoneum were ruled out. Symptoms resided within two weeks. CONCLUSION(S): The appearance of a periumbilical hematoma (Cullen's sign) following ultrasound-guided transvaginal oocyte retrieval reflects a retroperitoneal hematoma of a benign course.

Uterine leiomyoma among women who conceived following fertility treatment.

OBJECTIVE: The objective was to determine if uterine leiomyoma increases the risk of cesarean section (CS) among women who conceived following fertility treatment. STUDY DESIGN: The study population consisted of all women who conceived after fertility treatment with singleton gestation and who delivered between the years 1988 and 1999 in the Soroka University Medical Center. A comparison was performed between patients with and without uterine leiomyomas. The Mantel-Haenszel procedure was used to obtain the weighted odds ratio (OR) for CS, while controlling for confounding variables. RESULTS: During the study period 1,995 women conceived following fertility treatment. Of these, 63 patients had uterine leiomyomas (3.2%). Women treated for fertility with uterine leiomyomas had statistically significant higher rates of CS than those without uterine leiomyomas (61.9% vs. 28.1%, OR=4.2, 95% CI 2.4-7.2; P<0.001). Stratified analysis (the Mantel-Haenszel technique) was used to control for possible confounders, such as gestational diabetes, hypertensive disorders, maternal age, failure of labor to progress, placental abruption, malpresentation, hydramnios, oligohydramnios, and a previous CS. None of those variables changed the significant association or explained the higher incidence of CS in the uterine leiomyoma group. CONCLUSIONS: Uterine leiomyoma is an independent risk factor for CS among women who conceived following fertility treatment.

Relationship between the duration of sexual abstinence and semen quality: analysis of 9,489 semen samples.

OBJECTIVE: To evaluate the relationship between duration of sexual abstinence and various characteristics of normal and subnormal semen. DESIGN: A retrospective study based on computerized data. SETTING: Fertility and IVF unit at a university medical center. PATIENT(S): Nine thousand, four hundred eighty-nine semen samples from 6,008 patients were analyzed according to the World Health Organization (WHO) manual and grouped according to sperm concentration (10(6)/mL) into severe (0.2-4 x 10(6)), moderate (>4-10 x 10(6)), and mild (>10-19.99 x 10(6)) oligozoospermia, and normozoospermia (> or =20-250 x 10(6)) groups. MAIN OUTCOME MEASURE(S): In each group mean values of semen volume, sperm concentration, percentage of motile sperm and of normal morphology (according to WHO or Kruger criteria), total sperm count, and total motile sperm count per ejaculate were related to duration of abstinence. RESULT(S): Among the 3,506 oligozoospermic samples, the peak mean sperm motility of 30.3% was observed after 1 day of abstinence. Similarly, the mean percentage of normal morphology among mild-moderate oligozoospermic samples (n = 2,260) reached peak values of 7.4%-8.6% between 0-2 days of abstinence. The 5,983 normozoospermic samples showed a significant decrease in the percentage of sperm motility and normal morphology to mean values of 33.1% and 7.0%, respectively, on days 11-14 of sexual abstinence. CONCLUSION(S): Our data challenge the role of abstinence in male infertility treatments and suggest that to present the best possible semen samples, patients with male factor infertility should collect the semen after just 1 day of sexual abstinence. Patients presenting normal sperm analysis or sperm donors for cryopreservation purposes should be advised not to exceed 10 days of sexual abstinence.

Interleukin-6 expression during normal maturation of the mouse testis.

In this study, we examined the cellular origin and the expression levels of interleukin-6 (IL-6) during normal maturation of mouse testis. The levels of IL-6 (protein and mRNA) were higher in testicular homogenates of sexually immature than mature mice. Immunohistochemical staining of testicular tissues of sexually immature and adult mice show that testicular germ cells, at different stages of differentiation, Leydig cells/interstitial cells and peritubular cells express IL-6. Our results demonstrate, for the first time, overexpression of IL-6 in testicular tissues of immature mice, as compared to mature mice, as well as the expression of IL-6 in germ cells of testicular tissues of adult and sexually immature mice. Thus, our results may indicate the involvement of the endocrine system (gonadotropins and testosterone) in the regulation of IL-6, which is involved in the regulation of testicular development, functions and spermatogenesis.

Testicular interleukin-6 response to systemic inflammation.

Spermatogenesis is a highly controlled process of proliferation, meiosis, and differentiation. Systemic infection and chronic inflammation can impair testicular steroidogenesis and spermatogenesis. In this study, we examined the effect of systemic infection--intraperitoneal (i.p.) injection of lipopolysaccharide (LPS)--on the expression levels of IL-6 in the testis of sexually immature and adult mice. IL-6 levels in testicular homogenates of immature mice were significantly higher than in mature mice (both protein and RNA levels), before and after LPS injection. Injection of LPS (i.p.) into mature mice over 3 hours, significantly increased testicular IL-6 protein and mRNA levels (as demonstrated by ELISA and RT-PCR respectively) compared to the control group. Injection of LPS over 24 hours significantly increased IL-6 mRNA expression, but it did not significantly affect IL-6 protein levels in the homogenates. In contrast, stimulation of immature mice with LPS (2, 20 or 100 microg/mL) over 3 hours or LPS (2 or 20 microg/mL) over 24 hours, significantly increased testicular IL-6 (both protein and mRNA expression). The levels of testicular IL-6 (protein) in the homogenates were not significantly increased after stimulation with 100 microg/mL over 24 hours, but they were significantly increased at the mRNA level. Our results demonstrate, for the first time, the over-expression of IL-6 in testicular homogenates of mature and immature mice following systemic inflammation (i.p. injection of LPS). These results suggest the possibility of the involvement of systemic infection/inflammation, through the elevation of testicular IL-6, in testicular functions, which may affect male fertility. Also, high levels of IL-6 during pathological conditions, could play a role in protecting testicular tissue.

Expression of plasminogen activators in preimplantation rat embryos developed in vivo and in vitro.

BACKGROUND: Embryo implantation plays a major role in embryogenesis and the outcome of pregnancy. Plasminogen activators (PAs) have been implicated in mammalian fertilization, early stages of development and embryo implantation. The invasion of trophoblast cells into the endometrium during the implantation process can be blocked by inhibitors of serine proteases, illustrating the role of these enzymes in the invasion process. As in vitro developing embryos resulted in lower implantation rate than those developed in vivo we assume that a reduced PAs activity may lead to it. There is hardly any information regarding qualitative or quantitative differences in expression of PAs in preimplantation embryos, or comparisons between in vivo and in vitro developed embryos. The purpose of this study was to assess the expression of urokinase type (uPA) and tissue type (tPA) plasminogen activators in in vivo and in vitro preimplantation development in rat embryos using immunofluorescence confocal microscopy and computerized image analysis. METHODS: Zygotes, 2-cell, 4-cell, 8-cell, morula and blastocyst stages of development were flushed from the reproductive tract (control groups) of Wistar rats. Zygotes were flushed and grown in vitro to the above mentioned developmental stages and comprised the experimental groups. Immunofluorescence microscopy and computerized image analysis were used to evaluate both qualitative (localization) and quantitative expression of plasminogen activators. RESULTS: uPA and tPA were found to be expressed in rat embryos throughout their preimplantation development, both in vivo and in vitro. While uPA was localized mainly in the cell cytoplasm, the tPA was detected mainly on cell surface and in the perivitelline space. In blastocysts, both in vivo and in vitro, uPA and tPA were localized in the trophectoderm cells. Total uPA content per embryo was higher in the in vivo as compared with the in vitro developed embryos at all stages measured. Blastocyst uPA content was significantly low as compared with the four-cell, eight-cell, and morula stages. Total tPA content was higher in embryos developed in vivo than those developed in vitro except for the 4-cell and 8-cell stages. CONCLUSION: In vitro embryo development leads to lower PAs expression in a stage dependent manner as compared with in vivo developing controls. The enzymes studied vary probably in the ratio of their active and inactive forms as there is no correlation between their content and the activity observed in our previous study. The localization of both PAs in the blastocysts' trophectoderm supports the assumption that PAs plays a role in the implantation process in rats.

Blastocyst-stage embryo transfer in patients who failed to conceive in three or more day 2-3 embryo transfer cycles: a prospective, randomized study.

OBJECTIVE: To compare blastocyst-stage embryo transfers (ETs) with day 2-3 ETs in patients who failed to conceive in three or more day 2-3 IVF/ET cycles. DESIGN: Prospective, randomized. SETTING: Fertility unit in a university medical center. PATIENT(S): Fifty-four patients with an adequate ovarian response underwent oocyte retrievals. The patients were prospectively and randomly divided into blastocyst-stage and day 2-3 ET groups. INTERVENTION(S): Ovarian down-regulation was obtained using GnRH agonist, and controlled ovarian hyperstimulation was achieved using daily administration of gonadotropins. MAIN OUTCOME MEASURE(S): The rate of blastocyst formation, ET cancellations, pregnancies, implantation, multiple gestation, and live births. RESULT(S): The clinical pregnancy rates per oocyte retrieval were 21.7% and 12.9% per blastocyst and day 2-3 ETs, respectively. Although there was a significantly higher implantation rate for blastocyst embryos (21.2%) as compared with 48- to 72-hour embryos (6%), the multiple-pregnancy rate was not significantly different between both groups. An ET cancellation rate of 26% and 6.4% for blastocyst and day 2-3 ETs, respectively, was observed. The presence of two or more 8-cell embryos on day 3 in culture carried a high probability of obtaining blastocysts for transfer. CONCLUSION(S): This prospective randomized study suggests that in patients with an adequate ovarian response who failed to conceive in at least three IVF/ET cycles [1]. transfer of blastocyst-stage embryos carries a significantly higher implantation rate; [2]. the pregnancy rate per oocyte retrieval and ET are higher in the blastocyst-stage group, even if it did not reach statistical significance; [3]. a higher ET cancellation rate was observed in the whole blastocyst-stage group; [4]. the ET cancellation rate was reduced significantly if the decision to proceed to blastocyst transfer was made on day 3 after oocyte retrieval, which is a post hoc conclusion.

Differences in the implantation rates of rat embryos developed in vivo and in vitro: possible role for plasminogen activators.

OBJECTIVE: To assess the participation of urokinase-type (uPA) and tissue-type (tPA) plasminogen activators in embryo development and implantation. DESIGN: The study was set to compare the activities of PAs in embryos developed in vivo and in vitro and their implantation rates. Endometrial PA activity was studied as well. SETTING: University laboratories. PATIENT(S): None. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Plasminogen activator activity in embryos. RESULT(S): Whereas tPA activity in in vivo-developed embryos decreased from the zygote to the blastocyst stages, it did not change in embryos developed in vitro. The activity of uPA was lowest in two-cell-stage embryos, both in vivo and in vitro, and increased as embryos developed into blastocysts. The activity of uPA in the in vitro-developed embroys was lower in all developmental stages as compared with those developed in vivo. Whereas endometrial tPA activity did not change during the preimplantation period, uPA activity increased gradually toward the time of implantation. The implantation rate of the in vitro-developed embryos was much lower than that of those developed in vivo. CONCLUSION(S): Taken together, these data indicate that in vitro embryo development leads to lower PA activities as compared with in vivo development. The increased activity of uPA toward the blastocyst stage supports uPA's role in the implantation process. Moreover, the rise in endometrial uPA might also indicate its importance in this process.

Over expression of interleukin-1alpha, interleukin-1beta and interleukin-1 receptor antagonist in testicular tissues from sexually immature mice as compared to adult mice.

The levels of IL-1alpha, IL-1beta and IL-1Ra were higher in homogenates of testicular tissue from sexually immature than those from mature mice. Immunohistochemical staining of testicular tissues from sexually immature and adult mice show that differentiated germ cells express higher levels of IL-1alpha compared to Sertoli cells and Leydig cells/interstitial cells. Peritubular cells of sexually immature and adult mice did not express IL-1alpha. Testicular tissue cells of adult mice showed high levels of expression of IL-1beta, mainly in the cytoplasm and nucleus of the spermatogonia and in spermatocytes. Sertoli cells and Leydig/interstitial cells were also highly stained for IL-1beta. However, peritubular cells did not express IL-1beta. On the other hand, testicular tissue cells from sexually immature mice, showed high levels of IL-1beta, mainly in spermatocytes. Spermatogonia showed low levels of IL-1beta expression. Also, high levels of IL-1beta expression were detected in Leydig/interstitial cells. Peritubular cells clearly showed IL-1beta expression. Testicular tissue cells from adult mice, showed IL-1Ra expression in spermatogonia, Sertoli and Leydig/interstitial cells. IL-1Ra expression was clearly present in the Golgi apparatus of spermatogonia and Sertoli cells. However, peritubular cells did not show IL-1Ra expression. Testicular tissue cells from sexually immature mice, also showed high levels of IL-1Ra expression mainly in the cytoplasm and nucleus of the spermatogonia and Sertoli cells. In addition, Leydig/interstitial cells and peritubular cells also expressed IL-1Ra. Our results demonstrate, for the first time, the expression of IL-1beta in germ and Sertoli cells, and IL-1Ra in Leydig/interstitial cells of testicular tissues from adult and sexually immature mice, under in vivo conditions. In addition, the relative elevated levels of the IL-1 system in the testis of immature mice compared to mature mice may indicate its involvement in the spermatogenesis.

Effect of occupational exposures on male fertility: literature review.

The present review was aimed to determine the influence of working conditions, occupational exposures to potential chemical and physical reproductive toxic agents and psychological stress during work on male fertility. Significant associations were reported between impaired semen parameters and the following chemical exposures: metals (lead, mercury), pesticides (dibromochlorophane, 2,4-dichlorophenoxyacetic acid), ethylene glycol ethers and estrogens. The following physical exposures were shown to deteriorate sperm parameters: radiation (both ionized and microwaves) and heat. Psychological distress has another important contribution to infertility. Several studies indicated that stress has a negative impact on sperm parameters. Occupational parameters should be an important part of history taking among patients attending infertility clinics.

Potential association between male infertility and occupational psychological stress.

The purpose of this work was to investigate the influence of working conditions, occupational exposures to potential reproductive toxic agents, and psychological stress on male fertility. The study population consisted of 202 consecutive male patients attending a fertility clinic. Of those, 106 patients had attended the clinic because of a male infertility problem (case group), 66 patients had attended the clinic because of a female infertility problem (control group), and 30 patients had a combined infertility problem (male and female). Male infertility was associated with working in industry and construction as compared with other occupations (78.6% vs 58.3%, P = 0.044). Industry and construction workers were of lower educational level than the other workers (mean: 12.1 vs 13.4 years, P = 0.021). These patients also tended to smoke more than the other workers (OR = 2.53, 95% CI = 1.08 to 5.98), more often worked in shifts (OR = 3.12, 95% CI = 1.19 to 8.13), reported physical exertion in work (OR = 3.35, 95% CI = 1.44 to 7.80), and were more exposed to noise and welding (OR = 3.84, 95% CI = 1.63 to 9.14, OR = 4.40, 95% CI = 1.11 to 1.76, respectively). Male infertility (case group) was found to be statistically related to higher marks in all four measures of burnout as compared with the controls. The largest difference was obtained in the measure of cognitive weariness (mean: 2.9 vs 2.1, P < 0.001). In a multiple logistic regression analysis, industry and construction jobs (adjusted OR = 2.2, 95% CI 1.2 to 2.7) and cognitive weariness (adjusted OR = 1.8, 95% CI = 1.03 to 4.6) were found to be independent risk factors for male infertility problems. Male infertility was independently associated with industry and construction jobs as well as job burnout.