David Vantman

Mechanism of action of hormonal preparations used for emergency contraception: a review of the literature

This paper focuses on the research efforts undertaken to understand how emergency contraception (EC) methods act to prevent pregnancy and to identify what is known and what are the important gaps that need to be addressed. Divided into five sections the first section presents a discussion on the background of the review and a brief description of the mode of use efficacy and most common side effects of the Yuzpe regimen levonorgestrel (LNG) and mifepristone. Section 2 includes studies on the effects of postcoital steroid administration upon fertility in nonprimate animal models. Section 3 focuses on the effects of estrogens progestins or the antiprogestin mifepristone administered in the preovulatory period to macaques and the New World monkey Cebus apella. Section 4 highlights clinical studies on the effects of the Yuzpe regimen administered before and after the luteinizing hormone surge and on progesterone-regulated endometrial proteins. Finally section 5 identifies some of the most difficult areas of the literature that need to be researched.

A low sperm concentration does not preclude fertility in men with isolated hypogonadotropic hypogonadism after gonadotropin therapy

In order to define the minimal number of sperm needed for conception, we studied semen characteristics of men with isolated hypogonadotropic hypogonadism (IHH) who became sperm-positive during gonadotropin therapy. Twenty-two of 24 men (92%) proved fertile, initiating a total of 40 pregnancies. The mean (+/- standard error of the mean) sperm concentration at the time of conception was 16.7 +/- 4.0 X 10(6)/ml. However, 71% of pregnancies were conceived when the mean sperm concentration was less than 20 X 10(6)/ml; in 16%, the mean sperm concentration was less than 1 X 10(6)/ml. Mean total sperm count correlated highly with sperm concentration (r = 0.67, P less than 0.001). We conclude that men with IHH can initiate conception even when their sperm concentration is well below the conventional lower limit of 20 X 10(6)/ml.

Metformin augments the levels of molecules that regulate the expression of the insulin-dependent glucose transporter GLUT4 in the endometria of hyperinsulinemic PCOS patients

STUDY QUESTION Does treatment with the insulin sensitizer metformin modify the levels and activation of proteins related to the expression of the insulin-dependent glucose transporter (GLUT4), such as adenosine monophosphate-activated protein kinase (AMPK) and myocyte enhancer factor 2A (MEF2A), in endometria from hyperinsulinemic hyperandrogenemic polycystic ovary syndrome (PCOS h-Ins) patients? SUMMARY ANSWER In PCOS h-Ins patients, metformin increases endometrial levels of GLUT4 mRNA and protein levels by normalizing the quantity and activation of molecules that regulate GLUT4 expression to healthy values. These changes could improve endometrial metabolic function. WHAT IS ALREADY KNOWN PCOS is an endocrine-metabolic disorders closely associated with insulin resistance. In particular, the insulin signaling pathway is impaired in endometria from these patients and the concentration of GLUT4, as well as the molecules involved in its translocation to the cell surface, is decreased. However, there are limited data about the mechanisms that regulate the GLUT4 expression in the endometria and the effect of metformin on them. STUDY DESIGN, SIZE AND DURATION This is a case-control study in the setting of a research unit, approved by the Ethical Committees of our institution. The groups whose endometria were studied were PCOS h-Ins (n = 8); PCOS patients with hyperandrogenemia hyperinsulinemia taking only metformin for at least 3 months (PCOS-MTF, n = 8) and healthy fertile women at the time of hysterectomy because of benign pathology as controls (CE, n = 8). PARTICIPANTS/MATERIALS, SETTING, METHODS Steroids and sex hormone-binding globulin were measured and glucose and insulin levels were evaluated during an oral glucose tolerance test. Protein levels for αAMPK (catalytic subunit of AMPK), phosphorylated (p)-AMPKαThr(172) (activating phosphorylation site), MEF2A, p-MEF2AThr312 (activating phosphorylation site) and GLUT4 were assessed by western blot and immunohistochemistry. In addition, GLUT4 gene expression was evaluated by RT-PCR. MAIN RESULTS AND THE ROLE OF CHANCE We found significantly lower levels of MEF2A and p-MEF2AThr312 in PCOS h-Ins compared with CE endometria (P < 0.05). Also, we detected lower levels of p-AMPKαThr(172) in PCOS h-Ins endometria compared with the PCOS-MTF group (P < 0.05). The ratios of phospho-AMPK/total AMPK and phospho-MEF2A/total MEF2A were significantly increased in the PCOS-MTF compared with the PCOS h-Ins group (P < 0.05). The RT-PCR experiments showed lower levels of GLUT4 mRNA transcripts in PCOS h-Ins compared with PCOS-MTF-treated group (P < 0.05), the protein levels of GLUT4 were decreased in a similar way. LIMITATIONS, REASONS FOR CAUTION The limited number of patients included in this study who presented large clinical variability. Therefore, it would be necessary to recruit a greater number of patients to minimize our data dispersion in order to prove the clinical benefits of metformin described by others. WIDER IMPLICATIONS OF THE FINDINGS Since the insulin sensitizer metformin increases the expression of the GLUT4, it may improve endometrial physiology in PCOS patients and, therefore, promote better reproductive outcomes. These results suggest that in PCOS patients, metformin may act directly at the endometrial level and decrease insulin resistance condition by increasing the expression of GLUT4 and, in this way, indirectly restore endometrial function. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by Fondo Nacional de Desarrollo Científico y Tecnológico (grant number 1095127 to M.V.). None of the authors has any conflict of interest to declare.

Role of the transcriptional factors FOXO1 and PPARG on gene expression of SLC2A4 in endometrial tissue from women with polycystic ovary syndrome

Fifty to seventy percent of patients with polycystic ovary syndrome (PCOS) present hyperinsulinemia. On the other hand, reports indicate that forkhead box class O 1 (FOXO1) and peroxisome proliferator-activated receptor-gamma (PPARG) are involved in the insulin signaling pathway, regulating the gene expression of SLC2A4 (GLUT4). The negative effect of FOXO1 over PPARG transcription disappears when FOXO1 is phosphorylated (p-FOXO1) and excluded from the nucleus, whereas PPARG can suppress gene expression of SLC2A4. Scarce knowledge is available in endometrium of women with PCOS and hyperinsulinemia (PCOSE h-Ins) about the role of these factors. We aimed to evaluate whether the endocrine and metabolic status of PCOS modify the levels of gene and protein expression of FOXO1, PPARG, and SLC2A4 in the endometria from hyperinsulinemic PCOS women compared with controls. In endometria from control (CE, n=7) or PCOSE h-Ins (n=7), we determined the subcellular location and protein levels of p-FOXO1Ser319 and FOXO1/FOXO4 by immunohistochemistry and western blot respectively; gene and/or protein levels of PPARG and SLC2A4 were evaluated by RT-PCR and/or western blot. Cytoplasm location for FOXO1 and p-FOXO1Ser319 was immunodetected in both groups of endometria, showing significantly higher staining in PCOSE h-Ins for these proteins (P<0.05). In PCOSE h-Ins, gene and protein levels of PPARG were significantly higher than in CE, whereas SLC2A4 mRNA was decreased (P<0.05). In conclusion, the derepression of PPARG transcription by the high levels of p-FOXO1Ser319 could partially account for the lower levels of SLC2A4 found in PCOSE h-Ins, suggesting an alteration of the endometrial function in these patients.

Levels of Rabs and WAVE family proteins associated with translocation of GLUT4 to the cell surface in endometria from hyperinsulinemic PCOS women

BACKGROUND Polycystic ovary syndrome (PCOS) is an endocrine-metabolic disorder highly associated with insulin resistance and compensatory hyperinsulinemia. It is known that the insulin signaling pathway is impaired in endometria from PCOS hyperinsulinemic women, but no information is available about molecules associated with cell surface GLUT4 translocation. We therefore evaluated the protein levels of AS160 target molecules, Rab8A and Rab10, and the WAVE family proteins involved in the cortical-actin remodeling, Neural Wiskott-Aldrich Syndrome Protein (N-WASP) and WASP, in endometria from hyperinsulinemic PCOS women and controls. METHODS Protein levels were assessed by western blot, immunohistochemistry and immunofluorescence in proliferative (PE = 7) and secretory (SE = 7) phase endometria from control women and in endometria from hyperinsulinemic PCOS women (PCOS h-INS = 7). RESULTS Similar levels were detected for Rab10 in the three studied groups; however, Rab8A levels decreased in SE (P < 0.05) while higher levels were obtained in PCOSE h-INS compared with PE (P < 0.05). In the normal menstrual cycle, Neural Wiskott-Aldrich syndrome protein (N-WASP) and WASP levels were increased in SE versus PE (P < 0.05), but in PCOSE h-INS, the levels were diminished compared with PE (P < 0.05). CONCLUSIONS SE is characterized by protein expression changes associated with glucose uptake. In endometria from PCOS women with hyperinsulinemia, reduced levels of WAVE family proteins could compromise the cell surface GLUT4 exposure and the consequent glucose uptake in this tissue.

The conversion of dehydroepiandrosterone into androst-5-ene-3β,17β-diol (androstenediol) is increased in endometria from untreated women with polycystic ovarian syndrome

The changes in endometrial homeostasis found in women with polycystic ovarian syndrome (PCOS) could be associated with alterations in the intracrine metabolism of steroid hormones. The uptake of dehydroepiandrosterone-sulphate (DHEA-S), precursor of the intracrine pathway, is achieved by transporters, such as organic anion transporter polypeptides (OATPs), and molecules with oestrogenic activity, such as androst-5-ene-3beta,17beta-diol (androstenediol), can be generated. We aimed to determine androstenediol generation and the expression of OATPs in human endometria throughout the menstrual cycle and in endometria from PCOS women. Endometrial samples were obtained from control women in the proliferative phase (control endometria (CEp), n=7), secretory phase (CEs, n=7), and from PCOS patients (PCOSEp, n=7). The mRNA levels of OATP-B, OATP-D and OATP-E were measured by reverse transcriptase polymerase chain reaction (RT-PCR) and protein levels of OATP-E by immunofluorescence; 3beta-hydroxysteroid dehydrogenase (HSD) by immunohistochemistry/Western blot; the metabolism of DHEA to androstenediol was evaluated by thin layer chromatography-high-performance liquid chromatography (TLC-HPLC). Lower levels of OATP-E transcript were obtained in PCOSEp (p<0.05) compared with CEp, while OATP-E protein levels (p<0.05) and DHEA conversion to androstenediol (p<0.01) were higher in PCOSEp. Lower 3beta-(hydroxysteroid dehydrogenase) HSD protein levels were found in PCOSEp (p<0.05) (Western blot, immunohistochemistry). These results reveal a higher capacity of the endometria from PCOS women to metabolise DHEA to androstenediol, which, coupled with the high oestrogen sensitivity previously found in these endometria, may account for the increase in cell proliferation in PCOSEp already reported.

Absence of Y chromosome microdeletions in patients with cryptorchidism and hypospadias.

Microdeletions of the Y chromosome have been observed in some patients with cryptorchidism and severe defects of spermatogenesis. We investigated whether microdeletions of the Y chromosome may be present in patients with cryptorchidism and hypospadias. Peripheral blood was obtained from 20 male patients 5.8 +/- 4.1 years (range: 0.4-14 years) with cryptorchidism and hypospadias for somatic DNA analysis of Y chromosome using multiplex polymerase chain reaction. These patients had no identifiable genetic syndrome, other genitourinary malformations or an abnormal karyotype. We evaluated the presence or absence of amplification using a set of 34 different sequence-tagged sites (STS) in each patient. All patients showed normal length amplifications for each of the regions evaluated, suggesting that microdeletions of the Y chromosome are not a frequent cause of hypospadias associated with cryptorchidism.

Presence of angiotensin II and expression of angiotensin II type-2 receptor in human fallopian tube

OBJECTIVE To determine the presence of angiotensin II and angiotensin II type-2 receptor subtype messenger RNA (mRNA) in human fallopian tube. DESIGN Frozen fallopian tubes were used for all studies. SETTING Procedures were performed at the Institute of Maternal and Child Research, School of Medicine, University of Chile and San Borja-Arriarán Clinical Hospital, National Health Service, Santiago, Chile. PATIENT(S) Eight patients (30-38 years) undergoing surgical sterilization or hysterectomy for benign gynecologic conditions. INTERVENTION(S) Surgery was scheduled in the proliferative or secretory stage of the menstrual cycle. MAIN OUTCOME MEASURE(S) Localization of angiotensin II by immunofluorescence studies, expression of angiotensin II type-2 receptor mRNA, and concentrations of angiotensin II receptor subtypes and binding characteristics. RESULT(S) Angiotensin II was immunolocalized mainly in blood vessel endothelium and, to a lesser extent, in stromal cells. Both binding and angiotensin II type-2 receptor mRNA were detected at high levels. No differences were seen in receptor concentration, dissociation constants or median inhibitory concentration in fallopian tubes ipsilateral or contralateral to the corpus luteum. CONCLUSION(S) Angiotensin II and angiotensin II receptor are present in human fallopian tube. Angiotensin II type-2 receptor was the main receptor subtype. These results suggest that the renin-angiotensin system may play a regulatory role in functions of the fallopian tube.

Levels of Regulatory Proteins Associated With Cell Proliferation in Endometria From Untreated Patients Having Polycystic Ovarian Syndrome With and Without Endometrial Hyperplasia.

Polycystic ovarian syndrome (PCOS) has been associated with endometrial hyperplasia and cancer. The aim of this study was to establish whether the expression of proliferation regulatory proteins in the endometria of patients having PCOS, with or without hyperplasia, differs from control women. Control endometria (CE), patients having PCOS without and with endometrial hyperplasia (PCOSE and HPCOSE, respectively), and that of women with endometrial hyperplasia (HE) were used. The phosphorylated estrogen receptor form (pERα), similar to mother against decapentaplegic (SMAD) 2, SMAD3, and SMAD4, vascular epithelial growth factor (VEGF), and phosphorylated SMAD (pSMAD) 2 and pSMAD3 were detected by immunohistochemistry or Western blot. The results show higher levels of pERα in HE versus CE (P < .05), while higher VEGF levels were found in PCOSE and HE (P < .05) compared to CE; SMAD2 diminished in HE (P < .05) versus CE. Consequently, the higher levels of VEGF and pERα in PCOSE could represent early changes in the progression of PCOSE toward hyperplasia and cancer, whereas changes observed in SMAD proteins support the differential origin of the pathologies of HPCOSE and HE.

Hydrosalpinx fluid affects murine embryonic development in a coculture system with epithelial endometrial cells

OBJECTIVE The aim of the present investigation was to assess whether a coculture system protects from the effect of hydrosalpinx fluid (HF) on murine embryo development, evaluated through blastocyst cell number. DESIGN Controlled prospective study. SETTING Academic research center. PATIENT(S) Endometrium and HF from six patients and endometrium from six normal patients. INTERVENTION(S) Murine embryos were exposed to the absence or presence of different concentrations of human HF: 0% HF (control), 50% HF, 70% HF in human tubal fluid, and 100% HF, in a simple culture system (SCS), epithelial coculture system (ECS), and hydrosalpinx epithelial coculture system (HECS). MAIN OUTCOME MEASURE(S) Embryonic development at 72 hours and blastocyst cell number determined by the Tarcowsky method. RESULT(S) In SCS, 91.9% of the embryos reached the blastocyst stage, and no significant differences were shown in the presence of HF. However, significant differences were observed in the blastocyst cell number. Of the embryos cultured in ECS, 97.1% reached the blastocyst stage, and high concentrations of HF caused a decrease in embryonic development. A significant difference was observed between ECS and HECS in embryo development without HF. When HF was added, a significant decrease in blastocyst cell number was seen in embryos exposed to HECS compared with ECS. CONCLUSION(S) Our data suggest that normal and hydrosalpinx endometria do not protect from the deleterious effect of HF on embryo development at the concentrations evaluated. This effect is dose dependent and was determined through the blastocyst cell number.