Reproductive Sciences

The early discontinuation of the Women's Health Initiative trials of daily conjugated estrogens and medroxyprogesterone and of conjugated estrogens only was hailed as the “death to the use of hormone replacement regimens” in menopause. The analyses showed risks outweighing benefits of hormone therapy when given broadly to postmenopausal women. The expanding basic science and clinical research on the specific actions of sex steroids at the genomic and nongenomic level, however, shed new insight into these results. This review focuses on the vascular and metabolic effects of sex steroids to illustrate new advances. Understanding the mechanisms of sex steroid receptor action in a tissue-specific manner, ligand-specific dose responses, and the effects of steroid hormones in normal compared to diseased tissues may explain some of the outcomes in the clinical trials. Further research will clarify the potential benefits and risks of hormone therapy after menopause, both in individual patients and in selected populations.

The objective of this study is to test whether the activation of toll-like receptors (TLRs) 2 and 3 (innate immune receptors for gram-positive and viral pathogens, respectively) can induce preterm delivery. One uterine horn of preterm pregnant CD-1 mice at approximately 75% of gestation was injected with TLR-2 ligands (lipoteichoic acid [LTA] or peptidoglycan [PGN]) or the TLR-3 ligand polyinosinic:cytidylic acid (poly[I:C]). Preterm delivery was recorded. In a separate group of mice, tissue mRNAs were quantified by reverse transcriptase polymerase chain reaction 5 hours after treatment with PGN or poly(I:C). Intrauterine PGN and LTA induced preterm delivery, reaching 100% at maximal doses. Intraperitoneal PGN also induced preterm delivery but at lower rates (maximum = 55%). Intrauterine poly(I:C) induced preterm birth in up to 31% of mice. Poly(I:C) induced uterine interferon β and chemokine (C-C motif) ligand 5 (CCL5, also known as RANTES) but not interleukin 1β, tumor necrosis factor, or lipopolysaccharide-induced CXC chemokine. PGN did not alter these mRNAs when compared with saline. Neither treatment induced gene expression in fetal membranes. Activation of either TLR-2 or -3 can induce preterm delivery in the mouse. Activation of TLR-3 with poly(I:C) induces interferon β and the chemokine CCL5 in uterine tissues but not in fetal membranes.

Multidrug resistance phosphoglycoprotein (ABCB1) has been shown to limit maternal-fetal transfer by actively excluding ABCB1 substrates. The authors have previously demonstrated a marked decrease in placental ABCB1 expression in the human and mouse with advancing gestation. In the present study, it is hypothesized that the decrease in ABCB1 expression will result in increased transplacental transfer of ABCB1 substrates over the second half of gestation and that progesterone exhibits a regulatory role on placental ABCB1 expression and function. The authors demonstrate a significant increase in transplacental transfer of [³H]digoxin (an ABCB1 substrate) in late gestation (E18.5; P < .001) when compared to earlier embryonic days. Furthermore, maternal plasma progesterone levels did not influence expression or function of ABCB1. The authors conclude that the fetus is increasingly exposed to both endogenous and exogenous substrates of ABCB1 present in the maternal circulation with advancing gestation and that progesterone does not elicit a regulatory role on placental ABCB1 expression or function in vivo.

Maternal food restriction during pregnancy results in intrauterine growth-restricted (IUGR) newborns with significantly decreased plasma leptin levels. When nursed by ad libitum—fed controls, IUGR offspring exhibit hyperphagia with adult obesity, marked by increased percentage body fat and plasma leptin, suggesting altered anorexigenic pathways. The authors examined leptin signaling pathways and food intake responses to 2 putative anorexic effectors (leptin and sibutramine, a serotonin reuptake inhibitor) in IUGR offspring. From 10 days to term gestation and through lactation, control pregnant rats received ad libitum food, whereas study rats were 50% food restricted. Following birth, litter size was standardized, and all offspring were nursed by control dams. At 3 weeks of age, offspring were weaned to ad libitum laboratory chow. At ages 1 day and 3 weeks, hypothalamic leptin receptor (Ob-Rb) mRNA and total STAT3 protein expression were determined. In addition, phosphorylated STAT3 was measured in 1-day-old offspring administered peripheral leptin. In prepubescent and adult offspring, anorexic effects of leptin and sibutramine were determined. At 1 day of age, IUGR pups showed increased hypothalamic Ob-Rb mRNA and total STAT3 protein expression though reduced leptin activated phosphorylated STAT3. At 3 weeks of age, IUGR offspring had decreased hypothalamic Ob-Rb mRNA expression, although with continued elevated STAT3 protein levels. The IUGR offspring demonstrated resistance to anorexigenic agents, leptin (6 weeks and 6 months), and sibutramine (8 months), as evidenced by less reduction in food intake and less body weight loss than controls. The IUGR offspring demonstrate suppressed leptin-induced STAT3 phosphorylation and impaired anorexigenic response to 2 factors in the central satiety pathway. This reduced anorexigenic function, together with normal or perhaps enhanced orexigenic function, contributes to the development of programmed obesity in IUGR rat offspring.

Most of cervical intraepithelial neoplasia 1 (CIN 1) will regress and 12% to 40% of high-grade CIN may progress to squamous cell carcinoma (SCC) of the uterine cervix. However, the differentiation of CIN 1 and high-grade CIN is sometimes controversial among pathologists. Human telomerase reverse transcriptase (hTERT) is therefore applied to detect the differences among normal, CIN 1, high-grade CIN, and SCC tissues of uterine cervix. One hundred six cervical specimens were collected for immunohistochemical study of hTERT. These data were compared with the human papillomavirus (HPV) DNA status. Expression of hTERT in high-grade CIN increased significantly compared to that in CIN 1 ( P < .001). A positive relationship was found between high hTERT expression and degree of malignant transformation ( P < .001). Most of the cases with high hTERT expression tested positive for the high-risk HPV groups. High hTERT expression was detected in 88.73% of the samples with cervical high-grade CIN or SCC. Low hTERT expression was found in 94.29% of low-grade CIN or normal tissues. Furthermore, 96.92% of the cervical tissues with high hTERT expression were high-grade CIN or SCC. A total of 80.49% of samples with low hTERT expression were low-grade CIN or normal tissues. A significantly increased hTERT expression between CIN 1 and high-grade CIN exhibits a critical progression in cervical carcinogenesis. hTERT can be offered as additional molecular information correlated with more severe dysplasia and SCC. Furthermore, this increased hTERT expression is correlated whigh-risk HPVs infection.

Endometrial cancer is associated with both EGFR and HER2 receptor activation. The EGFR and HER2 genes could be disease susceptibility candidate genes for this cancer. This study was conducted to investigate a possible association between EGFR and HER2 gene polymorphisms and endometrial cancer and the influence of these polymorphisms on the clinical outcome of endometrial cancer patients in a Japanese population. The authors compare the genotype distributions and allele frequencies of the EGFR +2073 A/T and HER2 +655 A/G polymorphisms in 116 endometrial cancer patients and 213 controls using polymerase chain reaction—restriction fragment length polymorphism (RFLP) analysis. RFLP results were confirmed by direct DNA sequencing. Of the 116 patients, 76 (65.5%) could be followed up. Disease-free survival estimates were computed using the Kaplan-Meier method, and differences between survival periods were assessed using the log-rank test. No significant differences were observed in either genotype distributions or allele frequencies in the EGFR +2073 A/T and HER2 +655 A/G polymorphisms between endometrial cancer patients and controls. The stratification by histological types and staging failed to identify significant differences between endometrial cancer patients and controls. No statistical differences were noted between these polymorphisms and disease-free survival (Kaplan-Meier log-rank test P = .55 and .66, for the EGFR +2073 A/T and HER2 +655 A/G, respectively). These results suggest that the EGFR +2073 A/T and HER2 +655 A/G polymorphisms are not associated with endometrial cancer in a Japanese population. These conclusions are based on relatively small numbers and will require verification from additional independent studies.

Extracellular matrix plays a key role in cell growth, survival, and differentiation in a wide array of tissue types through integrin-mediated signaling pathways and its interaction with growth factors. This study investigates the role of extracellular matrix and its interaction with activin-A on in vitro growth and survival of mouse preantral follicles. Preantral follicles isolated from 14-day-old immature mouse ovaries were cultured either 3 dimensionally using basement membrane matrix (growth factor — reduced matrigel) or 2 dimensionally on cover slips coated with a single component of extracellular matrix (fibronectin, collagen, or laminin), on polylysine (negative control), or in standard culture plates in a serum-free culture medium with or without activin-A for 7 days. Follicles cultured in matrigel maintained well their 3-dimensional structure compared to those cultured conventionally. This observation was confirmed by analyzing 3-dimensional images of follicles cultured in matrigel and standard culture plate using confocal microscopy. Furthermore, follicles displayed higher growth and survival rates and exhibited antral space formation as early as day 5 of culture when activin-A was added to matrigel; in contrast, the addition of activin-A had no effect on the growth and survival of follicles cultured on individual extracellular matrix components after 7 days of culture. These data may suggest that 3-dimensional culture with extracellular matrix and activin-A provides a better milieu for in vitro growth and survival of preantral follicles in immature mice.

Deep infiltrating endometriosis is characterized by the presence of nodular lesions largely composed of fibromuscular tissue. Transforming growth factor beta 1 (TGF-β1) is the cytokine most causatively associated with disorders characterized by fibrosis throughout the body. Therefore, the hypothesis was tested that mechanisms increasing the fraction of biologically active TGF-β1, such as TGF-β 1 gene polymorphisms, lead to an increased risk of developing deep infiltrating endometriosis. The frequency of the -509C/T polymorphism of the TGF-β 1 gene was tested in women with deep infiltrating endometriosis (n = 72), gynecological patients without symptoms of endometriosis (n = 95) and healthy females (n = 93). Detection of the -509C/T polymorphisms was performed using PCR-restriction fragment length polymorphism analysis. We did not observe statistically significant differences in the frequency of the -509C/T polymorphism between the groups. Our study does not support an association between the −509C/T polymorphism of the TGF-β 1 gene and an increased risk of deep infiltrating endometriosis.

The objective of this study is to determine whether trichostatin A (TSA) can suppress the invasiveness of 2 endometriotic cell lines known to be invasive and E-cadherin negative. The membrane invasion culture system was used to assess cell invasion using invasive and a noninvasive bladder cancer cell lines as positive and negative controls, respectively. The E-cadherin mRNA levels and protein expression were evaluated by real-time reverse transcriptase polymerase chain reaction and Western blot analysis, respectively. The authors found that TSA attenuates the invasiveness of 2 cell lines in the presence or absence of tumor necrosis factor α (TNFα) stimulation. In addition, TSA treatment reactivates E-cadherin gene and protein expression in these cell lines. These results, along with recent findings that TSA suppresses proliferation, interleukin-1 β—induced cyclo-oxygenase 2 expression, and constitutive or TNFα-stimulated nuclear factor κ B activation in endometrial and endometriotic cells, makes histone deacetylase inhibitors a promising class of compounds for novel and more effective medical treatment of endometriosis, especially given the mounting evidence that endometrios be an epigenetic disease.

Activin A and follistatin are growth factors produced by several organs, comprising the endometrium, where they modulate cell and tissue differentiation. In this study, the authors tested whether activin A and follistatin are measurable in menstrual blood and whether their concentrations change in women with dysfunctional uterine bleeding (DUB). The authors evaluated healthy women with regular menstrual cycles (n = 15) and women with DUB (n = 12). Activin A and follistatin were measured in both menstrual and peripheral blood samples using highly sensitive enzyme immunoassays, whereas their respective mRNAs were quantified by real-time polymerase chain reaction in endometrial samples collected during the perimenstrual period. Activin A concentrations were 4-fold higher in menstrual than in peripheral serum of healthy women (mean ± SE, 4.24 ± 0.18 vs 1.00 ± 0.15 ng/mL, P < .001) and were significantly lower in women with DUB compared to healthy subjects (P < .001). Follistatin concentration was 8-fold higher in menstrual than in peripheral serum of healthy women (3.94 ± 0.49 vs 0.49 ± 0.04 ng/mL, P < .001) and was significantly lower in the menstrual serum of women with DUB compared to controls (P < .001). There was no correlation between menstrual and peripheral serum concentrations of both proteins. The endometrial expression of activin A and follistatin mRNA was lower in women with DUB compared to controls (P < .05). Both activin A and follistatin are measurable in high concentrations in human menstrual blood and are relatively lower in women with DUB. The quantitative assessment of activin A and follistatin in menstrual serum might be a putative clinical marker of endometrial function.