Higher FSH Concentrations Research Articles

Hemostatic Factors and Estrogen during the Menopausal Transition

It has been speculated that gender differences in cardiovascular disease (CVD) mortality can be attributed to the effects of estrogens on inflammation and hemostatic marker profiles. Therefore, we evaluated endogenous hormone concentrations, menopause transition stages, and adoption of exogenous hormone use in relation to hemostatic and inflammation marker concentrations in women. Longitudinally, we studied 3302 participants from the Study of Women's Health Across the Nation, aged 42-52 yr at baseline and self-identified as African-American (28%), Caucasian (47%), Chinese (8%), Hispanic (8%), or Japanese (9%). Serum samples from baseline and years 2001, 2003, and 2005 were assayed for estradiol and FSH. Hormone concentrations were related to CVD markers, including fibrinogen, factor VII-c, plasminogen activator inhibitor-1 (PAI-1), tissue plasminogen activator, and human serum C-reactive protein (hsCRP). Lower estradiol levels were associated with higher levels of PAI-1 and tissue plasminogen activator, but there were no significant relationships with fibrinogen, factor VII-c, or hsCRP. Higher FSH concentrations were associated with higher PAI-1 and factor VII levels, but lower fibrinogen and hsCRP levels. Transitions from premenopause and early perimenopause to postmenopause were not associated with significant differences in levels of hemostatic factors. The hsCRP concentrations were approximately 25% higher, and the PAI-1 concentrations approximately 20% lower among women who initiated hormone therapy, compared with nonusers. Endogenous estrogens may reduce CVD risk via modulation of fibrinolytic factors, but not coagulation or inflammatory markers. Notably, conclusions derived from studies of exogenous hormones and CVD risk may not parallel or explain the effects of endogenous hormones or perimenopausal hormone changes on CVD risk.

Ovulation induction with letrozole in gonadotrophin stimulation resistant women undergoing in vitro fertilization

To assess the clinical outcomes of aromatase inhibitors for stimulating poor responder women undergoing in vitro fertilization (IVF) protocol in whom high doses gonadotrophins treatment was unsuccessful. Retrospective analysis. 37 IVF patients with high basal concentrations of FSH(>13 mIU/mL) who had shown no response to vigorous gonadotrophin stimulation are reported. Patients were offered letrozole, 5 mg/day for 5 consecutive days starting on day 3 of the menstrual cycle. Ultrasonography was performed before initiation of treatment, on day 9 of menstrual cycle, and as required thereafter until the dominant follicle reached 18 mm in diameter. Ovulation was triggered with 10,000 IU of hCG, and oocyte pick-up was performed 34–36 hours later. Sixteen cycles (42%) were canceled because of no ovarian response. Five patients (24%) did not have IVF because of no oocyte retrieval. And four patients did not have ET because of failed fertilization. Twelve patients become an embryo transfer. Only one patient was pregnant. The clinical pregnancy rate was 1 of 12 (9%) per embryo transfer. Poor responders undergoing controlled ovarian hyperstimulation with letrozole in cycles with high basal FSH and who are vigorous gonadotrophin stimulation resistant will often show adequate ovarian response and reach oocyte retrieval and ET. The pregnancy rates, however, remain low. This intervention comparing to the cost of gonadotrophin protocols may have future implications for the treatment of poor responders.

Overview of the response of anoestrous ewes to the ram effect.

The present review summarises knowledge of the reproductive response of anoestrous ewes to the introduction of rams - in other words, the ram effect. The ovarian and endocrine response, the factors that determine whether ewes will respond or not (associated with both the stimulus and the receptivity of the ewes) and some aspects of practical management are discussed. Information on the use of the ram effect to stimulate post-partum, prepubertal and cyclic ewes is also given. New insights are provided on ovarian response patterns, including recently collected information on luteal responses. The existence of delayed ovulation (5-7 days after the introduction of the rams) followed by luteal phases of normal or short length, luteal cysts and luteinised follicles is reported after scanning the ovaries with ultrasound. Endocrine parameters for depth of anoestrus, such as LH pulsatility and FSH concentrations, and how the concentrations of these hormones should be considered are discussed. Particular attention is paid to the observation of spontaneous, higher LH pulsatility and higher FSH concentrations in anoestrous ewes that respond to rams with luteal phases than in those that fail to respond. The use of progestogen priming and single progestogen administration and the possible advantages for synchronisation of oestrus are also discussed. Other factors that should be considered before the ram effect is applied, such as the strength of the stimulus and some practical considerations, are also included.

Cryopreservation of pig granulosa cells: effect of FSH addition to freezing medium

We cryopreserved swine granulosa cells by a slow cooling rate system; FSH was added to the freezing medium to test its effectiveness in protecting the cells. After thawing, proliferative activity, viability, steroidogenesis and apoptosis were tested; moreover, we determined heat shock protein (HSP70) production, to investigate the recovery from stress and superoxide dismutase (SOD) and catalase activity to evaluate a possible impairment of the antioxidant pathway. E2 production was enhanced by cryopreservation in particular with FSH; on the contrary, P4 production was inhibited by the freezing process in particular without FSH. Only the higher FSH concentration (10 ng/ml) stimulated steroid secretion in freshly collected cells; P4 production by cells cryopreserved in the presence and in absence of FSH was increased by both 5 and 10 ng/ml while the lowest concentration was effective in stimulating E2 production only when FSH was added to freezing medium. Freezing did not modify proliferative activity, while apoptosis was higher in frozen than in fresh cells. HSP70 production was lower in cells cryopreserved in presence of FSH, whose antioxidant metabolism was also conserved: SOD and catalase activities were similar to control. In conclusion, cryopreservation does not seem to markedly affect granulosa cells, in particular if they are frozen in presence of FSH; the gonadotrophin somehow improves their performances after thawing, probably stimulating E2 production and the antioxidant metabolism.

Overview of the response of anoestrous ewes to the ram effect

The present review summarises knowledge of the reproductive response of anoestrous ewes to the introduction of rams – in other words, the ram effect. The ovarian and endocrine response, the factors that determine whether ewes will respond or not (associated with both the stimulus and the receptivity of the ewes) and some aspects of practical management are discussed. Information on the use of the ram effect to stimulate post-partum, prepubertal and cyclic ewes is also given. New insights are provided on ovarian response patterns, including recently collected information on luteal responses. The existence of delayed ovulation (5–7 days after the introduction of the rams) followed by luteal phases of normal or short length, luteal cysts and luteinised follicles is reported after scanning the ovaries with ultrasound. Endocrine parameters for depth of anoestrus, such as LH pulsatility and FSH concentrations, and how the concentrations of these hormones should be considered are discussed. Particular attention is paid to the observation of spontaneous, higher LH pulsatility and higher FSH concentrations in anoestrous ewes that respond to rams with luteal phases than in those that fail to respond. The use of progestogen priming and single progestogen administration and the possible advantages for synchronisation of oestrus are also discussed. Other factors that should be considered before the ram effect is applied, such as the strength of the stimulus and some practical considerations, are also included.

Overcoming poor in vitro nuclear maturation and developmental competence of domestic cat oocytes during the non-breeding season

The domestic cat experiences circannual variations in ovarian activity and intrafollicular oocyte quality. One result is poor nuclear and cytoplasmic maturation during in vitro maturation (IVM) conducted during the annual non-breeding season (July through November). In an attempt to overcome this seasonal phenomenon immature oocytes were collected from July through November and cultured in a conventional IVM medium (IVM1) or in IVM1 supplemented with different FSH concentrations and antioxidant (ascorbic acid or cysteine). Nuclear status of oocytes was assessed after IVM or IVF. Embryo stage and blastocyst quality were evaluated after 7 days of in vitro culture. Although the addition of antioxidant alone had no effect, the presence of 10 microg FSH ml(-1) improved nuclear maturation (75.4+/-4.1% versus 48.7+/-8.8% in IVM1; P<0.05) and fertilization success (47.9+/-3.2% versus 35.0+/-5.1% in IVM1; P<0.05). Furthermore, developmental competence of fertilized oocytes was enhanced (P<0.05) only in the presence of ascorbic acid (30.6+/-6.7%) or cysteine (33.6+/-5.1%) compared with IVM1 (8.1+/-8.8%). Consequently, blastocyst yield (17% of total oocytes cultured) was highest when oocytes were matured in medium containing higher FSH concentration and antioxidants. The results of this study demonstrate that meiotic and developmental competences are inherent to the immature cat oocyte collected during the non-breeding season. However, appropriate mechanisms (perhaps seasonal variation in FSH receptors or lack of antioxidant capacity of the cumulus-oocyte complex) are inadequate during this period of gonadal quiescence. Regardless, this compromised oocyte function during the non-breeding season can be overridden by altering in vitro culture conditions to include supplemental FSH and antioxidant.

Role of oestradiol in growth of follicles and follicle deviation in heifers.

Follicle deviation is characterized by continued growth of the largest (developing dominant) follicle and reduced growth of the smaller (subordinate) follicles. The aim of the present study was to test the following hypotheses: (1). oestradiol contributes to the depression of circulating FSH encompassing follicle deviation and (2). oestradiol plays a role in the initiation of deviation. Heifers were treated with progesterone (n = 5) or antiserum against oestradiol (n = 7) or given no treatment (control; n = 6). On the basis of previous studies, progesterone treatment would decrease LH and thereby the circulatory and intrafollicular concentrations of oestradiol and the antiserum would reduce the availability of oestradiol. Progesterone was given in six 75 mg injections at 12 h intervals beginning when the largest follicle of wave 1 first reached >or=5.7 mm (t = 0 h). Oestradiol antiserum (100 ml) was given in a single injection at t = 12 h. Follicles of the wave were defined as F1 (largest) and F2, according to the diameter at each examination. Blood samples were collected at 12 h intervals during t = 0-72 h. Treatment with progesterone lowered the circulatory concentrations of LH by 12 h after the start of treatment (P < 0.05), and concentrations remained low compared with those of controls during the treatment period. Treatment with oestradiol antiserum had no effect on LH. Both progesterone and the antiserum treatments increased the FSH concentrations compared with controls (P < 0.05), which supports the first hypothesis. The interval from t = 0 h to the beginning of deviation was longer in the progesterone- (51.0 +/- 7.6 h; P < 0.06) and antiserum (51.4 +/- 6.3 h; P < 0.05)-treated groups than in the controls (38.0 +/- 3.7 h), which supports the second hypothesis. There was no difference among groups in the diameters of F1 and F2 at deviation. Reduced diameter (P < 0.05 or P < 0.06) of both F1 and F2 occurred in both the progesterone- and antiserum-treated groups at t = 36 h and 48 h, compared with controls. Follicle retardation occurred in both the progesterone- and antiserum-treated groups despite the high FSH concentrations, whereas LH was altered only in the progesterone-treated group. Therefore, the follicle effect can be attributed to inadequate intrafollicular oestradiol. This interpretation implies a functional local role for oestradiol in the deviation process, independent of the systemic negative effect on FSH.

Aging women with polycystic ovary syndrome who achieve regular menstrual cycles have a smaller follicle cohort than those who continue to have irregular cycles

Objective To examine whether follicle loss due to ovarian aging is responsible for the occurrence of regular menstrual cycles in aging women with polycystic ovary syndrome (PCOS), the size of the FSH-sensitive follicle cohort was estimated by the exogenous follicle-stimulating hormone ovarian reserve test (EFORT) and related to the follicle count as measured by ultrasound. Design Prospective study. Setting Reproductive endocrinology unit of an academic medical center. Patient(s) Twenty-seven aging women with PCOS (35.8–49.4 years): 20 with regular menstrual cycles and 7 with oligomenorrhea or amenorrhea. Intervention(s) EFORT and transvaginal ultrasound. Main outcome measure(s) Baseline (cycle day 2, 3, or 4) FSH, androstenedione (A), T, E 2, and inhibin B levels, the E 2 and inhibin B increment after the EFORT, and the follicle count. Result(s) After correction for the body mass index (BMI), the inhibin B increment was higher in the irregular menstrual group, but the E 2 increment did not differ significantly between the two groups. Ultrasound showed a median follicle count of 8.5 (4.0–18.0) in women with regular menstrual cycles (n = 16), compared with 18.0 (8.0–35.0) in irregularly menstruating women (n = 7). The follicle count was significantly correlated to the FSH-induced E 2 increment (r = 0.656) as well as to the inhibin B increment (r = 0.654). The regularly menstruating group was significantly older, had a higher basal FSH concentration, and had lower androgens than the irregularly menstruating group. Conclusion(s) The smaller follicle count, the older age, the higher FSH concentration, and the lower FSH-induced inhibin B increment found in women with PCOS and a regular menstrual cycle confirm that a decrease in the size of the follicle cohort due to ovarian aging is largely responsible for the regular menstrual cycles in aging PCOS women.

Inter-relationships among gonadotrophins, reproductive steroids and inhibin in freemartin ewes.

Freemartins are sterile XX/XY chimaeras that occur as a result of placental fusion between male and female fetuses during early pregnancy. Freemartins occur predominantly in cattle, although the prevalence of ovine freemartinism is increasing. In this study, the reproductive endocrinology of ovine freemartins was compared with that of normal sheep. Freemartins had significantly (P < 0.001) higher basal concentrations of LH and FSH than did normal ewes or rams, although the response of LH to GnRH (10 microg) was similar in freemartins, ewes and rams. Resting concentrations of oestradiol were similar in freemartins and ewes and were increased in both after eCG administration. Testosterone concentrations were higher in freemartins than in ewes, but were unresponsive to GnRH or eCG. Administration of 62.5 mg progesterone or 25 lg oestradiol twice a day for 3 days suppressed LH concentrations to baseline values in freemartins, ewes and rams. In ewes, 500 microg oestradiol administered twice a day caused preovulatory surges in LH concentrations, but suppressed LH in freemartins to baseline values. Thus, LH secretion can potentially be regulated in freemartins by gonadal steroids. FSH concentrations in freemartins were not suppressed by doses of inhibin that were effective in ewes and rams. Therefore, freemartins behave in part like castrated animals, as they have high basal concentrations of LH and FSH, which can be stimulated by GnRH and suppressed by gonadal steroids. Conversely, inhibin does not suppress FSH concentrations in freemartins, and freemartins have circulating concentrations of steroids intermediate between those of castrated and normal animals.

In-vitro role of leptin on ovarian granulosa cell steroidogenesis.

Objective: Leptin receptor localized in ovary and testis suggests a likely target for leptin action. We tried to determine the biologic effects and role of leptin in vitro on granulosa cell function. Design: Controlled prospective study. Materials/Methods: Serum and follicular fluid leptin levels, BMI determination, and granulosa cells obtained during oocyte pick-up in 20 patients (mean age 34.0 ± 3.3 yrs) undergoing ART. Following culture with androstenedion (0.1 mM), at least 100 000 granulosa cells in each well were exposed to different concentrations of FSH (0.1, 1.0, 10, 100 ng/ml) with IGF-1 (30 ng/ml) and leptin (3 ng/ml). The second group of cells were exposed to increased concentrations of leptin with FSH (1.0 ng/ml) and IGF-1 (30 ng/ml). Control group was incubated with culture medium only. Estrogen and progesterone levels are measured by RIA in culture media. Results: Serum and follicular fluid leptin levels were similar among the patients (19.8 ± 8 and 17.6 ± 6.6 ng/ml). Leptin levels are correlated with BMI being greater in obese patients (p = 0.03). No correlation was found in relation to the factors leading to infertility. While leptin did not alter the additive effects of FSH and IGF in low FSH concentrations, the stimulatory effect of IGF-1 was blunted in high FSH concentrations. The estrogen levels which was increased with maximum FSH/IGF-1 concentrations are decreased around 35% with leptin. Progesterone levels did not decreased based on different concentrations of leptin. Conclusions: Our results support leptin affects granulosa cells more than luteinized cells, especially in those with greater BMI. Therefore, in women with increased leptin levels such as PCOS, the resultant defect in IGF sensitization in FSH stimulation of granulosa cells may alter its development. Supported by: SSK Tpecik/Center for Womens Health & Research.

The effect of FSH on male germ cell survival and differentiation in vitro is mimicked by pentoxifylline but not insulin.

High concentrations of FSH have been shown to boost in-vitro differentiation of germ cells from men with normal spermatogenesis and from some patients with in-vivo maturation arrest. This study shows that the differentiation-promoting effect of FSH is connected to protection against germ cell apoptosis and that both effects can be mimicked by the intracellular cyclic AMP (cAMP)-elevating drug pentoxifylline. On the other hand, a high concentration of insulin, supposed to act at the insulin-like growth factor I receptor, did not exert any effect either on differentiation or apoptosis of germ cells in vitro. These data show that the in-vitro effects of supraphysiological concentrations of FSH on human spermatogenesis are mediated by the classical FSH signal transduction pathway involving cAMP as a second messenger. Pentoxifylline may thus be useful as an alternative means for intracellular cAMP elevation in men with high circulating FSH concentrations leading to desensitization of the FSH receptor.

Immediate and delayed effects of heat stress on follicular development and its association with plasma FSH and inhibin concentration in cows

The aim of this study was to characterize the immediate effects of heat stress on plasma FSH and inhibin concentrations, and its involvement in follicular dynamics during a complete oestrous cycle, and to examine a possible delayed effect of heat stress on follicular development. Holstein dairy cows were oestrous synchronized and randomly assigned to either cooled (n = 7) or heat-stressed (n = 6) treatment groups. During a complete oestrous cycle, control cows, which were cooled, maintained normothermia, whereas heat-stressed cows, which were exposed to direct solar radiation, developed hyperthermia. At the end of this oestrous cycle (treated cycle), both groups were cooled and maintained normothermia for the first 10 days of the subsequent oestrous cycle. Throughout this period, follicular development was examined by ultrasonography, and plasma samples were collected. During the second follicular wave of the treated oestrous cycle, a significantly larger cohort of medium sized follicles (6-9 mm) was found in heat-stressed cows than in cooled cows (P &lt; 0.05). The enhanced growth of follicles in this wave in heat-stressed cows was associated with a higher plasma FSH increase which lasted 4 more days (days 8-13 of the oestrous cycle; P &lt; 0.05), and coincided with a decrease in the plasma concentration of immunoreactive inhibin (days 5-18 of the oestrous cycle; P &lt; 0.05). During the follicular phase (days 17-20 of the treated cycle), heat-stressed cows showed an increase in the number of large follicles (&gt;/= 10 mm), and the preovulatory plasma FSH surge was significantly higher in heat-stressed cows than in cooled cows (P &lt; 0.01). The effect of heat stress was also observed during the first follicular wave of the subsequent cycle: the postovulatory plasma FSH concentration was higher (P &lt; 0.01), but fewer medium follicles developed, and the first follicular wave decreased at a slower rate in previously heat-stressed cows than in cooled cows (0.40 and 0.71 follicles per day, respectively). This study shows both immediate and delayed effects of heat stress on follicular dynamics, which were associated with high FSH and low inhibin concentrations in plasma. These alterations may have physiological significance that could be associated with low fertility of cattle during the summer and autumn.

Immediate and delayed effects of heat stress on follicular development and its association with plasma FSH and inhibin concentration in cows

The aim of this study was to characterize the immediate effects of heat stress on plasma FSH and inhibin concentrations, and its involvement in follicular dynamics during a complete oestrous cycle, and to examine a possible delayed effect of heat stress on follicular development. Holstein dairy cows were oestrous synchronized and randomly assigned to either cooled (n = 7) or heat-stressed (n = 6) treatment groups. During a complete oestrous cycle, control cows, which were cooled, maintained normothermia, whereas heat-stressed cows, which were exposed to direct solar radiation, developed hyperthermia. At the end of this oestrous cycle (treated cycle), both groups were cooled and maintained normothermia for the first 10 days of the subsequent oestrous cycle. Throughout this period, follicular development was examined by ultrasonography, and plasma samples were collected. During the second follicular wave of the treated oestrous cycle, a significantly larger cohort of medium sized follicles (6-9 mm) was found in heat-stressed cows than in cooled cows (P < 0.05). The enhanced growth of follicles in this wave in heat-stressed cows was associated with a higher plasma FSH increase which lasted 4 more days (days 8-13 of the oestrous cycle; P < 0.05), and coincided with a decrease in the plasma concentration of immunoreactive inhibin (days 5-18 of the oestrous cycle; P < 0.05). During the follicular phase (days 17-20 of the treated cycle), heat-stressed cows showed an increase in the number of large follicles (>/= 10 mm), and the preovulatory plasma FSH surge was significantly higher in heat-stressed cows than in cooled cows (P < 0.01). The effect of heat stress was also observed during the first follicular wave of the subsequent cycle: the postovulatory plasma FSH concentration was higher (P < 0.01), but fewer medium follicles developed, and the first follicular wave decreased at a slower rate in previously heat-stressed cows than in cooled cows (0.40 and 0.71 follicles per day, respectively). This study shows both immediate and delayed effects of heat stress on follicular dynamics, which were associated with high FSH and low inhibin concentrations in plasma. These alterations may have physiological significance that could be associated with low fertility of cattle during the summer and autumn.

In-vitro spermatogenesis resumption in men with maturation arrest: relationship with in-vivo blocking stage and serum FSH.

We have shown previously that germ cells recovered from some men with maturation arrest can resume spermatogenesis in vitro and give rise to late elongated spermatids. This study relates the ability of germ cells to differentiate in vitro to the stage at which spermatogenesis is blocked in vivo and to the patient's serum FSH concentration. The presence of germ cells at different stages of spermatogenesis was assessed, before and after culture, by classical cytology, by fluorescence in-situ hybridization and by immunocytochemistry with a germline-specific marker. The proportion of cases of maturation arrest at the primary spermatocyte, secondary spermatocyte and spermatid stage in which in-vitro resumption of meiosis was achieved was 24.3% (9/37), 100% (3/3) and 51.1% (23/45) respectively. Serum FSH concentrations were higher than normal in most cases. However, lower values were measured in patients in whom in-vitro spermatogenesis was achieved compared with those in whom no progression was detected. These data show that, under the conditions of this study, germ cells from men with very high serum FSH concentrations (>20 IU/l) are less likely to resume spermatogenesis in vitro than those coming from men with only moderate increase (10-20 IU/l).

Maintenance of oestradiol production and expression of cytochrome P450 aromatase enzyme mRNA in long-term serum-free cultures of pig granulosa cells.

Studies were carried out to investigate the conditions required for maintenance of aromatase activity and expression in long-term cultures of pig granulosa cells. Cells from large (> 2 mm) and small (< or = 2 mm) follicles were cultured at 37 degrees C with 5% CO2 in McCoys 5a medium supplemented with 0.1% (w/v) BSA, testosterone (100 micrograms l-1), insulin (10 micrograms l-1) and long R3 insulin-like growth factor I (IGF-I) (100 micrograms l-1). Cells were cultured with five concentrations of USDA pFSH-I-2 (0-100 micrograms l-1) for 48, 96 or 144 h with or without fetal calf serum (FCS). The number of cells and oestradiol, progesterone and inhibin production were measured. In marked contrast to oestradiol production from cells cultured in plates precoated with FCS, 1 microgram FSH l-1 was optimal for the maintenance of high oestradiol production by granulosa cells from large follicles after 144 h of serum-free culture. Culture with FCS promoted cell proliferation, reduced oestradiol production, and supported FSH-dependent (P < 0.01) increased progesterone and inhibin production indicating cellular luteinization. Northern blot analysis of total RNA from cells cultured with 1 microgram FSH l-1 detected 2.5 and 1.8 kb transcripts encoding aromatase cytochrome P450 (P450arom) and cholesterol side-chain cleavage cytochrome P450 (P450scc), respectively. Transcript expression was hormone sensitive, irrespective of the presence of FCS. High concentrations of FSH (100 micrograms l-1) stimulated expression of P450scc, but inhibited P450arom expression as the cells luteinized after 144 h of culture. This serum-free system, which maintains the aromatase enzyme complex, is fundamental if physiologically relevant observations are to be made of the mechanisms regulating follicle hierarchy development from long-term cultures of pig cells.

Human Spermatogenesis in Vitro: Respective Effects of Follicle-Stimulating Hormone and Testosterone on Meiosis, Spermiogenesis, and Sertoli Cell Apoptosis

In spite of the availability of abundant data about in vitro spermatogenesis in laboratory animals, studies on human in vitro spermatogenesis are scarce. This study employed a relatively simple culture system, involving all cell types of seminiferous tubules, to analyze the effects of FSH and testosterone (T) on different characteristics of human germ and Sertoli cells in culture. By using fluorescence in-situ hybridization, we show that in vitro reduction of germ cell ploidy can be stimulated by FSH but not by T. FSH, but not T, also induced unexpectedly rapid (24–48 h) morphological changes resembling spermiogenesis, although individual changes (spermatid nucleus condensation and protrusion, cell body elongation, and flagellar growth) proceeded in an uncoordinated way and mostly resulted in the development of abnormal forms of elongated spermatids. Though ineffective alone, T potentiated the effects of FSH on meiosis and spermiogenesis. These effects of T were probably caused by the prevention of Sertoli cell apoptosis, an effect that could not be mimicked by FSH. These data show that, in the presence of high concentrations of FSH and T, human spermatogenesis can proceed in vitro with an unusual speed, but the resulting gametes are morphologically abnormal. The potential practical relevance of these findings to assisted reproduction remains to be assessed.

Human spermatogenesis in vitro: respective effects of follicle-stimulating hormone and testosterone on meiosis, spermiogenesis, and Sertoli cell apoptosis.

In spite of the availability of abundant data about in vitro spermatogenesis in laboratory animals, studies on human in vitro spermatogenesis are scarce. This study employed a relatively simple culture system, involving all cell types of seminiferous tubules, to analyze the effects of FSH and testosterone (T) on different characteristics of human germ and Sertoli cells in culture. By using fluorescence in-situ hybridization, we show that in vitro reduction of germ cell ploidy can be stimulated by FSH but not by T. FSH, but not T, also induced unexpectedly rapid (24-48 h) morphological changes resembling spermiogenesis, although individual changes (spermatid nucleus condensation and protrusion, cell body elongation, and flagellar growth) proceeded in an uncoordinated way and mostly resulted in the development of abnormal forms of elongated spermatids. Though ineffective alone, T potentiated the effects of FSH on meiosis and spermiogenesis. These effects of T were probably caused by the prevention of Sertoli cell apoptosis, an effect that could not be mimicked by FSH. These data show that, in the presence of high concentrations of FSH and T, human spermatogenesis can proceed in vitro with an unusual speed, but the resulting gametes are morphologically abnormal. The potential practical relevance of these findings to assisted reproduction remains to be assessed.

The effect of pulsatile gonadotropin-releasing hormone and estradiol administration on luteinizing hormone and follicle-stimulating hormone concentrations in pituitary stalk-sectioned ovariectomized pony mares

Hourly pulses of gonadotropin-releasing hormone (GnRH) or bi-daily injections of estradiol (E2) can increase luteinizing hormone (LH) secretion in ovariectomized, anestrous pony mares. However, the site (pituitary versus hypothalamus) of positive feedback of estradiol on gonadotropin secretion has not been described in mares. Thus, one of our objectives involved investigating the feedback of estradiol on the pituitary. The second objective consisted of determining if hourly pulses of GnRH could re-establish physiological LH and FSH concentrations after pituitary stalk-section (PSS), and the third objective was to describe the declining time trends of LH and FSH secretion after PSS. During summer months, ovariectomized pony mares were divided into three groups: Group 1 (control, n = 2), Group 2 (pulsatile GnRH (25 μg/hr), n = 3), and Group 3 (estradiol (5 mg/12 hr), n = 3). All mares were stalk-sectioned and treatment begun immediately after stalk-section. Blood samples were collected every 30 min for 8 h on the day before surgery (DO) and 5 d post surgery (D5) to facilitate the comparison of gonadotropin levels before and after pituitary stalk-section. Additionally, jugular blood samples were collected every 12 hr beginning the evening of surgery, allowing for evaluation of the gonadotropin secretory time trends over the 10 d of treatment. On Day 10, animals were euthanized to confirm pituitary stalk-section and to submit tissue for messenger RNA analysis (parallel study). Plasma samples were assayed for LH and FSH by RIA. Mean LH secretion decreased from Day 0 to Day 5 in Groups 1 and 3, whereas LH secretion tended (P < 0.08) to decrease in Group 2 mares. On Day 5, LH was higher (P < 0.01) in Group 2 (17.26 ± 3.68 ng/ml; LSMEANS ± SEM), than either Group 1 (2.65 ± 4.64 ng/ml) or group 3 (4.28 ± 3.68 ng/ml). Group 1 did not differ from Group 3 on Day 5 (P < 0.40). Similarly, mean FSH levels decreased in all groups after surgery, yet Group 2 mares had significantly (P < 0.001) higher FSH concentrations (17.66 ± 1.53 ng/ml) than Group 1 or Group 3 (8.34 ± 1.84 and 7.69 ± 1. 63 ng/ml, respectively). Regression analysis of bi-daily LH and FSH levels indicated that the time trends were not parallel. These findings indicate: 1) Pituitary stalk-section lowered LH and FSH to undetectable levels within 5 d after surgery, 2) pulsatile administration of GnRH (25 μg/hr) maintained LH and FSH secretion, although concentrations tended to be lower than on Day 0, and 3) E2 did not stimulate LH or FSH secretion.

High In Vitro Maturation Rate of Immature Mouse Eggs Can be Obtained by Using a Medium Containing Higher Concentrations of FSH and hCG

High In Vitro Maturation Rate of Immature Mouse Eggs Can be Obtained by Using a Medium Containing Higher Concentrations of FSH and hCG

P-239 High in vitro maturation rate of immature mouse eggs can be obtained by using a medium containing higher concentrations of FSH and hCG

P-239 High in vitro maturation rate of immature mouse eggs can be obtained by using a medium containing higher concentrations of FSH and hCG