Environmental epigenetic interaction of gametes and early embryos†.

In recent years, the developmental origins of diseases have been increasingly recognized and accepted. As such, it has been suggested that most adulthood chronic diseases such as diabetes, obesity, cardiovascular disease, and even tumors may develop at a very early stage. In addition to intrauterine environmental exposure, germ cells carry an important inheritance role as the primary link between the two generations. Adverse external influences during differentiation and development can cause damage to germ cells, which may then increase the risk of chronic disease development later in life. Here, we further elucidate and clarify the concept of gamete and embryo origins of adult diseases by focusing on the environmental insults on germ cells, from differentiation to maturation and fertilization.

Aberrant SERPINE1 DNA methylation is involved in carboplatin induced epithelial-mesenchymal transition in epithelial ovarian cancer.

AIM: Resistance to platinum-based therapeutic agents is the major contributor to epithelial ovarian cancer (EOC) mortality. There is an urgent need to better understand the underlying mechanisms. Here we investigated the role of serpins in EOC chemoresistance and related mechanisms, and found that SERPINE1 played an important role in chemoresistance in A2780cp cells. MATERIALS AND METHODS: A2780cp and A2780s cells were used in our study. Microarray screening was used to identify the gene expression change under carboplatin treatment. A cell-counting kit-8 was used to detect the sensitivity of ovarian cancer cells after treatment. The expression of SERPINE1 was silenced by siRNA. The levels of SERPINE1 and epithelial-mesenchymal transition (EMT)-related proteins were confirmed by Western blot. MassArray EpiTYPER quantitative DNA methylation analysis was introduced to evaluate the methylation of the promoter of SERPINE1. RESULTS: Microarray data showed that SERPINE1 and SERPINE2 increased most dramatically under carboplatin treatment in A2780cp cells. Carboplatin treatment could significantly increase the expression of SERPINE1 and induce the EMT process, with decreased expression of E-cadherin and increased expression of Vimentin, Snail and Twist. Knockdown of SERPINE1, but not SERPINE2, in A2780cp cells could inhibit the EMT process. We also found that hypomethylation in the promoter of SERPINE1 might result in the increased expression of SERPINE1 and subsequent EMT process in A2780cp cells. CONCLUSION: Our study suggested that SERPINE1 may be a promising therapeutic target for chemoresistance.

Doxorubicin-induced epithelial-mesenchymal transition through SEMA 4A in hepatocellular carcinoma.

Semaphorins are essential for the functions in the regulation of cell migration. SEMA 4A has been proven to play a prominent role in immune function and angiogenesis. However, whether SEMA 4A is involved in HCC chemoresistance is unclear. We investigated the role of SEMA 4A in HCC chemoresistance and the underlying mechanisms. We tested the doxorubicin sensitivity of the Huh7, and Hep-G2 HCC cell lines. Immunofluorescence and Western blot were used to detect the location and expression of EMT-related protein, such as, E-cadherin, Vimentin, and SEMA4A expression. Microarray data showed that SEMA 4A and SEMA 3F increased most dramatically under DOX treatment. Kncokdown of SEMA 4A in hepatoma cells can reduce EMT process. Expectedly, depletion of SEMA 4A also reversed EMT and increased the DOX sensitivity. SEMA 4A confers doxorubicin resistance on HCC by inducing epithelial-mesenchymal transition (EMT).

Successive and cyclic oral contraceptive pill pretreatment improves IVF/ICSI outcomes of PCOS patients and ameliorates hyperandrogenism and antral follicle excess.

OBJECTIVES: To evaluate different oral contraceptive pill (OCP) pretreatment associated differential in-vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) outcomes of polycystic ovary syndrome (PCOS) patients and explore enhanced hormonal balance induced by the pretreatment. METHODS: This retrospective study included 500 PCOS women and 565 normal ovulating counterparts undergoing IVF/ICSI. The PCOS patients were divided into three groups based on the OCP pretreatment regimens: non-OCP (without OCP pretreatment), unsuccessive OCP (the period of successive pretreatment ≤2 months) and successive OCP (the period of successive pretreatment ≥3 months) groups. Comprehensive hormonal and ultra-sonographic assessments were performed before/after IVF pretreatment. Confounding factors affecting pregnancy outcomes were analyzed with logistic regression. RESULTS: PCOS patients with significant endocrine disorders had reduced implantation and pregnancy rates and increased miscarriage rate. Successive, not unsuccessive OCP pretreatment, significantly improved the implantation and pregnancy rates, and reduced the incidence of monotocous small-for-gestational age infants, which was accompanied by remarkably decreased hyperandrogenism and antral follicles. CONCLUSION: PCOS is an independent risk factor for poor IVF outcome. Successive, not unsuccessive, OCP cyclical pretreatment could improve pregnancy outcome of PCOS patients, associated with reduction of hyperandrogenism and antral follicle excess.

Preliminary proteomic analysis on the alterations in follicular fluid proteins from women undergoing natural cycles or controlled ovarian hyperstimulation.

PURPOSE: To study the differences in protein expression profiles of follicular fluid (FF) between controlled ovarian hyperstimulation (COH) and natural ovulatory cycles. METHODS: Twelve infertile women undergoing in vitro fertilization and embryo transfer (IVF-ET), with matched clinical information, were retrospectively recruited in the IVF center of our university hospital, including six undergoing COH and another six with natural cycles. FF was sampled from dominant follicles with mature oocytes. Protein expression profiles in each FF sample were analyzed respectively using two-dimensional gel electrophoresis. Differentially expressed proteins were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and validated by western blotting. Differentially expressed proteins were further analyzed using Ingenuity Pathway Analysis (IPA) software. RESULTS: Two proteins were downregulated and 11 proteins were upregulated (change ≥1.5-fold, P < 0.05) in the COH group. We identified one down-egulated and seven upregulated proteins using MALDI-TOF MS. Four differentially expressed proteins, including transferrin, complement component C3 (C3), haptoglobin and alpha-1-antitrypsin (AAT), were further validated by rate nephelometry and western blotting analyses. The IPA analysis revealed a significant network involved in the humoral immune and inflammatory responses. CONCLUSIONS: The eight differentially expressed proteins were related to immune and inflammatory responses in the ovary. Our results provide new insights into the influence of COH on follicular (spp) development and IVF outcomes.

Altered thyroid hormone profile in offspring after exposure to high estradiol environment during the first trimester of pregnancy: a cross-sectional study.

BACKGROUND: The increasing number of babies conceived by in vitro fertilization and embryo transfer (IVF-ET) shifts concern from pregnancy outcomes to long-time health of offspring. Maternal high estradiol (E2) is a major characteristic of IVF-ET and lasts throughout the first trimester of pregnancy. The fetal thyroid develops during this period and may thus be affected by exposure to the supra-physiological E2. The aim of this study is to investigate whether the high E2 maternal environment in the first trimester increases the risk of thyroid dysfunction in children born following IVF-ET. METHODS: A cross-sectional survey design was used to carry out face-to-face interviews with consecutive children attending the hospital. A total of 949 singletons born after fresh embryo transfer (ET) (n=357), frozen ET (n=212), and natural conception (NC) (n=380), aged 3 to 10 years old, were included. All children were thoroughly examined. Meanwhile, another 183 newborns, including 55 fresh ET, 48 frozen ET, and 80 NC were studied. Levels of serum T3, FT3, T4, FT4, and TSH and levels of maternal E2 at different stages of the first trimester were examined. RESULTS: The mean serum E2 levels of women undergoing fresh ET during the first trimester of pregnancy were significantly higher than those of the women undergoing frozen ET or following NC. The thyroid hormone profile, especially the levels of T4, FT4, and TSH, were significantly increased in 3- to 10-year-old children conceived by fresh ET compared to NC. The same tendency was confirmed in newborns. However, levels of T4 and TSH in the frozen ET group were nearer to that of the NC group. Furthermore, levels of T4 and FT4 in fresh ET were positively correlated with maternal serum levels of E2 during early pregnancy. CONCLUSIONS: The maternal high E2 environment in the first trimester is correlated with increased risk of thyroid dysfunction. Frozen ET could reduce risks of thyroid damage in children conceived by IVF. Further studies are needed to confirm these findings and to better determine the underlying molecular mechanisms and clinical significance. TRIAL REGISTRATION: ChicCTR-OCC-14004682 (22-05-2014).

Follicle-stimulating hormone orchestrates glucose-stimulated insulin secretion of pancreatic islets.

Follicle-stimulating hormone (FSH) is involved in mammalian reproduction via binding to FSH receptor (FSHR). However, several studies have found that FSH and FSHR play important roles in extragonadal tissue. Here, we identified the expression of FSHR in human and mouse pancreatic islet ß-cells. Blocking FSH signaling by Fshr knock-out led to impaired glucose tolerance owing to decreased insulin secretion, while high FSH levels caused insufficient insulin secretion as well. In vitro, we found that FSH orchestrated glucose-stimulated insulin secretion (GSIS) in a bell curve manner. Mechanistically, FSH primarily activates Gαs via FSHR, promoting the cAMP/protein kinase A (PKA) and calcium pathways to stimulate GSIS, whereas high FSH levels could activate Gαi to inhibit the cAMP/PKA pathway and the amplified effect on GSIS. Our results reveal the role of FSH in regulating pancreatic islet insulin secretion and provide avenues for future clinical investigation and therapeutic strategies for postmenopausal diabetes.

Small-conductance calcium-activated K(+) channels 3 (SK3) regulate blastocyst hatching by control of intracellular calcium concentration.

BACKGROUND: The present study was designed to investigate the expression of small-conductance calcium-activated K(+) channels 3 (SK3) in preimplantation embryos and to explore their role in the underlying mechanism of blastocyst hatching. METHODS: Human preimplantation embryos were donated by patients who achieved successful pregnancy with in vitro fertilization. Mouse preimplantation embryos in different stages were collected and cultured with or without siRNA cell injection. The expression of SK3 was examined by RT-PCR, quantitative real-time PCR, western blot and immunofluorescence. Functional expression of SK3 was investigated using the patch-clamp technique. [Ca(2+)]i was measured by fluorescent imaging. Embryos were cultured in vitro to investigate the effect of SK3 knockdown or apamin, an SK3 inhibitor, on blastocyst hatching and F-actin formation. RESULTS: In human blastocysts, the level of SK3 expression was significantly lower in blastocysts that failed to hatch than in blastocysts that hatched successfully. In mouse embryos, SK3 mRNA and protein were not found in zygotes, but were detected from the 2-cell stage onward, with the highest levels observed in blastocysts. SK3 was predominately located in the trophectoderm cell membrane of expanded blastocysts. SK3 knockdown in trophectoderm cells not only suppressed the SK3 current, but also reduced [Ca(2+)]i elevation and membrane potential hyperpolarization induced by thapsigargin. Although the formation of expanded blastocysts was not affected, blastocyst hatching and F-actin formation were significantly inhibited after SK3 knockdown in trophectoderm cells. CONCLUSIONS: SK3-mediated [Ca(2+)]i elevation and membrane potential hyperpolarization in trophectoderm cells are important for blastocyst hatching, and defects in SK3 expression may contribute to infertility.

Associations Between Asthma and Polycystic Ovary Syndrome: Current Perspectives.

A potential correlation between polycystic ovary syndrome (PCOS) and asthma, used to be identified as diseases originating from two independent systems, has been supported by increasing evidence. From an epidemiological perspective, mounting studies have confirmed that women suffering from PCOS exhibit increased susceptibility to asthma. Meanwhile, PCOS and asthma seem to share several mutual pathological conditions, such as metabolic disorders, hormonal fluctuation, proinflammatory state, etc. Here, we further elucidate the correlation between asthma and PCOS by focusing on the internal common pathophysiology and adverse influences on women's health. Understanding the internal connection between PCOS and asthma may shed light on developing new prevention and control strategies to fight against these conditions.

COVID-19 Lockdown Increased the Risk of Preterm Birth.

Purpose: To estimate whether the city-specific lockdown in Shanghai induced by the COVID-19 pandemic affected preterm birth rates among uninfected pregnant women in different trimesters. Methods: The population-based retrospective cohort study was conducted in the International Peace Maternity and Child Health Hospital (IPMCH) in Shanghai, China. Pregnant women without COVID-19 received perinatal healthcare during lockdown (from January 24, 2020 to March 24, 2020) and non-lockdown (from January 24, 2019 to March 24, 2019) period and giving birth to a live infant at IPMCH were enrolled. 1:1 propensity score matching and Inverse probability of treatment weighting were used to evaluate preterm birth (<37 weeks), very preterm birth (<34 weeks), preterm birth with premature rupture of membranes (PROM-PTB), spontaneous preterm birth with intact membranes (S-PTB), and medically induced preterm birth (MI-PTB) between two groups. Results: 8,270 pregnant women were in the lockdown group, and 9,815 were in the non-lockdown group. Pregnant women in second trimester during lockdown had a higher risk of PTB than those during the non-lockdown period [OR: 1.43 (CI 1.01-2.02), ARD: 1.7% (CI 0.04-3.4%), p = 0.045]. Furthermore, pregnant women in third trimester during lockdown had a higher risk of PROM-PTB than those during the non-lockdown period [OR: 1.64 (CI 1.09-2.47), ARD: 0.9% (CI 0.2-1.6%), p = 0.02]; no group differences were found related to rates of VPTB, S-PTB or MI-PTB. Conclusion: In this cohort study in China, we found that there was an increased risk in preterm birth for non-infected women in COVID-19 lockdown who were in their second trimester.

American, European, and Chinese practice guidelines or consensuses of polycystic ovary syndrome: a comparative analysis.

Polycystic ovary syndrome (PCOS) is the most common metabolic and endocrine disorder in women. However, there is no agreement concerning how to diagnose and treat PCOS worldwide. Three practice guidelines or consensuses, including consensus from the European Society of Human Reproduction and Embryology (ESHRE)/the American Society for Reproductive Medicine (ASRM) in Rotterdam, diagnosis criteria and consensus in China, and clinical practice guideline from the Endocrine Society (ES) in the United States are widely recognized. The present paper may provide some guidance for clinical practice based on a comparative analysis of the above three practice guidelines or consensuses.

Isocorydine suppresses doxorubicin-induced epithelial-mesenchymal transition via inhibition of ERK signaling pathways in hepatocellular carcinoma.

Doxorubicin (DOX) is a conventional and effective chemotherapeutic used in the treatment of hepatocellular carcinoma (HCC). However, doxorubicin administration may induce EMT, which results in the development of chemoresistance in HCC. Recent studies report that Isocorydine (ICD) selectively inhibits human cancer stem cells (CSCs), which have an important role in the development of chemoresistance. In this study, we observed that ICD co-administration enhanced DOX cytotoxicity in HCC cells, enabling the inhibition of DOX-induced epithelial-mesenchymal transition (EMT). Microarray data analysis revealed substantially decreased ERK signaling after ICD treatment. Additionally, we observed decreased IC50 for DOX upon ERK knockdown. Finally, we confirmed the enhanced efficacy of treatment with a combination of DOX and ICD in xenograft models. Collectively, the present study unveils the benefit of using DOX in combination with ICD for chemotherapy against HCC, revealing a novel potential anti-cancer strategy.

Aberrant expression and DNA methylation of lipid metabolism genes in PCOS: a new insight into its pathogenesis.

Background: Polycystic ovary syndrome (PCOS), whose etiology remains uncertain, is a highly heterogenous and genetically complex endocrine disorder. The aim of this study was to identify differentially expressed genes (DEGs) in granulosa cells (GCs) from PCOS patients and make epigenetic insights into the pathogenesis of PCOS. Results: Included in this study were 110 women with PCOS and 119 women with normal ovulatory cycles undergoing in vitro fertilization acting as the control group. RNA-seq identified 92 DEGs unique to PCOS GCs in comparison with the control group. Bioinformatic analysis indicated that synthesis of lipids and steroids was activated in PCOS GCs. 5-Methylcytosine analysis demonstrated that there was an approximate 25% reduction in global DNA methylation of GCs in PCOS women (4.44 ± 0.65%) compared with the controls (6.07 ± 0.72%; P < 0.05). Using MassArray EpiTYPER quantitative DNA methylation analysis, we also found hypomethylation of several gene promoters related to lipid and steroid synthesis, which might result in the aberrant expression of these genes. Conclusions: Our results suggest that hypomethylated genes related to the synthesis of lipid and steroid may dysregulate expression of these genes and promote synthesis of steroid hormones including androgen, which could partially explain mechanisms of hyperandrogenism in PCOS.

Transcutaneous Electrical Acupoint Stimulation Improves the Outcomes of In Vitro Fertilization: A Prospective, Randomized and Controlled Study.

OBJECTIVES: To explore whether transcutaneous electrical acupoint stimulation (TEAS) can improve the outcomes of in vitro fertilization (IVF). DESIGN: A prospective, randomized, and controlled study. SETTING: IVF center in a university hospital. PARTICIPANTS: Four hundred and eighty-one infertile patients with bilateral tubal blockage who were referred for IVF. Patients were randomized into four groups. INTERVENTION: TEAS was administered for 30min, respectively, at 24h before TVOR and two hours before ET. The acupoints included SP10 (Xuehai, bilateral), SP8 (Diji, bilateral), LR3 (Taichong, bilateral), ST36 (Zusanli, bilateral), EX-CA1 (Zigong, bilateral), RN4 (Guanyuan), PC6 (Neiguan, bilateral), and RN12 (Zhongwan). Based on different frequencies of TEAS, patients were grouped into a TEAS-2Hz group, a TEAS-100Hz group and a TEAS-2/100Hz group. Patients in the control group only received routine IVF treatment and no TEAS was applied on them. PRIMARY AND SECONDARY OUTCOME MEASURES: The number of mature oocytes, normally fertilized oocytes and good-quality embryos were used to evaluate oocyte developmental competence of the patients. Data of clinical pregnancy rate (CPR), implantation rate (IR), and live birth rate (LBR) were also obtained. The levels of neuropeptide Y (NPY), transforming growth factor alpha and granulocyte colony-stimulating factor in the follicular fluids were measured with enzyme-linked immunosorbent assay (ELISA). RESULTS: No significant differences were found between the control, TEAS-2Hz, TEAS-100Hz and TEAS-2/100Hz groups on the numbers of metaphase II oocytes, normally fertilized zygotes, early cleavage embryos or good quality embryos (P > .05). However, the CPR, IR and LBR of the TEAS-2/100Hz group were significantly higher than those of the other groups, respectively (P < .05). The NPY levels in the follicular fluids of TEAS-2/100Hz group were significantly higher than those of the other groups (P < .05). CONCLUSION: TEAS using a frequency of 2/100Hz could help to improve the IVF outcomes partly by increasing NPY levels in the follicular fluids.

Bisphenol A Exposure May Induce Hepatic Lipid Accumulation via Reprogramming the DNA Methylation Patterns of Genes Involved in Lipid Metabolism.

Accumulating evidence suggests a role of bisphenol A (BPA) in metabolic disorders. However, the underlying mechanism is still unclear. Using a mouse BPA exposure model, we investigated the effects of long-term BPA exposure on lipid metabolism and the underlying mechanisms. The male mice exposed to BPA (0.5 µg BPA /kg/day, a human relevant dose) for 10 months exhibited significant hepatic accumulation of triglycerides and cholesterol. The liver cells from the BPA-exposed mice showed significantly increased expression levels of the genes related to lipid synthesis. These liver cells showed decreased DNA methylation levels of Srebf1 and Srebf2, and increased expression levels of Srebf1 and Srebf2 that may upregulate the genes related to lipid synthesis. The expression levels of DNA methyltransferases were decreased in BPA-exposed mouse liver. Hepa1-6 cell line treated with BPA showed decreased expression levels of DNA methyltransferases and increased expression levels of genes involved in lipid synthesis. DNA methyltransferase knockdown in Hepa1-6 led to hypo-methylation and increased expression levels of genes involved in lipid synthesis. Our results suggest that long-term BPA exposure could induce hepatic lipid accumulation, which may be due to the epigenetic reprogramming of the genes involved in lipid metabolism, such as the alterations of DNA methylation patterns.

Protein extracts of Crassostrea gigas alleviate CCl4-induced hepatic fibrosis in rats by reducing the expression of CTGF, TGF-ß1 and NF-κB in liver tissues.

Hepatic fibrosis may contribute to liver carcinoma and the mortality of patients with hepatic fibrosis is gradually increasing. However, no definitive treatment has been established for hepatic fibrosis. The hepatic fibrotic process is reversible and can be controlled; therefore, the creation of novel and effective therapeutic methods to prevent or reverse the disease is required. The aim of the present study was to identify whether protein extracts from Pacific oysters (PEPO) could alleviate the hepatic fibrosis induced by CCl4 and to examine the mechanisms involved. A total of sixty rats were randomly divided into the following experimental groups: The normal control group; the hepatic fibrosis model group; the high­dose; medium­dose; and low­dose PEPO groups; and the colchicine group. The results indicated that compared with those of the model group, PEPO treatment significantly decreased the serum levels of alanine aminotransferase, aspartate aminotransferase, γ­glutamyltransferase, alkaline phosphatase, hyaluronic acid, laminin, collagen type IV and procollagen III in rats with hepatic fibrosis. The hematoxylin and eosin staining demonstrated that PEPO markedly alleviated hepatic fibrosis. The experiments using immunohistochemistry, western blotting and quantitative PCR indicated that protein and mRNA expression levels of connective tissue growth factor (CTGF), transforming growth factor ß1 (TGFß­1) and nuclear factor κB (NF­κB) in the liver tissues were significantly reduced by PEPO treatment. Therefore, it was concluded that PEPO successfully alleviated hepatic fibrosis induced by CCl4 and reversed the effects of hepatotoxicity by regulating the serum levels of enzymes and decreasing the expression levels of CTGF, TGF­ß1 and NF­κB in liver tissues. These findings may provide a novel treatment option for patients with hepatic fibrosis in the future.

The effects of diabetes on male fertility and epigenetic regulation during spermatogenesis.

The effects of diabetes mellitus include long-term damages, dysfunctions, and failures of various organs. An important complication of diabetes is the disturbance in the male reproductive system. Glucose metabolism is an important event in spermatogenesis. Moreover, glucose metabolism is also important for maintaining basic cell activity, as well as specific functions, such as motility and fertilization ability in mature sperm. Diabetic disease and experimentally induced diabetes both demonstrated that either type 1 diabetes or type 2 diabetes could have detrimental effects on male fertility, especially on sperm quality, such as sperm motility, sperm DNA integrity, and ingredients of seminal plasma. Epigenetic modifications are essential during spermatogenesis. The epigenetic regulation represents chromatin modifications including DNA methylation, histone modifications, remodeling of nucleosomes and the higher-order chromatin reorganization and noncoding RNAs. If spermatogenesis is affected during the critical developmental window, embryonic gonadal development, and germline differentiation, environmentally-induced epigenetic modifications may become permanent in the germ line epigenome and have a potential impact on subsequent generations through epigenetic transgenerational inheritance. Diabetes may influence the epigenetic modification during sperm spermatogenesis and that these epigenetic dysregulation may be inherited through the male germ line and passed onto more than one generation, which in turn may increase the risk of diabetes in offspring.

Androgens as double-edged swords: Induction and suppression of follicular development.

Androgens, which are mediated via the androgen receptor (AR), play important roles in normal follicular development and female fertility. However, just like a double-edged sword, besides the positive effects of androgen on follicular development, abnormal androgen levels, especially as in hyperandrogenism, seriously suppress normal follicular development. A crucial balance exists between the importance of androgens in follicular development and their negative effects when in excess. As the first meiotic division and epigenetic reprogramming are two critical events in oogenesis, abnormal androgen levels or deficiency in androgen/AR signaling in the ovary may affect these vital events. Oocytes have a tendency to develop genomic instability, thus resulting in an increasing incidence of unpredictable adult diseases. Although many studies have explored the effects of androgens and AR on follicular development, the conclusions are controversial and there has been no thorough review of this topic. This review focuses on the roles of androgens in the physiological process of follicular development, summarizes new insights into the roles of androgens in the arrested development of follicles, and discusses the potential risk of adult diseases originating from abnormal follicular androgen levels or androgen receptor signals, which may determine areas for future studies.

Small-conductance, calcium-activated potassium channel 3 (SK3) is a modulator of endometrial remodeling during endometrial growth.

BACKGROUND: Small-conductance, Ca(2+)-activated K(+) channel 3 (SK3) has been shown to be expressed in porcine endometrium. However, the roles of SK3 in human endometrium during the menstrual cycle and early pregnancy are unknown. OBJECTIVE: The objective of the study was to investigate the expression and function of SK3 in human endometrium and the mechanism involved. METHODS: We determined the expression of SK3 in human endometrium by RT-PCR, Western blotting, and immunofluorescence. Using electrophysiological and fluorescent imaging techniques, we investigated the effects of SK3 on the membrane potential and the concentrations of cytosolic calcium, respectively. The effects of SK3 on endometrial thickness and pregnancy outcome were also investigated. Knockdown of endometrial SK3 was used to examine the effects of SK3 on cell migration, cytoskeleton formation, and calcium concentration in the cytosol. RESULTS: SK3 channels are present in human endometrium. In vivo experimental and clinical data demonstrated that the reduced expression of SK3 was associated with a thin endometrium and unsuccessful pregnancy outcomes. Knockdown of human endometrial SK3 attenuated the rise in cytosolic calcium and membrane hyperpolarization induced by thapsigargin, a Ca(2+)-ATPase inhibitor, cell migration, and F-actin assembly. Knockdown of endometrial SK3 in mice also resulted in a thin endometrium and unsuccessful pregnancy outcome. CONCLUSIONS: These observations demonstrate that SK3 channels are expressed in human endometrial cells. Reduced SK3 expression attenuates endometrial cell migration and is associated with unsuccessful pregnancy outcomes.