Reproductive Sciences

Ovulation induction (OI) is a cornerstone of human assisted reproduction treatments (ART). Current OI protocols are based on the human follicular dynamics model known as propitious moment theory (PMT), by which follicles continuously grow from the primordial pool without any pattern, and follicular fate depend on the occurrence of a gonadotropin surge. Recently, a new paradigm of human follicular dynamics called follicular waves was revealed using sequential ultrasound examination of 1 interovulatory interval. Instead of random growth, follicles develop in coordinated groups or waves, occurring 2 to 3 times during an interovulatory interval. Follicular waves are common in several other mono-ovulatory species, like equines and bovines. In fact, this model was applied to the development of several OI protocols in veterinary medicine, especially in cows. It has been shown that synchronization of OI with the emergence of a follicular wave increases substantially success rates in animals, even with single embryo transfer. Veterinarians have already developed mechanisms to control wave emergence through mechanical or chemical ablation of the dominant follicle or corpus luteum. Considering the follicular dynamics similarities between humans and bovines regarding the follicular wave phenomenon, we hypothesize that synchronization of follicular wave emergence with ovarian stimulation produces more competent oocytes and embryos and will enhance ART efficiency in humans. At the end of this article, we propose 2 theoretical approaches to induce the emergence of a follicular wave in women: (1) a mechanical strategy by aspiration of the dominant follicle and (2) a pharmacological strategy by administering estradiol and progesterone.

We have found that estrogen promotes suppression of feeding and a lean body mass while activating the arcuate nucleus proopiomelanocortin (POMC)-expressing neurons. These neurons, when activated, suppress appetite and increase energy expenditure. Because the increased activation of POMC neurons by estradiol was associated with increased glutamate receptor presence that enable calcium influx, we analyzed the expression of the calcium-binding protein, parvalbumin, in these hypothalamic neurons. We observed that estrogen treatment of female mice resulted in induction of parvalbumin-immunoreactivity in arcuate nucleus neurons, a large number of which was POMC-expressing. These data indicate that the increased excitatory activity induced by estradiol in the arcuate nucleus in support of suppression of appetite is associated with calcium overload of these neurons. Although parvalbumin may protect these cells from calcium overload-associated neuronal degeneration, maintenance of calcium entry may lead to increased vulnerability of POMC neurons during the course of sustained satiety.

MicroRNAs (miRs) are known to repress target genes at posttranscriptional level and play important roles in the maturation of cells. However, the expression profiles of miRs during follicular maturation have not been fully elucidated. This study was designed to investigate the expression profiles of miRs in murine follicles according to human chorionic gonadotropin (hCG) treatment and vitamin C status during in vitro culture. Ovaries were removed from the 12-day-old wild-type and vitamin C-deficient (L-gulonogammalactone oxidase knockout, Gulo—/—) C57BL6 mice. Preantral follicles were isolated and cultured in 20 µL droplets of culture media supplemented with follicle-stimulating hormone and luteinizing hormone (FSH + LH). After their full maturation, follicles were divided into 2 groups: with and without hCG treatment. Real-time polymerase chain reaction (PCR) was performed using oocytes and granulosa cells (G-cells) to evaluate the miRs known to be expressed mainly in the mouse ovary. After the addition of hCG, miR profiles showed divergent changes between oocytes and G-cells. These profiles significantly differed from those of hCG(—) group. Compared to wild type, Gulo—/— mice showed altered miR profiles in matured oocytes and G-cells. Conclusively, hCG supplementation and vitamin C status alter the miR expression profiles in oocytes and G-cells during in vitro growth of murine follicles.

Rationale: The mechanism of atherogenesis includes leukocyte adhesion to endothelial cells followed by migration into the subendothelial space. The polysialylated neural cell adhesion molecules (PSA-nCAMs) are a group of hydrophilic neural cell adhesion molecule (NCAM) isoforms that inhibit NCAM: NCAM association, thereby blocking cell: cell adhesion. During previous studies, we demonstrated that sialylation of specific NCAMs are upregulated at proestrus in the rat and that PSA-nCAM is expressed by the rat vascular endothelium. Methods and Results: In this study, we sought the presence of PSA-nCAM in human vessels and regulation of its expression in estradiol-treated human umbilical vascular endothelial cells (HUVEC). Immunoreactive PSA-nCAM (ir-PSA-nCAM) was shown in blood and lymph vessels of adult rats and human brain, skin, liver, lung, cervix, endometrium, and ovary. Staining for ir-PSA-nCAM was present on the glycocalyceal surface of estradiol-treated HUVEC, but not in the presence of the estrogen receptor (ER)-blocker fulvestrant. Western blotting confirmed these findings. Conclusions: PSA-nCAM is widely present in the glycocalyx of human and rat vascular endothelium. It also is expressed by HUVEC, in which it is induced by estradiol. The estrogen-regulated presence of vascular PSA-nCAM could diminish NCAM-dependent interactions between vessels and circulating leukocytes, thereby impeding vascular inflammation and atherogenesis, and, contributing to estrogen-induced cardioprotection. This hypothesized action is presently under study.

Although pains of various kinds top the list of complaints from women with endometriosis and are the most debilitating of the disease, little is known about the mechanism/mechanisms of endometriosis-associated pains. To test the hypothesis that women with endometriosis have generalized hyperalgesia which may be alleviated by a successful surgery, we recruited 100 patients with surgically and histologically confirmed endometriosis and 70 women without, and tested their responses to pain stimulations. Before the surgery, all patients rated their dysmenorrhea severity by Visual Analog scale (VAS) and went through an ischemic pain test (IPT) and an electrical pain test (EPT). The controls were also administrated with IPT/EPT. Three and 6 months after surgery, all patients were administrated with IPT/EPT and rated their severity of dysmenorrhea. We found that patients with endometriosis had significantly higher IPT VAS scores and lower EPT pain threshold than controls, but after surgery their IPT scores and EPT pain threshold were significantly and progressively improved, along with their dysmenorrhea severity. Thus, we conclude that women with endometriosis have generalized hyperalgesia, which was alleviated by surgery. Consequently, central sensitization may be a possible mechanism underlying various forms of pain associated with endometriosis, and its recognition should have important implications for the development of novel therapeutics and better clinical management of endometriosis.

Vascular endothelial growth factor (VEGF) has been implicated in the regulation of vesicular transport of amniotic fluid via caveolae across the amnion. This study tested the hypothesis that VEGF regulates caveolar function by stimulating caveolin-1 expression and phosphorylation in ovine amniotic epithelial cells (oAECs). Using primary cultures of oAECs, caveolin-1 was identified by immunofluorescent staining. Caveolin-1 messenger RNA (mRNA) abundance was determined by Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and protein by Western blotting. The effects of VEGF ₁₆₅ on caveolin-1 expression and phosphorylation were determined. Caveolin-1 immunoreactivity was detected in oAECs. In response to 10 ng/mL VEGF ₁₆₅, caveolin-1 mRNA levels increased whereas the protein levels were unaffected. Furthermore, VEGF stimulated caveolin-1 phosphorylation, an effect abrogated by the inhibition of c-Src protein kinase. These data suggest that VEGF upregulates caveolin-1 activity through c-Src signaling pathways. Our observations support the hypothesis that VEGF regulates amniotic fluid transport across the amnion by stimulating caveolin-1 activity to mediate caveolar function in amnion cells.

The current study investigates tissue-specific prostaglandin secretion and cyclooxygenase 2 (COX-2) induction in full-thickness human gestational membranes. Gestational membranes were collected from healthy, nonlaboring cesarean deliveries at 37 to 39 weeks gestation and cultured in 2-chamber Transwell devices. Lipopolysaccharide exposure (100 ng/mL for 8 hours) elevated prostaglandin E₂ and F_2α concentrations in the amniotic chamber medium regardless of whether exposure was to the amniotic, decidual, or both sides of the membranes. However, prostaglandin E₂ and F_{2 α} concentrations in the decidual chamber medium were elevated compared with controls only if the decidual side was exposed directly to lipopolysaccharide. Whereas prostaglandin F_2α concentrations increased to similar extents in the amniotic and decidual chambers regardless of lipopolysaccharide exposure modality, prostaglandin E₂ concentrations were 22-fold higher on the amniotic side than the decidual side after lipopolysaccharide stimulation of the amnion. These findings demonstrate the propagation of prostaglandins, prostaglandin precursors, or other factors in the direction of the decidua to the amnion, but the reverse situation was not evident. Immunostaining for COX-2 was related to the side of lipopolysaccharide exposure, that is, exposure to the amnion caused immunostaining in cells of the collagen layers of the amnion and chorion, whereas exposure to the decidual side caused staining in decidual cells. These findings suggest that the inflammatory effect of lipopolysaccharide on COX-2 induction occurs within a localized area of exposure and that prostaglandins or their precursors move across the tissues of the gestational membranes by currently undefined transport mechanisms.

This study examined possible relationships between homocysteine and markers used in first-trimester screening for Down syndrome. Pregnancies were categorized into 4 groups according to quartile ranking of maternal plasma homocysteine concentration. Of the 595 pregnancies, 147 were assigned to group 1 (homocysteine level 0.6-3.5 μmol/L), 156 to group 2 (homocysteine level 3.6-4.5 μmol/L), 142 to group 3 (homocysteine level 4.6-5.6 μmol/L), and 150 pregnancies to group 4 (homocysteine level 5.7-12.6 μmol/L). No significant difference in mean nuchal translucency and mean free β-human chorionic gonadotropin (free-βhCG) multiples of the median (MoM) levels were observed. However, the mean pregnancy-associated plasma protein A (PAPP-A) MoM levels were significantly decreased in inverse relationship with homocysteine level among all 4 groups (F = 31.127, P < .001). If homocysteine is assayed as part of the first-trimester maternal serum testing, it is important to adjust for homocysteine concentration when using PAPP-A serum level for calculating the risk of fetal aneuploidy.

The aim of this study was to develop a dynamic culture medium containing FSH, LH and EGF to promote the in vitro development of oocytes obtained from goat preantral follicles to complete maturation and to improve the capacity of these oocytes for in vitro fertilization (IVF) and embryo production. For experiment I, preantral follicles were cultured for 18 days in medium supplemented with increasing concentrations of FSH (T1 - control) or in control medium added LH alone or in association with EGF: T2 (LH 50 ng/ml), T3 (LH 50 ng/ml + EGF 50 ng/ml), T4 (LH 50 ng/ml + EGF 100 ng/ml), T5 (LH 100 ng/ml), T6 (LH 100 ng/ml + EGF 50 ng/ml) and T7 (LH 100 ng/ml + EGF 100 ng/ml). For experiment II, preantral follicles were cultured only in the culture medium used in T7, and after 18 days, their oocytes underwent in vitro maturation (IVM) followed by IVF. At the end of the culture period, T3, T4 and T7 had a positive influence on the daily follicular growth rate. Oocytes grown in T4 and T7 had a meiosis resumption percentage significantly superior to the other treatments. Two embryos were obtained, in which preantral follicles in medium supplemented with 100 ng/ml LH and 100 ng/ml EGF (T7). In conclusion, our sequential culture system was able to promote the in vitro growth of preantral follicles, promoting their oocyte maturation and caprine embryo production from preantral follicles.

Objective: To evaluate the feasibility for confirming the preservation of the parasymphathetic nerve pathway innervating the bladder during nerve-sparing radical hysterectomy (RH). Methods: A total of 20 patients underwent nerve-sparing RH. Intraoperative electrical stimulation (IES) were performed on the root of pelvic splanchnic nerve (PSN) trunk while recording the electromyographic (EMG) activity of the vesical detrusor. The average duration achieving residual urine ≤50 mL and urodynamic study (UDS) was observed. Results: Evoked potentials were recorded when stimulating, in 18 patients who were referred IES-positive. Its duration was 9.89 days. The UDS results indicated that all voided normally. The remaining 2 IES-negative cases with no evoked potentials had longer duration and the micturitions were performed using abdominal pressure. Conclusion: During nerve-sparing RH, IES based on the measurement of EMG activity is a useful tool for confirmation of the preservation of parasymphathetic nerve pathway innervating the bladder and prediction of the postoperative bladder function.