UPA and LNG in emergency contraception: the information by EMA and the scientific evidences indicate a prevalent anti-implantation effect.

RATIONALE AND OBJECTIVES: Emergency contraceptives pills (ECPs) are described as drugs that work by either inhibiting or delaying ovulation without affecting implantation. In our opinion, as we aim at demonstrating, both EMA documents and the experimental papers indicate that they prevalently inhibit embryo-implantation. LNG-ECPs: literature: LNG-ECPs never prevent ovulation when are taken in the most fertile days (EMA-EPAR on ellaOne® p. 9, first table). Conversely, they prevent the formation of an adequate corpus luteum. When they are taken pre-ovulatory ovulations occur regularly, but pregnancies do not appear. Taken after ovulation, they seem ineffective in preventing pregnancies. UPA-ECPs: literature: EllaOne® prevents ovulation only when is taken in the first fertile day. Thereafter, its anti-ovulatory effect drops sharply and becomes insignificant (8%) 36 h before ovulation, in the most fertile days (Brache); its decreasing anti-ovulatory effect cannot explain a consistently high effectiveness in preventing pregnancies (≥80%) that does not decrease depending on which of the 5 d it is taken after unprotected intercourse. Besides, ovulation occurs regularly in 91.7% of women taking ellaOne® weekly, for eight consecutive weeks (EMA-CHMP-Assessment Report 'EMA/73099/2015': study HRA2914-554, p. 7). Lastly, Lira-Albarrán administered ellaOne® to women in the most fertile pre-ovulatory days: they had normal ovulation, but their endometrium, evaluated through samples obtained in the implantation window, became inhospitable: the expression of 1183 genes was exactly the opposite of that observed in the receptive pro-gestational endometrium. This agrees with information by EMA-CHMP-Assessment Report 'EMEA/261787/2009' (p. 8): after UPA administration 'the proteins necessary to begin and maintain pregnancy are not synthesized'. CONCLUSIONS: Emergency Contraceptives work prevalently by preventing embryo-implantation. People shall receive correct information.

Does levonorgestrel emergency contraceptive have a post-fertilization effect? A review of its mechanism of action.

BACKGROUND: Recent studies have identified that levonorgestrel administered orally in emergency contraception (LNG-EC) is only efficacious when taken before ovulation. However, the drug does not consistently prevent follicular rupture or impair sperm function. OBJECTIVE: The present systematic review is performed to analyze and more precisely define the extent to which pre-fertilization mechanisms of action may explain the drug's efficacy in pregnancy avoidance. We also examine the available evidence to determine if pre-ovulatory drug administration may be associated with post-fertilization effects. CONCLUSION: The mechanism of action of LNG-EC is reviewed. The drug has no ability to alter sperm function at doses used in vivo and has limited ability to suppress ovulation. Our analysis estimates that the drug's ovulatory inhibition potential could prevent less than 15 percent of potential conceptions, thus making a pre-fertilization mechanism of action significantly less likely than previously thought. Luteal effects (such as decreased progesterone, altered glycodelin levels, and shortened luteal phase) present in the literature may suggest a pre-ovulatory induced post-fertilization drug effect. LAY SUMMARY: Plan B is the most widely used emergency contraceptive available. It is important for patients and physicians to clearly understand the drug's mechanism of action (MOA). The drug was originally thought to work by preventing fertilization. Recent research has cast doubt on this. Our review of the research suggests that it could act in a pre-fertilization capacity, and we estimate that it could prevent ovulation in only 15 percent or less of cases. The drug has no ability to alter sperm function and limited ability to suppress ovulation. Further, data suggest that when administered pre-ovulation, it may have a post-fertilization MOA.

Insulin and body weight but not hyperandrogenism seem involved in seasonal serum 25-OH-vitamin D3 levels in subjects affected by PCOS.

PCOS patients were frequently characterized by lower plasma vitamin D levels. The mechanisms involved in this dysfunction remains still debated, therefore we evaluated the role of androgen, insulin and body weight on the serum VitD levels in women with or without PCOS. Eighty one patients 18-42 yrs old were studied into "SUMMER" and "WINTER" seasonal period: thirty seven PCOS, seventeen no-ovarian hyperandrogenic (noPCOS), twelve functional hypothalamic amenorrhea (FHA) and finally fifteen healthy (Con). Patients were further divided into: lean (L), obese (O), normo- (nINS) and hyperinsulinemic (hINS). All hormonal and metabolic parameters were measured at 1-7 days of the menstrual cycle. Our results show that VitD levels were lower in PCOS and in noPCOS than in FHA and Con, in particular in (O) and (hINS) PCOSs. Both in summer and in winter, PCOSs had basal VitD levels significantly lower than FHA and Con, whereas they were similar to noPCOS. Yet, LhINS and OPCOS had VitD levels lower than Con and noPCOS. VitD levels were comparable in LnINS PCOS and Con. In conclusion, PCOSs had levels of VitD lower than controls. Weight and hyperinsulinemia had a significant influence on these values. Finally, over 70% of our healthy patients had VitD deficiency.

Ulipristal acetate: critical review about endometrial and ovulatory effects in emergency contraception.

The effectiveness of emergency contraception (EC) is usually estimated by comparing the number of observed pregnancies to that of expected pregnancies after unprotected intercourse. Second-generation selective progesterone receptors modulators have been developed and evaluated for EC use. Among these compounds, ulipristal acetate (UPA) has been proven to share the same antiprogestin activity as mifepristone, and as with mifepristone, UPA has been demonstrated to be effective up to 120 hours after unprotected intercourse. The UPA is more effective than levonorgestrel (LNG) in preventing the appearance of clinically evident pregnancies. The LNG delays ovulation only when taken at the beginning of the fertile period; taken later, it is ineffective on ovulation, while it has been proven to impair the subsequent luteal function. The effectiveness of LNG decreases as time elapses and is limited to 72 hours after unprotected intercourse. The UPA maintains consistent effectiveness for 5 days after unprotected intercourse, and this effectiveness is independent on which of these 5 days it is taken. The ability of UPA to delay ovulation decreases progressively as ovulation approaches and is null at the time of the luteinizing hormone (LH) peak: 1 to 2 days before ovulation, UPA behaves as a placebo. The persistent effectiveness of the drug cannot be due to antiovulatory action, as it decreases sharply as LH approaches its peak level. The effectiveness is most likely due to the dramatic endometrial effects of the drug that are produced regardless of when it is taken. These effects are consistently present, as the threshold for altering endometrial morphology is lower than the threshold for altering folliculogenesis.

Magnetic resonance-guided focused ultrasound myomectomy: safety, efficacy, subsequent fertility and quality-of-life improvements, a systematic review.

We performed a systematic review about studies reporting data of myomectomy performed by magnetic resonance-guided focused ultrasound (MRgFUS) technique in order to define its safety, feasibility, indications, complications, and impact on uterine fibroid symptom and health-related quality of life (UFS-QOL) and fertility. Outcomes were considered according to fibroids shrinkage, nonperfused volume (NPV), NPV ratio, and uterine fibroid symptoms assessed with UFS-QOL questionnaire (baseline 3, 4, 6, and 12 months). We analyzed 38 eligible studies reporting outcomes about 2500 patients (mean age 43.67 years). The MRgFUS results a safe, efficient, and cost-effective minimal invasive technique for treatment of uterine fibroids. Increasing experience, device improvements, and availability for a larger number of patients are enhancing the outcomes, while the obstetrical ones should be more extensively explored. The MRgFUS could be considered as a minimal invasive alternative to traditional surgical or radiological procedures for the treatment of symptomatic uterine myomas improving both QOL and subsequent fertility.

Cyclic variations of bone resorption mediators and markers in the different phases of the menstrual cycle.

Female hormones are very important in regulating bone homeostasis; the drop of estrogen levels occurring at menopause is linked to a dramatic prevalence of bone resorption on formation. Only a small number of studies investigated the relationship between changes in circulating female sex hormones and the markers and mediators of bone homeostasis and they showed conflicting results. To explore such relationships we studied 20 young fertile healthy women, aged between 19 and 32 years. None had received hormone treatment for at least 6 months. We assayed luteinizing hormone, follicle-stimulating hormone, progesterone and 17ß-estradiol, as well as the levels of osteoprotegerin (OPG), C-terminal telopeptide of collagen type I (CTx) and RANKL (receptor activator of nuclear factor-B ligand) in samples drawn from every subject at four different times during the menstrual cycle when estrogens are lowest, at the start of the cycle: T 0 (2-4th day); when estrogens are highest, in the pre-ovulatory period: T 14 (12-14th day); when progesterone activity is highest, in the advanced luteal phase: T 26 (24-26th day); and again at the start of the next cycle: T 01 (2-4th day). We observed that CTx levels are highest at the start of the cycle, decreased significantly from T 0 to T 26 (pfwe = 0.0455) and then increased from T 26 to T 01 (pfwe = 0.0415); OPG, on the other hand, which was also highest at the start of the cycle, decreased significantly from T 0 to T 14 (pfwe = 0.02) and then increased, though not significantly, from T 14 to T 01; no variation was observed in RANKL values at any time. We observed inverse correlations between estradiol and OPG levels, which became highly significant at T 01 between estradiol nadir and OPG peak levels (pfw = 0.0095). Furthermore, the increase of estradiol from T 0 to T 14 was negatively correlated with the concomitant decrease of OPG (pfwe = 0.0277), as was the fall of estradiol from T 26 to T 01 with the OPG peak levels, both at T 01 (pfw = 0.0045) and at T 0 (pfwe = 0.0381). We also observed direct correlations between the OPG levels and the variations of progesterone in the preceding intervals, but they never attained statistical significance. We conclude that OPG and CTx fluctuation during the menstrual cycle are likely due to the physiological variations of sex steroids levels.

Insulin receptor and glucose transporters mRNA expression throughout the menstrual cycle in human endometrium: a physiological and cyclical condition of tissue insulin resistance.

The expression of insulin receptor (IR), together with that of glucose transporters 1 and 4 (GLUT1-4) and of Insulin Growth Factor-I and -II (IGF-I,-II) in the endometrium of healthy and young women in both phases of menstrual cycle was assessed. Sixteen out of 20 healthy and normal menstruating volunteers were studied. Endometrial samplings were performed in every subject, twice in the same cycle, during the follicular and luteal phase respectively. The mRNA expression of IR, GLUT1-4, IGF-I and -II were evaluated by real-time quantitative RT-PCR and immunostaining reactions. Our results indicate that IR, GLUT1-4, IGF-I and -II mRNAs were expressed in both phases of the endometrial cycle: GLUT4 and IGF-I mRNA expression were significantly higher in the follicular phase and localized at the epithelial and stromal cell level, respectively, whereas IR, GLUT1 and IGF-II mRNA expression were mostly present in the secretory phase and mainly localized at the stromal level. An inverse tendency of IR and GLUT4 mRNA expression was respectively observed from follicular to luteal phase. In conclusion our data suggest that IR, glucose transporters and IGFs are significantly and differently expressed at the endometrial level throughout the menstrual cycle and that human endometrium cyclically undergoes through a transitory condition from normal to an insulin-resistance state.

How do levonorgestrel-only emergency contraceptive pills prevent pregnancy? Some considerations.

Controversial opinions exist about the possible mechanisms throughout emergency contraception prevents pregnancy. Recently, the International Federation of Gynaecology and Obstetrics and the International Consortium for Emergency Contraception released a Joint Statement declaring that 'inhibition or delay of ovulation should be their primary and possibly only mechanism of action'. They still added that 'Review of the evidence suggests that LNG-ECPs cannot prevent implantation'. Concerning levonorgestrel-only emergency contraceptive pills effects on ovulation, the Statement based on seven reference papers which considered a total of only 142 patients, divided into still different subgroups. Basing on their same references we got quite different conclusions.

Culture of cryopreserved ovarian tissue: state of the art in 2008.

This paper reviews the most relevant literature from the past 10 years on different techniques for the culture of fresh and cryopreserved ovarian tissue from animals and humans. Information on strategies for culturing whole ovarian tissue and isolated follicles are provided as well as an updated and comprehensive view of the role that growth factors have in mediating and regulating in vitro folliculogenesis.

Healthy early preantral follicle can be obtained in a culture of frozen-thawed human ovarian tissue of 32 weeks.

The objective of this study was to report morphological and functional evidence of a well-preserved preantral follicle recovered from human frozen-thawed ovarian tissue in a long-term culture. The tissue was originally obtained from a 26-year-old woman with breast cancer. The ovarian cortex was collected by laparoscopy and frozen/thawed and cultured for 32 weeks in minimum essential medium alpha-MEM, supplemented with insulin transferrine selenite (ITS), human serum (HS), antibiotics, follicle-stimulating hormone (FSH). and N-acetyl cysteine (NAC). Thawed tissue samples were examined by light microscopy (LM), transmission electron microscopy (TEM), and real-time RT-PCR. LM examination of cortical pieces after 32 weeks of culture showed a healthy early preantral follicle; TEM and real-time PCR confirmed its good state of preservation. The synergy in action of NAC and FSH plays an important role in follicle growth of ovarian tissue cultures. For the first time a well-preserved preantral follicle was found in a culture of frozen-thawed human ovarian tissue.

Oral 17beta-estradiol and sequential progesterone in menopause: effects on insulin-like growth factors and their binding proteins.

OBJECTIVE: We evaluated the acute effects of low-dose oral estradiol and sequential progesterone on the insulin-like growth factor (IGF)/growth hormone (GH) axis, IGF-binding proteins (IGFBPs) 1 and 3, and plasma levels of sex hormone-binding globulin (SHBG) in postmenopausal subjects. STUDY DESIGN: Thirty healthy normal-weight women (mean age: 54.2 +/- 5.7 years) spontaneously postmenopausal for at least 6 months were enrolled. None had used hormone replacement therapy (HRT). Appropriate investigations excluded renal, glucose, lipid and coagulation abnormalities. Breast X-ray and endometrial ultrasound examinations excluded organic pathologies. They received oral cyclical HRT for 1 year, based on the administration of oral estradiol (1 mg/day) for 28 consecutive days plus progesterone (200 mg/day) from day 15 to day 28; out of the whole group, 15 subjects received progesterone orally (group A), while in 15 progesterone was administered transvaginally (group B). On the day before treatment (T0), on day 14 (T14) and on day 28 (T28) of the first cycle, plasma levels of estradiol, progesterone, SHBG, GH, IGF-I and -II, IGFBP-1 and -3, insulin and C-peptide were assayed in all patients. The same parameters were evaluated at T14 and T28 during the 12th month of treatment. RESULTS: At T14, we observed significant increases in the levels of estradiol (from 20 +/- 16 to 115 +/- 71 pg/ml, p < 0.001), SHBG (from 132 +/- 42 to 182 +/- 55 nmol/l, p < 0.001) and IGFBP-1 (from 92 +/- 57 to 127 +/- 87 ng/ml, p < 0.004), while the level of IGF-I decreased (from 197 +/- 138 to 129 +/- 85 ng/ml, p < 0.003). At T28, progesterone levels were significantly higher in the women receiving it orally than transvaginally (8.4 +/- 6.1 vs. 3.7 +/- 3.2 ng/ml, p < 0.025). However, while oral progesterone did not affect the estrogen-induced variations, transvaginal progesterone abrogated the increase in the levels of IGFBP-1. The levels of IGF-II, IGFBP-3, GH, glucose, C-peptide and insulin did not change at any time. At 1 year, the values maintained the same trends. The estrogen-induced variations of SHBG were correlated directly with those of estradiol (r = 0.48) and inversely with those of IGF-I (r = -0.424). CONCLUSIONS: Low-dose oral estradiol reduces plasma levels of IGF-I and increases IGFBP-1 and SHBG concentrations, while GH is unchanged. These effects, significant and immediate, lead us to hypothesize a direct action of estradiol on hepatocytes.

Evidence for the presence of glucose transporter 4 in the endometrium and its regulation in polycystic ovary syndrome patients.

Glucose transporter 4 (GLUT4) seems to be involved in the mechanism of insulin resistance in polycystic ovary syndrome (PCOS) patients (PCOSs) in both muscular and adipose tissue. The observation that insulin stimulates glucose oxidation in endometrial cells led us to investigate the presence of GLUT4 in this tissue and whether a defect of GLUT4 is present at the endometrial level in PCOSs. We also investigated whether body weight influences GLUT4 expression in this syndrome. GLUT4 mRNA content was examined by real-time quantitative RT-PCR and immunostaining reaction in the endometrial tissue of nine normal subjects, nine lean and eight obese hyperinsulinemic (h-INS), and eight lean and 10 obese normoinsulinemic (n-INS) PCOSs. GLUT4 mRNA and its positive immunostaining reaction were present in epithelial cell level in the endometrium of both normal and PCOS subjects. Significantly higher levels of GLUT4 were observed in normal and lean n-INS PCOSs in comparison with other groups. In both n-INS and h-INS obese PCOSs, GLUT4 was significantly lower than in lean subjects. However, obese n-INS and lean h-INS PCOSs showed a similar low GLUT4 expression, whereas obese h-INS PCOSs showed the lowest expression when compared with other groups. In conclusion, our data demonstrate that GLUT4 is present in the endometrium of normal and PCOS subjects and that hyperinsulinism and obesity seem to have a negative effect on endometrial GLUT4 expression in PCOS.

Obesity reduces the expression of GLUT4 in the endometrium of normoinsulinemic women affected by the polycystic ovary syndrome.

GLUT4 is the most important glucose transporter in insulin-dependent tissues. A decrease of its expression by the adipocytes was reported in polycystic ovary syndrome (PCOS), regardless of obesity and glucose tolerance. In PCOS, abnormal menstrual cycles, abnormal insulin secretory patterns, and obesity, which are risk factors for endometrial diseases, frequently coexist. The endometrial effects of insulin are direct through specific insulin receptors. However, it is unknown whether the endometrium expresses GLUT4 and can be considered an insulin-regulated tissue. In this study, we investigated this question, and we investigated whether obesity modulates this expression in PCOS normoinsulinemic patients. We assayed GLUT4 in the endometrial samples from 18 normoinsulinemic PCOS patients and 9 controls in the advanced follicular phase of the cycle; 10 patients were lean and 8 obese, and all were aged between 23 and 32 years. Most tissue was immediately frozen for RT-PCR; some tissue was saved for histology and immunohistochemistry. GLUT4 mRNA expression was measured in three samples for every patient and expressed as mean +/- SE of an arbitrary unit. In obese PCOS subjects, endometrial GLUT4 expression was significantly lower than in the lean ones (24.0 +/- 6.8 vs. 65.2 +/- 24.4; P < 0.005) and the controls (53.2 +/- 10.7). Lean PCOS and control subjects showed similar values. GLUT4 immunostaining was strong in the epithelial and absent in the stromal cells. We demonstrated endometrial GLUT4 expression. The similar results in lean PCOS and control subjects suggest that endometrial GLUT4 expression is not affected by PCOS itself, whereas it is reduced by obesity in PCOS patients.