A correlative study of the restorative effects of endogenous and exogenous hormones on the Leydig cells, the testis and the epididymis of the regressed hedgehog Effects of hormones on hedgehog Leydig cells

The study of the effects of morphogenesis at puberty on the Leydig cells in the testis of the young hedgehog and of the subsequent changes due to the seasonal varisations, has been done. Furthermore, the restorative changes induced by the exogenous hormones in the Leydig cells and the related sex organs of the regressed hedgehogs have also been studied. It was observed that the Leydig cells from the undifferentiated mesenchyme cell-like nature in the young hedgehog, develop into an adult form possessing large number of lipids, a well-developed Golgi apparatus, complex mitochondria and extensive smooth endoplasmic reticulum. The depletion of the lipids and other regression associated changes are found in the interstitial Leydig cells but not in those situated under tunica albuginea and the latter probably function as lipid storing cells during regression. Pituitary extract, either alone or in combination, but not testosterone, could restore completely the structure of the regressed Leydig cells. Similarly, the restoration of the complete process of spermatogenesis and the structure and function of the epididymis in the regressed hedgehog was found to be dependent upon the synergistic action of both testosterone and the gonadotrophic hormones.     

Endocrinology: Gonadotrophins and prolactin rise after bilateral oophorectomy for benign conditions

The aim of this study was to analyse the changes in folliclestimulating hormone (FSH), luteinizing hormone (LH) and prolactinconcentrations in the 3 months following oophorectomy in pre-menopausalwomen operated on for benign gynaecological conditions. Includedin this analysis were 21 women (mean age 47 years, range 46–52)who underwent bilateral oophorectomy plus hysterectomy for fibroidsor ovarian cysts. Plasma concentrations of FSH, LH and prolactinwere measured before and on days 2, 4, 6, 14 and 30 after surgery;in 10 cases measurements were made on day 60, and in five caseson day 90 after surgery. Hormone concentrations were measuredin duplicate daily samples, and immunoenzymatic assay kits wereused for all the immunoassays. The FSH and LH concentrationsincreased constantly after surgery. Mean prolactin concentrationsalso increased from 12.1 ng/ml before surgery to 31.5 ng/mlon day 14 after bilateral oophorectomy, but decreased thereafterto 18.2 ng/ml on day 30, 10.9 ng/ml on day 60 and 6 ng/ml onday 90. In conclusion, transient (2–3 weeks) increasedprolactin concentrations are observed after surgical castration.     

Synthesis of gonadotrophins and its regulation in the pituitary gland

Regulation of the synthesis of pituitary gonadotrophins LH andFSH has been studied in the rat using either cell-free translationof pituitary mRNAs, or hybridization techniques with syntheticoligodeoxynucleotides or cloned complementary DNAs. Gonadectomygreatly increases and supplementing gonadectomized rats withgonadal steroids diminishes the rate of synthesis of the gonadotrophinsubunits. Hybridization experiments suggest that gonadal steroidsregulate the expression of the genes coding for pituitary gonadotrophinsubunit precursors. Using the incorporation of labelled methionineby pituitary cells in culture, followed by specific immunoprecipitationof LH-related subunits and SDS-poly-acrylamide gel analysisof immunoprecipitated peptides, there was evidence that gonadotrophinreleasing hormone (GnRH) significantly enhances the radioactivityincorporated into both - and LH-subunits. This effect is specific,it is not a secondary effect due to the release of LH. A cyclicAMP (cAMP) analogue, 8-Br-cAMP, as well as forskolin and choleragen,which are cAMP generators and a diacylglycerol analogue, tetradecanoylphorbolacetate (TPA), mimic the stimulatory action of GnRH on the synthesisof the polypeptide chains of LH. However, no evidence has beenobtained that either cAMP or diacylglycerols mediate this GnRHeffect. These results suggest that the synthesis of pituitarygonadotrophins is under a double control of gonadal steroidsand GnRH which exert opposite effects, inhibitory for steroidsand stimulatory for GnRH. The negative control by steroids occursat the genomic level, while the positive effect of GnRH proceedsvia different mechanisms which remain to be elucidated.     

The regulatory role of FSH in steroid formation by luteinized human granulosa cells in culture

Granulosa cells were aspirated from human pre-ovulatory folliclesfollowing a combined clomiphene-gonadotrophin stimulation inan in-vitro fertilization (IVF) programme. The cells were culturedfor 8 days in medium M199 containing 10% bovine fetal calf serumunder 5% CO₂ in air. Pure human FSH and human LH were addedalone or in combination to the culture in various concentrationsand the progesterone (P) and oestradiol-17 (E₂) levels in themedium were measured every second day by a conventional RIAtechnique. In the presence of FSH or LH the formation of P increased2- to 3-fold with the pronounced effect after 4 to 6 days inculture. Addition of testosterone (T) (3 ? 10^–7 M) tothe culture medium affected neither basal nor gonadotrophinstimulated P formation. In this system, only minute amountsof E₂ were formed and neither FSH nor LH stimulated its formation.When the medium was fortified with T, basal E₂ formation increased50- to 100-fold. FSH further stimulated this conversion significantlyafter 6 and 8 days of culture, while LH had no significant influence.     

Ovarian stimulation with HMG: results of a prospective randomized phase III European study comparing the luteinizing hormone-releasing hormone (LHRH)-antagonist cetrorelix and the LHRH-agonist buserelin. European Cetrorelix Study Group

In this prospective and randomized study, 188 patients received the luteinizing hormone-releasing hormone (LHRH) antagonist cetrorelix, and 85 patients the LHRH agonist buserelin to prevent endogenous luteinizing hormone (LH) surges during ovarian stimulation in in-vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) cycles. Ultimately, 181 patients (96.3%) in the cetrorelix group, and 77 (90.6%) in the buserelin group, reached the day of the human chorionic gonadotrophin (HCG) injection. The mean number of human menopausal gonadotrophin (HMG) ampoules administered and the mean number of stimulation days with HMG were significantly less in the cetrorelix group than in the buserelin group (P < 0.01). A rise in LH and progesterone concentrations was observed in three of the 188 patients (1.6%) who received cetrorelix. On the day of the HCG administration, more follicles of a small diameter (11-14 mm) were observed in the buserelin group than in the cetrorelix group (P = 0. 02) and the mean serum oestradiol concentration was significantly higher in patients who received buserelin than in those who received cetrorelix (P < 0.01). Similar results were observed in fertilization, cleavage and pregnancy rates in the two groups. In conclusion, the use of the LHRH antagonists might be considered more advantageous because of the short-term application needed to inhibit gonadotrophin secretion, so allowing a reduction in the treatment time in a clinically significant manner.     

The LHRH antagonist cetrorelix: a review

In those clinical situations in which an immediate and profound suppression of gonadotrophins is desired, LHRH agonists have the disadvantage of producing an initial stimulatory effect on hormone secretion. Therefore, the use of GnRH antagonists which cause an immediate and dose-related inhibition of LH and FSH by competitive blockade of the receptors is much more advantageous. One of the most advanced antagonist produced to date is Cetrorelix, a decapeptide which has been shown to be safe and effective in inhibiting LH and sex-steroid secretion in a variety of animal species and in clinical studies as well. Clinical trials in patients suffering from advanced carcinoma of the prostate, benign prostate hyperplasia, and ovarian cancer are currently in progress and have already shown the usefulness of this new treatment modality. In particular, the concept that a complete suppression of sex-steroids may not be necessary in indications such as uterine fibroma, endometriosis and benign prostatic hyperplasia represents a promising novel perspective for treatment of these diseases. Following completion of phase III trials in controlled ovarian stimulation for IVF regimens, Cetrorelix was given marketing approval and, thus, became the first LHRH antagonist available clinically.     

Embryo quality in natural versus stimulated IVF cycles

BACKGROUND: The impact of controlled ovarian stimulation (COS) on oocyte and subsequent embryo quality remains controversial. In the present study we have compared embryo quality in natural and stimulated cycles in the same group of patients. METHODS: This retrospective study was comprised of patients with a regular menstrual cycle who had IVF after COS using rFSH in a long GnRH agonist protocol. In all stimulated cycles the patients had fresh embryos transferred and surplus good quality embryos cryopreserved. Subsequently the same patients were treated with a modified FER cycle (mFER) where thawing of the frozen embryos was combined with aspiration of the dominant follicle in the natural cycle. The embryo cleavage stage and quality score were compared between the stimulated and the natural cycle for the patients having an embryo in the natural cycle. RESULTS: In 177 cases patients returned for mFER in a natural cycle. Spontaneous ovulation had occurred in 35 cycles. In 17 cycles no oocyte was retrieved at aspiration and in 125 cycles 128 oocytes were aspirated. In the stimulated cycles from these patients we had obtained 950 embryos (cleavage rate 70.4%) versus 85 embryos (cleavage rate 66.4%) (P = 0.34) in the natural cycles. Comparing the embryos in the natural and stimulated cycles in all patients having an embryo in the natural cycle, we found no difference in the distribution between the different cleavage stages. Of the cleaved embryos, 53% in the stimulated cycles had >or=4 cells versus 59% in the natural cycles after 2 days culture (P = 0.31). In the stimulated cycles 61% of the embryos had <10% fragmentation at the time of transfer on day 2, compared to 69% in the natural cycles (P = 0.15). CONCLUSION: The administration of exogenous gonadotrophins was not reflected in cleavage capacity or quality assessment of the resulting embryos.     

Controlled ovulation of the dominant follicle: a critical role for LH in the late follicular phase of the menstrual cycle

BACKGROUND: A method was sought to control ovulation of the dominant follicle and to test the importance of LH during the late follicular phase of the menstrual cycle. Menstrual cycles of rhesus monkeys were monitored, and treatment initiated at the late follicular phase (after dominant follicle selection, before ovulation). METHODS: The 2-day treatment consisted of GnRH antagonist plus either r-hFSH and r-hLH (1:1 or 2:1 dose ratio) or r-hFSH alone. In addition, half of the females received an ovulatory bolus of hCG. RESULTS: When treatment was initiated at estradiol levels >120 pg/ml, neither the endogenous LH surge, ovulation nor luteal function were controlled. However, when treatment was initiated at estradiol levels 80-120 pg/ml using either 1:1 or 2:1 dose ratios of FSH:LH, the LH surge was prevented, and ovulation occurred following hCG treatment. FSH-only treatment also prevented the LH surge, but follicle development appeared abnormal, and hCG failed to stimulate ovulation. CONCLUSIONS: Control over the naturally dominant follicle is possible during the late follicular phase using an abbreviated GnRH antagonist, FSH+LH protocol. This method offers a model to investigate periovulatory events and their regulation by gonadotrophins/local factors during the natural menstrual cycle in primates.     

Developmental potential of embryos produced by in-vitro fertilization from gonadotrophin-releasing hormone antagonist-treated macaques stimulated with recombinant human follicle stimulating hormone alone or in combination with luteinizing hormone

We previously demonstrated, in luteinizing hormone (LH)-deficientmacaques, that follicular growth and maturation occurred withadministration of exogenous (recombinant human) follicle stimulatinghormone (r-hFSH) alone, and that the oocytes recovered fertilizedat a notably higher rate than their counterparts from animalsreceiving both r-hFSH and r-hLH (Zelinski-Wooten et al., 1995).Here, the developmental potential of embryos produced from animalstreated with r-hFSH alone or in combination with r-hLH was evaluated.Embryos (n = 127) were cryopreserved, thawed and either co-culturedon buffalo rat liver cells until the hatched blastocyst stageor transferred to synchronized recipients. Although embryosfrom each treatment group demonstrated a similar ability todevelop to hatched blastocysts with a definitive inner cellmass, a significant difference was seen in cryosurvival (56versus 78%) and in developmental rate to the hatched blastocyst(12 versus 10 days) between embryos from the r-hFSH alone andthe combination group respectively. Pregnancies resulted followingoviductal embryo transfers in both groups, with corpus luteumrescue occurring on days 12–16 of the luteal phase. Insummary, r-hFSH alone during the pre-ovulatory interval is adequatefor the gametogenic events required to produce embryos thatdevelop either in vitro or in vivo; however, exposure to r-hLHmay improve embryo viability and the rate of development.