Modulation of gonadotrophin induced steroidogenic enzymes in granulosa cells by d-chiroinositol.

BACKGROUND: d-chiroinositol (DCI) is a inositolphosphoglycan (IPG) involved in several cellular functions that control the glucose metabolism. DCI functions as second messenger in the insulin signaling pathway and it is considered an insulin sensitizer since deficiency in tissue availability of DCI were shown to cause insulin resistance (IR). Polycystic ovary syndrome (PCOS) is a pathological condition that is often accompanied with insulin resistance. DCI can positively affects several aspect of PCOS etiology decreasing the total and free testosterone, lowering blood pressure, improving the glucose metabolism and increasing the ovulation frequency. The purpose of this study was to evaluate the effects of DCI and insulin combined with gonadotrophins namely follicle-stimulating hormone (FSH) and luteinizing hormone (LH) on key steroidogenic enzymes genes regulation, cytochrome P450 family 19 subfamily A member 1 (CYP19A1) and cytochrome P450 side-chain cleavage (P450scc) in primary cultures of human granulosa cells (hGCs). We also investigated whether DCI, being an insulin-sensitizer would be able to counteract the expected stimulator activity of insulin on human granulosa cells (hGCs). METHODS: The study was conducted on primary cultures of hGCs. Gene expression was evaluated by RT-qPCR method. Statistical analysis was performed applying student t-test, as appropriate (P < 0.05) set for statistical significance. RESULTS: DCI is able to reduce the gene expression of CYP19A1, P450scc and insulin-like growth factor 1 receptor (IGF-1R) in dose-response manner. The presence of DCI impaired the increased expression of steroidogenic enzyme genes generated by the insulin treatment in gonadotrophin-stimulated hGCs. CONCLUSIONS: Insulin acts as co-gonadotrophin increasing the expression of steroidogenic enzymes genes in gonadotrophin-stimulated granulosa cells. DCI is an insulin-sensitizer that counteracts this action by reducing the expression of the genes CYP19A1, P450scc and IGF-1R. The ability of DCI to modulate in vitro ovarian activity of insulin could in part explain its beneficial effect when used as treatment for conditions associated to insulin resistance.

The anti-Müllerian hormone (AMH) acts as a gatekeeper of ovarian steroidogenesis inhibiting the granulosa cell response to both FSH and LH.

PURPOSE: Anti Müllerian Hormone (AMH) has a negative and inhibitory role in many functions of human granulosa-lutein cells (hGCs) including notoriously the reduction of the aromatase CYP19A1 expression induced by follicle-stimulating hormone (FSH). No data have been provided on the possible role of AMH in modulating the response to luteinizing hormone (LH) (alone or combined with FSH) as well as its effect on other enzymes involved in steroidogenesis including aromatase P450scc. The aim of this study was to investigate the role of AMH as regulator of the basal and stimulated steroids production by hGCs. METHODS: Primary culture of hGCs were incubated with hormones AMH, LH, and FSH, alone or in combination. The CYP19A1 and P450scc messenger RNA (mRNA) expression, normalized by housekeeping ribosomal protein S7 (RpS7) gene, was evaluated by reverse transcriptase quantitative PCR (RT-qPCR). Each reaction was repeated in triplicate. Negative controls using corresponding amount of vehicle control for each hormone treatment were performed. RESULT: AMH did not modulate the basal mRNA expression of both aromatase genes at any of the concentrations tested. Meanwhile, the strong mRNA induction of CYP19A1 and P450scc generated by a 24-h gonadotropin treatment (alone and combined) was suppressed by 20 ng/ml AMH added to culture medium. CONCLUSIONS: These findings contribute in clarifying the relationship between hormones regulating the early phase of steroidogenesis confirming that AMH is playing a suppressive role on CYP19A1 expression stimulated by gonadotropin in hGCs. Furthermore, a similar inhibitory effect for AMH was observed on P450scc gene expression when activated by gonadotropin treatment.

Effect of oral contraceptives containing estradiol and nomegestrol acetate or ethinyl-estradiol and chlormadinone acetate on primary dysmenorrhea.

OBJECTIVE: To study the three cycles effect on primary dysmenorrhea of the monophasic 24/4 estradiol/nomegestrol acetate (E2/NOMAC) and of the 21/7 ethinyl-estradiol/chlormadinone acetate (EE/CMA) oral contraceptive. The tolerability and the effect of both preparations on metabolism and health-related quality of life were also evaluated. DESIGN: Prospective observational cohort study. SETTING: Tertiary gynecologic center for pelvic pain. PATIENTS: Subjects with primary dysmenorrhea requiring an oral contraceptive, who spontaneously selected either E2/NOMAC (n = 20) or EE/CMA (n = 20). MAIN OUTCOME MEASURES: Visual Analogue Scale (VAS) score for dysmenorrhea, Short Form-36 questionnaire for health-related quality of life, lipoproteins and days of menstrual bleeding (withdrawal bleeding during oral contraceptive). RESULTS: Mean age and body mass index (BMI) were similar between the two groups. The final analysis was performed on 34 women, 15 in E2/NOMAC and 19 in EE/CMA group. Compliance with treatment was significantly higher with EE/CMA (100%) than E2/NOMAC (75%) (p = 0.02). Both treatments significantly (p < 0.0001) reduced VAS of primary dysmenorrhea, similarly (E2/NOMAC by a mean of 74.7%, EE/CMA by a mean of 78.4%; p = 0.973). Only E2/NOMAC significantly increased SF-36 score (p = 0.001), both in physical (p = 0.001) and mental domains (p = 0.004). The mean number of days of menstrual bleeding was significantly reduced in E2/NOMAC group (from 4.86 ± 1.20 d to 2.64 ± 1.59 d, p = 0.0005 versus baseline, p = 0.007 versus EE/CMA group). BMI did not vary in either group. E2/NOMAC did not change lipoproteins and apoproteins while EE/CMA increased total cholesterol (p = 0.0114), HDL-cholesterol (p = 0.0008), triglycerides (p = 0.002), apoprotein-A1 (Apo-A1; p = 0.0006) and apopoprotein-B (Apo-B; p = 0.008), decreasing LDL/HDL ratio (p = 0.024). CONCLUSIONS: Both oral contraceptives reduced similarly primary dysmenorrhea, with E2/NOMAC also reducing withdrawal bleedings and being neutral on lipid metabolism.

Nitric Oxide concentration in endometrial washing throughout the menstrual cycle.

PURPOSE: Nitric oxide (NO) intrauterine production has been shown to have an important role in the reproductive system in females. The objective of the present study was to evaluate NO concentration in endometrial washing throughout the menstrual cycle. METHODS: Observational study at Institute of Obstetrics and Gynecology, Mother-Infant Department, University Hospital. The study included 40 healthy fertile women, aged 21-40, with regular menstrual cycle, undergoing endometrial washing by hydrosonography for the assessment of NO concentration. RESULTS: Nitric oxide concentration in endometrial washing were low in early to mid proliferative phase (4.73 ± 1.57 mcM/L), but significantly higher (p < 0.05) in late proliferative phase (7.30 ± 3.37 mcM/L) early secretory phase (8.05 ± 1.97 mcM/L) and late secretory phase (8.69 ± 4.12 mcM/L) of menstrual cycle. CONCLUSIONS: Endometrial washing by hydrosonography is a simple, minimally invasive, and effective tool to use in the evaluation of cyclical NO intrauterine production. Nitric oxide concentrations increased during the late proliferative and secretory phase of menstrual cycle.

Route of administration of contraceptives containing desogestrel/etonorgestrel and insulin sensitivity: a prospective randomized study.

BACKGROUND: The study was conducted to investigate whether hormonal contraceptives administered via the oral and vaginal route exert a similar effect on insulin sensitivity (SI). STUDY DESIGN: This is a prospective, randomized study performed in the University Hospital. Subjects were healthy lean young women, needing a hormonal contraceptive, randomly allocated to receive for 6 months (a) an oral contraceptive (OC) containing 30 mcg ethinylestradiol (EE)/150 mcg desogestrel (DSG) (high-estrogen group; n=12), (b) an OC containing 20 mcg EE/150 mcg DSG (low-estrogen group; n=12) and (c) a vaginal ring contraceptive releasing, per day, 15 mcg EE/120 mcg etonorgestrel, the active DSG metabolite (n=12). SI and glucose utilization independent of insulin (Sg) were evaluated by the minimal model method. Modifications of total, high-density lipoprotein (HDL) and low-density lipoprotein cholesterol and triglycerides were also evaluated. RESULTS: Sg did not vary with any treatment. SI decreased during OCs (5.74+/-0.49 vs. 3.86+/-0.44; p=.0005), independently of the high/low-estrogen dose. SI did not decrease during vaginal ring use (4.64+/-1.03 vs. 5.25+/-1.36; p=.57; p=.019 vs. oral). Total cholesterol and HDL cholesterol increased (p=.02) during OCs, independently of the dose. Triglycerides increased during both oral (p=.01) and vaginal (p=.032) contraceptive use. CONCLUSIONS: The present data indicate that in contrast to OC use, vaginal contraception with the ring does not deteriorate SI. The vaginal ring may represent an appropriate choice for long-term contraception in women at risk for developing diabetes mellitus or metabolic syndrome.

Levonorgestrel administration in emergency contraception: bleeding pattern and pituitary-ovarian function.

BACKGROUND: This study was conducted to evaluate the effects of levonorgestrel administration for emergency contraception (EC) on bleeding pattern and pituitary-ovarian function. STUDY DESIGN: In 69 women with a reported stable menstrual cycle length of 24-34 days, we investigated bleeding patterns following EC administration in the follicular (n=26), periovulatory (n=14) and luteal (n=29) phase. In a subgroup of 8 women, hormonal evaluation and ultrasonography were performed. RESULTS: EC taken in the follicular, but not in the periovulatory or luteal phase, significantly shortened cycle length by 10.9+/-1 days. The subsequent cycle was not affected. EC taken in the late preovulatory phase, prior to the gonadotrophin surge, suppressed ovulation (n=7), while ovulation was not blocked when EC was given during an ongoing luteinizing hormone (LH) pulse (n=1). CONCLUSIONS: Our data indicate that EC given before the onset of the luteinizing hormone (LH) surge inhibits ovulation and hastens the end of the current menstrual cycle. Subsequently, the length of the following menstrual cycle returned as prior to treatment. By contrast, levonorgestrel administered after the expected ovulation has no effect on menstrual cycle length.

Effects of two different oral contraceptives on homocysteine metabolism in women with polycystic ovary syndrome.

PURPOSE: This study was conducted to evaluate the effects of two different oral contraceptives (OCs) on homocysteine (Hcy) metabolism in 20 women with polycystic ovary syndrome (PCOS). METHODS: Women were randomly allocated to receive either the biphasic OC containing 40/30 mug ethynylestradiol (EE)+25/125 mug desogestrel (DSG; n=10) or the monophasic OC containing 35 mug EE and 2 mg cyproterone acetate (CPA; n=10). Investigations were performed before and after 6 months of treatment. Fasting vitamin B(12), folate, Hcy and insulin sensitivity (SI), and glucose utilization independent of insulin (Sg), by the minimal model method, were evaluated. RESULTS: Folate and vitamin B(12) were not significantly modified by either OC. EE/DSG decreased SI (2.53+/-0.35 vs. 1.68+/-0.45; p<.05), without modifying Hcy (9.54+/-0.7 micromol/L vs. 9.18+/-0.6 micromol/L). EE/CPA improved SI (1.47+/-0.38 vs. 3.27+/-0.48; p<.04) and decreased Hcy (9.8+/-1.9 micromol/L vs. 7.9+/-0.9 micromol/L; p<.05). This study indicates that in women with PCOS, EE/CPA, but not EE/DSG, improves IS and decreases fasting Hcy.

Effect on insulin sensitivity of Implanon vs. GnRH agonist in women with endometriosis.

OBJECTIVES: To evaluate the influence of two medical treatments for endometriosis on insulin sensitivity. STUDY DESIGN: After surgery, 26 women with endometriosis were randomly allocated to a 6-month treatment with a GnRH agonist (Leuprorelin 3.75 mg/28 days) or a subdermal progestin implant (etonogestrel 68 mg). Insulin sensitivity (SI) and glucose utilization independent of insulin (Sg) were investigated at baseline and after 6 months by a frequently sampled intravenous glucose tolerance test (FSIGT) associated with the minimal model method. RESULTS: Both therapies tended to decrease SI, but the effect did not reach statistical significance in the GnRH agonist group (5.43+/-1.29 vs. 3.99+/-0.8) and was significant in the etonogestrel group (5.74+/-1.12 vs. 3.95+/-0,78; p=.046). Sg, fasting glucose, insulin, C-peptide and C-peptide/insulin were not modified by either treatment. CONCLUSIONS: The modifications of glucose-insulin metabolism induced by the GnRH agonist are of no relevance for the short-term use of this molecule. Even if the modification induced by the etonogestrel implant is subtle and of no major impact, it should be taken into consideration for the long-term treatment of individuals with abnormalities of glucose-insulin metabolism.