Follicle-stimulating Hormone Treatment Research Articles

In vitro embryo production using ovaries removed from culled cows

The objective was to utilize in vitro maturation and fertilization to maximize embryo production from culled dairy cows. Ovaries were removed (via a colopotomy technique) from culled Holstein cows (n = 21) 2 d after standing estrus (T1), 2 d after treatment with 40 mg follicle stimulating hormone (FSH) given at standing estrus (T2), or at an unknown stage of the estrous cycle (T3).Treatment with FSH increased (P < 0.05) oocyte recovery rate (mean ± SEM, 14.6 ± 2.2, 25.4 ± 3.4, and 11.1 ± 2.6 oocytes ovary–1), for groups T1, T2 and T3, respectively, and the number of blastocysts obtained (1.6 ± 0.5, 4.4 ± 0.4 and 1.6 ± 0.6 blastocysts ovary–1). Culled dairy cows of high genetic merit are potential embryo donors, with an average of six to eight transferable embryos obtained from each FSH-treated cow. Key words: FSH treatment, oocyte recovery, IVM, IVF, culled cows

Effects of gonadotropins on bovine oocytes matured in TCM-199

The effects of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) during in vitro maturation of bovine oocytes in TCM-199 without serum were evaluated. Bovine oocytes with compact cumulus cells were collected from slaughterhouse-derived ovaries and cultured in Hepes-buffered TCM-199 supplemented with 5 mg/mL BSA, 1 μg/mL estradiol-17β, FSH (0, 0.015, 0.05, 0.15, 1.5 or 15 ng/mL; Experiment 1), LH (0, 0.14, 1, 7 or 49 μg/mL; Experiment 2) and combinations of 1 or 10 ng/mL FSH and 1 or 10 μg/mL LH (Experiment 3) at 39°C in 5% CO2 in air. After 22 h of maturation, cumulus expansion was estimated by scoring from 0 (no expansion) to 4 (full expansion of cumulus mass). In vitro fertilization was done with Percoll (45/90%) separated bull sperm at 1 × 106 sperm/mL in tert-TALP with 5 U/mL heparin. At 18 to 20 h post-insemination, presumptive zygotes were transferred to a chemically defined medium (CDM-1) supplemented with 0.5% BSA and nonessential amino acids for 72 h and then moved to CDM-2, additionally supplemented with essential amino acids. Zygotes were cultured at 39°C in 5% CO2, 5% O2 and 90% N2 for 8 days. During Experiments 1 and 2, cumulus expansion increased in proportion to concentrations of FSH and LH. Cleavage rates and development to blastocysts were not significantly different among FSH and LH treatments. In Experiment 3, cumulus expansion of bovine oocytes was maximal when 1 ng/mL FSH and 1 μg/mL LH were added to IVM medium, but cumulus expansion again was not related to developmental ability, although cleavage rates were improved slightly (P<0.05) by the combination of LH and FSH. Blastocyst quality, estimated by the size of inner cell mass, was not different between combinations of FSH and LH, and the numbers of nuclei were not different. Although expansion of cumulus cells surrounding bovine oocytes was altered in response to FSH and/or LH in semi-defined medium, cumulus expansion was not related to rates of cleavage or subsequent embryonic development in vitro. The effects of LH on cumulus expansion can be explained by as little as 1 part per 10,000 contamination with FSH.

The ERK-dependent signalling is stage-specifically modulated by FSH, during primary Sertoli cell maturation.

Primary cultures of Sertoli cells provide an interesting model to study how signalling pathways induced by a single hormone in a single cell type evolve, depending on the developmental stage. In vivo, follicle-stimulating hormone (FSH) induces proliferation of Sertoli cells in neonate and controls the subsequent differentiation of the entire population. Molecular mechanisms underlying Sertoli cell pleiotropic responses to FSH have long been investigated. But to date, only cAMP-dependent kinase (PKA) activation has been reported to account for most FSH biological activities in male. Here, we demonstrate that FSH activates the ERK MAP kinase pathway following dual coupling of the FSH-R both to Gs and to Gi heterotrimeric proteins, in a PKA- and also Src-dependent manner. This activation is required for FSH-induced proliferation of Sertoli cells isolated 5 days after birth. Consistently, we show that the ERK-mediated FSH mitogenic effect triggers upregulation of cyclin D1. In sharp contrast, at 19 days after birth, as cells proceed through their differentiation program, the ERK pathway is dramatically inhibited by FSH treatment. Taken together, these results show that FSH can exert opposite effects on the ERK signalling cascade during the maturation process of Sertoli cells. Thus, signalling modules triggered by the FSH-R evolve dynamically throughout development of FSH natural target cells.

Effects of follicle stimulating hormone (FSH) on follicular development, oocyte retrieval, and in vitro fertilization (IVF) in ewes during breeding season and seasonal anestrus

Administration of FSH increases the number of developing follicles, and affects oocyte health and cleavage rate. To determine the optimal level of FSH treatment, studies were conducted during the normal breeding season and seasonal anestrus. In Experiment 1, ewes were implanted with SyncroMate-B (SMB; norgestomet) for 14 days during the breeding season. Beginning on day 12 or 13 after SMB implantation, ewes were treated with saline (control; n=10), or treated with FSH for two days (2D; n=9) or three days (3D; n=10). In Experiment 2, conducted during seasonal anestrus, ewes were implanted with SMB for 14 days (n=23) or were not implanted (n=26). The SMB-implanted and nonimplanted ewes were assigned to one of three treatments as in Experiment 1: control (n=13), 2D (n=21) or 3D (n=15). In Experiments 1 and 2, ewes were laparotomized to count the number of follicles ≤ 3 mm and >3 mm and to retrieve oocytes. Healthy oocytes from each treatment were used for IVF. In Experiment 3, ewes (n=6) were implanted twice with SMB for 14 days during seasonal anestrus. Ewes were injected with FSH for 2 days, and the oocytes were collected and fertilized as in Experiments 1 and 2. In Experiment 1, FSH-treatment increased (P < 0.05) the number of follicles > 3 mm, the number of oocytes retrieved from follicles ≤ 3 mm and > 3 mm, the proportion of healthy oocytes, and the number of oocytes used for IVF. Oocytes from control and 2D ewes had greater (P < 0.01) cleavage rates than 3D ewes (68% and 71% vs. 42%). In Experiment 2, implanted and nonimplanted ewes had similar (P > 0.05) numbers of follicles, total oocytes, and healthy oocytes; therefore, data were combined. The FSH treatment increased (P < 0.01) the number of follicles >3 mm, and the number of oocytes recovered from follicles >3 mm. The recovery rate of oocytes and the percentage of healthy oocytes were similar for control and FSH-treated ewes. The cleavage rate in Experiment 2 ranged from 4 to 16%. In Experiment 3, the cleavage rate for ewes treated twice with SMB was 27% which tended to be greater (P < 0.07) than for the 2D ewes that received one SMB implant in Experiment 2. These data indicate that FSH increased the number of developing follicles and the number of healthy oocytes retrieved from ewes during the breeding season and seasonal anestrus. However, cleavage rates during seasonal anestrus were lower than during the normal breeding season in both FSH-treated and control ewes. Treatment of ewes for 2 days with FSH resulted in a greater cleavage rate than treatment of ewes for 3 days.

Follicle-Stimulating Hormone Regulates Steroidogenic Enzymes in Cultured Cells of the Chick Embryo Ovary

This investigation addresses the potential regulation of enzymes involved in the biosynthesis of steroid hormones during early stages of gonadal development by follicle-stimulating hormone (FSH). Gonadal cells of 10-day-old chick embryo and cells of the left ovary of 18-day-old chick embryo were cultured for 60 h in a defined medium with or without the addition of FSH (2.0 IU/ml). At the end of the culture, cells were recovered and evaluated by biotransformation of tritiated steroid precursors and mRNA levels were evaluated by RT-PCR. The production of estrone from androstenedione was increased in the FSH-treated cells, both human FSH (hFSH) and recombinant human FSH (rhFSH), indicating a stimulatory effect on aromatase (P450arom). Similarly, the intensity of the band corresponding to P450arom mRNA was higher in hFSH and rhFSH than in control and chorionic gonadotropin (hCG) groups. The P450arom stimulation was observed in the ovary of 10- and 18-day-old chick embryo. The transformation of dehydroepiandrosterone to androstenedione was taken as evidence of 3β-hydroxysteroid dehydrogenase function. This enzyme was stimulated in the cultured ovarian cells of 18-day-old chick embryos treated with hFSH and rhFSH compared with controls. The production of pregnenolone in the mitocondrial fraction of 18-day-old chick embryo ovary was increased when cultured with hFSH and rhFSH. This observation together with the increase in the band intensity corresponding to mRNA of P450 cholesterol side-chain cleavage indicates stimulation by FSH treatment; hCG produced a similar effect. Somatic cells of the medullary cords are proposed to be FSH target cells in the ovary of the chick embryo.

Treatment of chronic anovulation resistant to clomiphene citrate (CC) by using gonadotrophin-releasing hormone agonist (GnRH-a) ovarian suppression followed by recombinant follicle-stimulating hormone (r-hFSH) treatment (Suprecur, Puregon)

We performed a prospective study to assess the effects of a starting dose of 50 units of recombinant follicle stimulating hormone (Puregon) during gonadotrophin-releasing hormone agonist therapy (Suprecur) on follicular development in patients with clomiphene citrate-resistant polycystic ovary syndrome (PCOS). In a prospective clinical trial, 25 women with clomiphene citrate-resistant PCOS who wanted to become pregnant were treated by a low-dose step-up protocol of SC recombinant follicle stimulating hormone administration during gonadotrophin-releasing hormone agonist therapy, monitored by transvaginal ultrasonography and retrospectively by serum endocrine assays taken at each monitoring visit. Cancellation of cycles, ovulation, rate and size of follicles growth, serum E 2 concentration and pregnancy were recorded. All patients exhibited a response: 22 patients ovulated, of whom 10 conceived. Using the low-dose protocol during gonadotrophin-releasing hormone agonist therapy permitted induction of ovulation safely and successfully in patients with clomiphene citrate-resistant PCOS who were patients previously difficult to treat with the conventional ovulation induction protocol.

Pure follicle-stimulating hormone as an adjuvant therapy for selected cases in male infertility during in-vitro fertilization is beneficial

Objectives: Since research has demonstrated the possibility of hormonal therapy for male infertility, we conducted a study to analyze the efficacy of pure follicle-stimulating hormone (pFSH) treatment in patients with idiopathic, severe oligoteratoastheno-spermia (OTA) syndrome, or failed fertilization before referral to an intracytoplasmic sperm injection (ICSI) in an in-vitro fertilization (IVF) program. Study design: A retrospective, clinical study was carried out on 178 men with OTA syndrome. Group I comprised 76 patients selected for treatment with pFSH. Group II comprised 102 men who served as the controls. Pure FSH was administered intramuscularly to the patients in group I. Upon cessation of therapy, an IVF treatment cycle was carried out. Results: After treatment with FSH, sperm motility was the only parameter which significantly improved in Group I (34% vs. 23%, and 15% vs. 24% in the subgroups of Group 1, respectively; P<0.05). In-vitro fertilization pregnancy rates were similar in both groups. The highest spontaneous pregnancy rates were achieved in FSH-treated, severe OTA patients. Conclusions: Patients with severe male factor infertility may benefit from pFSH in terms of sperm motility, fertilization by IVF and spontaneous pregnancy rates. Selection criteria for FSH treatment are proposed.

Alteration of follicle-stimulating hormone and testosterone regulation of messenger ribonucleic acid for Sertoli cell proteins in the rat during the acute phase of spinal cord injury.

The detrimental effects of spinal cord injury (SCI) on spermatogenesis in the rat can be attenuated by exogenous testosterone (T) but enhanced by exogenous follicle-stimulating hormone (FSH). These results suggest that T-dependent cellular events may be involved in testicular injury after SCI and that such events may be associated with modification of FSH effects on Sertoli cell function. The current study compared the responses of Sertoli cells to exogenous T and FSH after SCI or sham surgery using steady-state levels of Sertoli cell protein mRNA transcripts as markers of responsiveness. Rats underwent sham surgery or SCI and then were treated for 7 or 14 days with T-filled silastic capsules (2 x 5 cm) and/or daily injections of 0.1 units of porcine FSH. Vehicle-treated control rats received 5-cm empty capsules and daily injections of saline vehicle. Two weeks after sham surgery, levels of mRNA for the androgen receptor (AR), FSH receptor (FSHR), androgen-binding protein (ABP), or sulfated glycoprotein (SGP)-2 in the testis were unaffected by T or FSH alone. Testosterone alone, however, significantly decreased transferrin (Trf) mRNA levels in the testis (P: < 0.01). The combination of T and FSH treatments resulted in significant decreases in levels of the above transcripts (P: < 0.05; P: < 0.01). Seven days after SCI, the testes of vehicle-treated SCI rats had higher levels of AR and SGP-2 mRNA than did those of sham control rats (P: < 0.01); such effects were transient and disappeared by Day 14 post-SCI. Testosterone treatment of SCI rats for 7 days resulted in decreases in mRNA levels for AR and Trf in the testes (P: < 0.01) but increased testicular levels of mRNAs for FSHR and SGP-2 in SCI rats. Follicle-stimulating hormone treatment for 7 days prevented the increase in AR mRNA that was seen in the testis of untreated SCI rats and increased levels of ABP and SGP-2 mRNAs in SCI rats (P: < 0.01). Follicle-stimulating hormone treatment of SCI rats did not affect FSHR mRNA levels by itself, but it blocked the stimulatory effect of T on FSHR and SGP-2 mRNAs. Fourteen days after SCI, testicular AR mRNA levels were not affected by T alone, but they increased in those rats that received FSH with or without concurrent T treatments (P: < 0.05). In contrast to their effects in sham control rats, T or FSH alone or in combination resulted in significant increases in testicular levels of ABP, SGP-2, and FSHR mRNAs (P: < 0.05). At this time, Trf mRNA in the testis of SCI rats was also suppressed by T (P: < 0.05), as it did in sham control rats, but Trf mRNA was increased by the FSH (P: < 0.01) that had inhibited this transcript in the testes of sham control rats. The effects of FSH on the Sertoli cell transcripts in SCI rats were either attenuated or blocked when T was given concurrently. In addition, testicular and serum T levels in those SCI rats that received FSH (alone or in combination with T) for 14 days were significantly increased, an effect that was not seen after sham surgery. These findings demonstrate that hormonal regulation of both Sertoli and Leydig cells was altered during the acute phase of SCI. Such changes may modify the functions of both cell types, thereby affecting the endocrine and/or paracrine microenvironment within the seminiferous epithelium. These effects could impair the functional capacity of Sertoli cells and contribute to impairment of spermatogenesis after SCI.

Tumor stroma as the main source of inhibin production in ovarian epithelial tumors.

Elevated serum inhibin levels have been found in ovarian cancer patients; however, the source of the elevated inhibin is uncertain. Previous studies of activin in human ovarian cancer suggest that activin may promote the growth of the tumor. The aims of this study were to examine the source of elevated inhibin from ovarian epithelial tumors (OETs) and to preliminarily investigate the role of the gonadotropin-inhibin/activin relationship in the development of OET. The protein and mRNA expression of alpha and betaA subunits of inhibin/activin were examined by immunohistochemistry (IHC) and reverse transcription polymerase chain reaction (RT-PCR) in 120 OETs, including 30 benign cystadenomas, 30 borderline tumors, and 60 carcinomas. Stromal and epithelial cells were microdissected from 23 OETs to further examine the expression of alpha and betaA subunits by RT-PCR. Dimeric inhibin A and activin A production were measured by using the two-site ELISA from three OET cell lines in culture under treatment of follicle-stimulating hormone (FSH) and luteinizing hormone (LH). betaA subunit was expressed in the epithelial component of 100% of the cystadenomas, in 80% of borderline tumors, and in 75% of the carcinomas, but not in tumor stroma. Inhibin alpha expression was not found in the epithelium of all OETs studied, but focal inhibin alpha immunoreactivity was seen in the tumor stroma (mainly luteinized stromal cells) in the majority of cases. Dimeric activin A was produced by all of the three OET cell lines with a 1.5-1.9-fold increment after FSH stimulation. However, activin A production was not augmented by LH treatment. No inhibin A was produced by the three OET cell lines with or without gonadotropin stimulation. The stroma of OET is the major source in the production of inhibin alpha (monomer). Dimeric inhibin A production may be the result of combined efforts of the tumor stroma (alpha subunit) and epithelium (betaA subunit). Cellular, compartmental expression of inhibin and activin subunits may play a role in the development of OET, although the mechanism remains undefined. The unopposed activin A production stimulated by FSH in OET cell lines suggests that activin production may represent one of the cellular mechanisms of growth promotion by FSH.

Differential regulation by FSH and IGF-I of extracellular matrix IGFBP-5 in bovine granulosa cells: effect of association with the oocyte

Inhibition of insulin-like growth factor (IGF)-I induced DNA synthesis in bovine oocyte–cumulus complexes (OCCs) caused by follicle-stimulating hormone (FSH) has been linked to changes in the extracellular matrix which do not occur in mural granulosa cells (MGCs). We investigated regulation by IGF-I and FSH of secreted and extracellular matrix entrapped IGF-binding proteins. OCCs and MGCs from bovine ovaries were cultured in media supplemented with IGF-I and FSH for 24 h. Culture media and extracellular matrix were analysed for IGF-binding proteins by Western ligand blot and immunoblot and found to contain principally IGFBP-3 and -5. The combined treatment of IGF-I and FSH increased the concentration of IGFBP-3 in OCC and MGC conditioned media by 4- and 6-fold, respectively. Treatment of OCCs and not MGCs with IGF-I and FSH together increased extracellular matrix IGFBP-5 by 2.5-fold. The differential regulation of extracellular matrix IGFBP-5 in OCCs compared to MGCs suggest involvement of changes in the extracellular matrix brought about by IGF-I and FSH in overall regulation of IGF-I in the ovarian follicle.

FSH in the treatment of oligozoospermia

The aim of this study was to individuate parameters able to distinguish oligozoospermic subjects who will respond to follicle-stimulating hormone (FSH) therapy. A group of 135 oligozoospermic subjects was divided in three groups considering basal FSH and inhibin B concentrations: group A (normal FSH and inhibin B) characterized by moderate hypospermatogenesis sometimes associated to partial spermatidic arrest; group B (high FSH and normal inhibin B) characterized by hypospermatogenesis associated or not to spermatogonial/spermatocytic arrest; group C (high FSH and low inhibin B) characterized by severe hypospermatogenesis. Seventy-eight patients were treated with FSH at the dose of 75 IU on alternate days while 57 were treated with the same dose every day for 3 months. After FSH treatment a significant increase in ejaculated sperm concentration was observed only in oligozoospermic subjects with normal basal FSH and inhibin B plasma levels (group A) showing a testicular cytological picture of moderate hypospermatogenesis. In these subjects no differences in sperm production were observed between the two protocols of therapy. In the remaining patients of group A, characterized by hypospermatogenesis associated with maturation arrest at spermatidic level and in group B and C, no increase in sperm concentration was observed after therapy. These data suggest that FSH treatment may have a role in oligozoospermic subjects only when the spermatogenetic alterations consist in germ cell depopulation without maturative disturbances and with normal FSH concentrations.

Secretion of IGF-1 by ovine granulosa cells: effects of growth hormone and follicle stimulating hormone.

Insulin-like growth factor-1 (IGF-1) is implicated in follicle development and is considered to mediate the actions of growth hormone (GH) and gonadotrophins at the ovarian level. However, the expression and secretion of IGF-1 by the ovary are controversial, partly because of species and cell-type specificity. The present study investigated whether IGF-1 is produced by ovine granulosa cells and whether its production is regulated by GH and follicle stimulating hormone (FSH). Follicles (>/=4.0 mm) were obtained from ewes during seasonal anoestrus. Granulosa cells were cultured for a total period of 96 h in Dulbecco's modified Eagle's medium (DMEM)/Ham's F-12 medium supplemented with BSA (0.1%, w:v), transferrin (0.5 microg/ml) and testosterone (100 ng/ml). In the first set of experiments, cells were incubated in the presence of bovine calf serum (BCS) (2.5%) for the initial 48 h of culture. The cells were then cultured for the next 48 h in medium without BCS, but containing either GH (0, 2, 20, and 200 ng/ml) or FSH (0, 20, 200, and 2000 ng/ml). The medium was assayed for oestradiol (E), progesterone (P) and IGF-1. There were six wells per treatment and the experiment was carried out four times. Control granulosa cells maintained both IGF-1 and E secretion, with only low levels of progesterone output. In all experiments, both GH and FSH produced significant (P<0.001) dose-related increases in E, IGF-1 and P secretion into the medium. The maximum responses to GH (20 or 200 ng/ml) were 402% for E and 528% for IGF-1 compared with controls. The maximum responses to FSH (200 or 2000 ng/ml) were 460% for E and 514% for IGF-1. The objective of the second set of experiments was to determine the effect of the progestogenic status of cells on IGF-1 production. Granulosa cells were cultured both in the presence and absence of BCS (2.5% in the medium) during the initial 48 h of culture. For the next 48 h, cells were cultured in serum-free medium. Addition of BCS to the medium during the initial 48 h of culture stimulated progesterone production. However, it did not affect either IGF-1 or oestradiol secretion between 49 and 96 h of culture, or the cell numbers at the end of culture. In conclusion, (1) IGF-1 is secreted by granulosa cells irrespective of their progestogenic status and (2) concomitant increases in E and IGF-1 production by granulosa cells as a result of GH and/or FSH treatment suggest a role for GH and FSH in the regulation of ovarian function.

Follicle-stimulating hormone treatment for men with idiopathic oligoteratoasthenozoospermia before in vitro fertilization: the impact on sperm microstructure and fertilization potential

Objective: To assess the effect of FSH on sperm fertilization potential and sperm intracellular structure in men with oligoteratoasthenozoospermia and a proven low fertilization rate in IVF. Design: Prospective, randomized, partial crossover study. Setting: IVF Unit, Golda Campus, Rabin Medical Center, Petah Tikva, Israel. Patient(s): Forty normogonadotropic, normogonadal men with oligoteratoasthenozoospermia and at least one previous IVF attempt in which fertilization failed or the fertilization rate was <30%. Intervention(s): The men were randomly assigned to treatment with daily injections of 75 IU of FSH or 150 IU of FSH for at least 60 days before IVF treatment. A control group of men underwent an IVF cycle without treatment and then were randomly assigned to join group 1A or 1B for an additional IVF cycle with treatment. Main Outcome Measure(s): LH, FSH, and testosterone levels during FSH treatment, evaluation of ultramorphologic changes in sperm by electron microscopy, and comparison of fertilization rates in the control and study groups. Result(s): After treatment with 75 IU or 150 IU of FSH, the mean fertilization rates were 19.7% and 20.5%, respectively, compared with a 5.8% fertilization rate in the study control cycles. Conclusion(s): Prolonged treatment with FSH results in a significant increase in fertilization rates. This effect may be related to improvements in subcellular components of the sperm.

Follicle-stimulating hormone and testosterone stimulation of immature and mature Sertoli cells in vitro: inhibin and N-cadherin levels and round spermatid binding.

The in vitro response of Sertoli cells isolated from adult rat testes to testosterone (T) and follicle-stimulating hormone (FSH) treatment was investigated. Sertoli cells from >70-day-old Sprague-Dawley rats were isolated by a combined enzymatic treatment followed by the removal of the majority of contaminating germ cells with immobilized peanut agglutinin lectin. Sertoli cells were then cultured for 6-10 days, forming a confluent layer with a cell viability of >83% and 74-77% purity. The contaminating cells were peritubular cells (4-6%), pachytene spermatocytes (4-5%), round spermatids (<2%), elongated spermatids (<1%), and degenerating germ cells (14.8%). The proportion of degenerating germ cells decreased from 14.8% to 8.6% between days 6 and 10 in culture. After a prestimulation culture period of 4 days, FSH treatment over a 2-day period resulted in a dose-related increase of inhibin with a median effective dose (ED50) value of 36.7+/-20.4 ng/ml in comparison with an ED50 value of 4.4+/-0.9 ng/ml obtained with immature Sertoli cell cultures from 20-day-old rats. Mature Sertoli cells, in contrast to immature Sertoli cells, were unresponsive to combined FSH + T treatment for the production of the cell adhesion protein N-cadherin. FSH treatment promoted the in vitro binding of round spermatids isolated from adult testis to adult Sertoli cells in coculture. It is concluded that mature Sertoli cells in culture are responsive to FSH in terms of inhibin production and round-spermatid binding. The lack of an FSH + T-induced increase in N-cadherin or round spermatid binding is attributed to either a reduced sensitivity, or an alteration in the regulation of mature Sertoli cells by FSH + T.

Inhibin B plasma concentrations in oligozoospermic subjects before and after therapy with follicle stimulating hormone.

The aim of this study was to investigate inhibin B and follicle stimulating hormone (FSH) secretion in a large group of oligozoospermic subjects affected by different degrees of testicular damage, before and after FSH treatment. A total of 135 oligozoospermic subjects (sperm count < 20 x 10(6)/ml) were evaluated for seminal parameters and FSH, luteinizing hormone (LH), testosterone and inhibin B plasma concentrations. Testicular structure was analysed with bilateral fine needle aspiration cytology. Inhibin B showed an inverse correlation with FSH, no correlation with sperm concentration and a significant relationship with intratesticular spermatid number, demonstrating that testicular spermatids play an important role in the control of inhibin B production. Twenty-five subjects with sperm counts < 10 x 10(6)/ml were treated with FSH; 11 of these had basal FSH and inhibin B plasma concentrations in the normal range (group A), while in seven subjects FSH was elevated (> 7 IU/l) with normal inhibin B (group B), and in seven patients FSH was high and inhibin B reduced (< 80 pg/ml) (group C). During treatment, in group A patients inhibin B plasma concentrations increased significantly after 2, 3 and 4 weeks of FSH administration and declined thereafter to pre-treatment concentrations. Groups B and C did not show any modification during the treatment. In the same period, in group A FSH increased significantly after 2, 3 and 4 weeks and subsequently declined. In groups B and C, FSH increased significantly after 2 weeks and remained elevated during the following period. The results of the present study confirm the significant inverse correlation between inhibin B and FSH plasma concentrations in subjects with disturbed spermatogenesis, and demonstrate that inhibin B reflects Sertoli cell function and their interaction with spermatids. FSH and inhibin B concentrations are an expression of the spermatogenic status of seminiferous tubules. FSH treatment seems to modify inhibin B plasma concentrations only in subjects with normal basal FSH and inhibin B, independently from the effects of this therapy on sperm production.

The effect of follicle-stimulating hormone (FSH) on the expression of FSH receptor in cultured rat granulosa cells

The acquisition of FSH receptors during folliculogenesis is believed to be a key event in the subsequent development of the follicle. The regulation by FSH of FSH receptor expression and function were further studied using cultured granulosa cells of diethylstilbestrol (DES)-primed immature rats. Incubation of rat granulosa cells with FSH led to a reduction in FSH receptor levels for a short time (6 h), followed by an increase in FSH receptor levels that reached maximum of around 150% of the initial level within 3 days after the addition of FSH. FSH stimulation caused a reduced cAMP response to subsequent FSH treatment and a time course experiment demonstrated that this response was detectable within 30 min of exposure to FSH and reached a plateau after 4 h to 24 h. The recovery of FSH responsiveness in cAMP production of granulosa cells was seen after 48 h of FSH-free interval. Treatment with forskolin (FSK) enhanced the effect of subsequent FSH on the production of intracellular cAMP. Treatment with PMA did not affect the response to subsequent FSH treatment. These data showed that the FSH is essential for the suppression of the FSH receptor function in the adenylyl cyclase pathway. Desensitization of cellular response to continuous agonist stimulation may occur because of changes in the numbers of FSH receptor, as well as changes in the functional properties of the effector system.

Quantitative ultramorphological (QUM) analysis of human sperm: diagnosis and management of male infertility.

The advantages of quantitative ultramorphological (QUM) sperm analysis in the diagnosis and treatment of male infertility are presented. QUM methodology is based on three elements: (1) complementary SEM and TEM observations of 7 sperm cell subcellular organelles: acrosome, postacrosomal lamina, nucleus, neck, axoneme, mitochondrial sheath, and outer dense fibers; (2) systematic classification of the specific ultramorphological malformations into 4 pathological and the normal categories, which indicate the morphological state of each subcellular organelle; and (3) comparison between well-defined reference groups with opposite fertility status or treatment conditions. QUM analysis has enabled the establishment of two indices that optimally express the in vivo and in vitro male fertility potential: The Natural Fertility Index (NFI), which allowed an accurate prediction (97% sensitivity and 90% specificity) of 80% of the naturally fertile and suspected infertile male patients, and the in vitro fertilization (IVF) score, which enabled prediction of 76% of the nonfertilizing and 90% of the fertilizing IVF groups. Validation tests confirmed these data. QUM also enabled assessment of ultramorphological indications for varicocele and radiation exposure: Both male factor etiologies indicated a persistent effect on the natural fertility potential, as expressed by structural changes in the nucleus. Varicocele was found to cause defects in the sperm head organelles related to early spermatid development, whereas ionizing radiation resulted in amorphous head shape. Criteria for specific non-in vitro therapeutic interventions such as varicocelectomy, follicle-stimulating hormone (FSH) administration, and acupuncture treatment were established. A varicocele index, which enabled the correct classification of 79 and 89% of the patients pre- and post-high ligation, respectively, was suggested to be a good indicator for varicocele which affects the fertility potential. Males exhibiting idiopathic impairment of sperm acrosome and nucleus were found to be potential responders to FSH treatment, whereas patients exhibiting low sperm activity proved to be good candidates for acupuncture treatment. Indications for selecting the optimal appropriate assisted reproduction technique (ART) procedure were found: Patients with a low Natural Fertility Index should be recommended for ART. A first choice ART selection should be performed according to an ART index based on the ultramorphological examination of the tail axoneme. The above index enabled correct prediction of 78% of the patients who achieved pregnancy following conventional ART (intrauterine insemination or IVF) and 74% of those whose wives conceived only following intracytoplasmic sperm injection. QUM sperm analysis is clinically informative, nontraumatic, and in the long run also cost-effective. This analysis should be performed when the male infertility factor cannot be clearly diagnosed by routine tests and prior to the first ART trial.

Quantitative ultramorphological analysis of human sperm: fifteen years of experience in the diagnosis and management of male factor infertility.

The advantages of quantitative ultramorphological (QUM) sperm analysis in the diagnosis and treatment of male infertility are presented. The QUM methodology is based on three elements: complementary scanning electron microscopy and transmission electron microscopy observations of 7 sperm cell subcellular organelles (acrosome, postacrosomal lamina, nucleus, neck, axoneme, mitochondrial sheath, and outer dense fibers); systematic classification of the specific ultramorphological malformations into 4 pathological and the normal categories, indicating the morphological state of each subcellular organelle; and comparison between well-defined reference groups with opposite fertility status or treatment conditions. QUM has established 2 indices for the in vivo and in vitro male fertility potential: (1) Natural Fertility Index (NFI), with accurate prediction (97% sensitivity and 90% specificity) of 80% of the male patients; and (2) IVF score, with prediction of 76% of the nonfertilizing and 90% of fertilizing IVF groups. QUM has enabled assessment of ultramorphological indications for varicocele and radiation exposure. Varicocele causes defects in sperm head organelles related to early spermatid development, whereas ionizing radiation causes amorphous head shape. QUM established criteria for specific non-in-vitro therapeutic interventions, including varicocelectomy, follicle-stimulating hormone (FSH) administration, and acupuncture. The varicocele index enabled correct classification of 79 and 89% of patients with and without varicocele. Males with idiopathic impairment of sperm acrosome and nucleus are potential responders to FSH treatment, whereas patients exhibiting low sperm activity are candidates for acupuncture treatment. Patients with a low Natural Fertility Index are recommended for an assisted reproduction technique (ART). based on the ultramorphology of the tail axoneme. Patients who achieved pregnancy following intrauterine insemination or in vitro fertilization and those whose wives conceived only following intracytoplasmic sperm injection were classified with accuracy of 78 and 74%, respectively. QUM sperm analysis is clinically informative, nontraumatic, and cost-effective, and is recommended when the male infertility factor cannot be clearly diagnosed by routine tests prior to first ART trial.

Pretreatment with follicle stimulating hormone promotes the numbers of human oocytes reaching metaphase II by in-vitro maturation.

The objective of this study was to investigate the effect of follicle stimulating hormone (FSH) priming on the in-vitro maturation (IVM) of human oocytes from healthy ovaries using a chemically defined culture system. Seventeen patients donating oocytes for research received a truncated course of 600 IU FSH over 5 days and a further control group of nine patients received no FSH treatment. Mid-follicular phase cumulus-enclosed oocytes (n = 160) were aspirated from follicles < or =4 mm diameter under transvaginal ultrasound guidance and were cultured for 48 h in microdrops of medium containing 10 mIU/ml FSH and 100 mIU/ ml human chorionic gonadotrophin (HCG). The results demonstrated that human oocytes will efficiently undergo IVM under serum-free conditions. After mild FSH stimulation, a greater number of cumulus-enclosed oocytes was collected, and following culture, a lower rate of degeneration was observed. Significantly more oocytes completed nuclear maturation to metaphase II following FSH stimulation (71.1 versus 43.5%). In conclusion, a truncated course of FSH stimulation in vivo improved the oocyte maturation rate in vitro, giving a mean of 4.8+/-0.7 metaphase II oocytes per patient compared with only 2.1+/-0.7 from control patients, thus yielding more mature oocytes for future IVF treatment.

Histomorphometric and progesterone receptor immunohistochemical analysis in the oviduct of newly hatched chicks treated with follicle-stimulating hormone during embryonic development.

In this study we evaluated the histomorphology and ultrastructure of the oviduct of newly hatched chicks, as well as the immunohistochemical expression of progesterone receptor (PR) in this tissue after follicle-stimulating hormone (FSH) treatment on days 13, 15 and 17 of embryonic development. Results indicated a marked difference in the histology of the oviduct of newly hatched chicks treated with FSH. Magnum mucosa from these animals presented a pseudostratified epithelium with evaginations from the lumen into the epithelium and from the latter into the stroma beneath where tubular glands are formed. In contrast magnum mucosa from control animals presented columnar epithelium with no evaginations. In magnum epithelium FSH also induced the formation of cilia and microvilli projections into the lumen as well as an increase in the wall and lumen areas and in the density of nuclei per unitarea. PR immunoreactivity was only observed in the oviduct of FSH treated animals. PR was located in the nucleus of epithelial luminal cells, mucosal stromal cells and smooth muscle cells. These findings suggest that FSH induces an adequate hormonal milieu for the cytodifferentiation and PR gene expression in the chick oviduct.