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It has been shown that both prostaglandin I2 (PGI2) and PGE2 are essential for mouse implantation, whereas only PGE2 is required for hamster implantation. To date, the expression and regulation of cyclooxygenase (COX) and prostaglandin E synthase (PGES), which are responsible for PGE2 production, have not been reported in the rat. The aim of this study was to examine the expression pattern and regulation of COX-1, COX-2, membrane-associated PGES-1 (mPGES-1), mPGES-2 and cytosolic PGES (cPGES) in rat uterus during early pregnancy and pseudopregnancy, and under delayed implantation. At implantation site on day 6 of pregnancy, COX-1 immunostaining was highly visible in the luminal epithelium, and COX-2 immunostaining was clearly observed in the subluminal stroma. Both mPGES-1 mRNA and protein were only observed in the subluminal stroma surrounding the implanting blastocyst at the implantation site on day 6 of pregancy , but were not seen in the inter-implantation site on day 6 of pregnancy and on day 6 of pseudopregnancy. Our data suggest that the presence of an active blastocyst is required for mPGES-1 expression at the implantation site. When pregnant rats on day 5 were treated with nimesulide for 24 h, mPGES-1 protein expression was completely inhibited. cPGES immunostaining was clearly observed in the luminal epithelium and subluminal stromal cells immediately surrounding the implanting blastocyst on day 6 of pregnancy. mPGES-2 immunostaining was clearly seen in the luminal epithelium at the implantation site. Additionally, immunostaining for prostaglandin I synthase (PGIS) was also strongly detected at the implantation site. In conclusion, our results indicate that PGE2 and PGI2 should have a very important role in rat implantation.

Preeclampsia is a gestational hypertensive disease; however, preeclampsia remains poorly understood. Bioinformatics analysis was applied to find novel genes involved in the pathogenesis of preeclampsia and identified CLDN1 as one of the most differentially expressed genes when comparing patients with preeclampsia and healthy controls. The results of the qRT-PCR, Western blotting and immunohistochemistry experiments demonstrated that CLDN1 was significantly downregulated in the chorionic villi in samples from patients with preeclampsia. Furthermore, knockdown of CLDN1 in HTR-8/SVneo cells resulted in the inhibition of proliferation and induction of apoptosis, and overexpression of CLDN1 reversed these effects. In addition, RNA-seq assays demonstrated that the gene BIRC3 is potentially downstream of CLDN1 and is involved in the regulation of apoptosis. Knockdown of CLDN1 confirmed that the expression level of BIRC3 was obviously decreased and was associated with a significant increase in cleaved PARP. Interestingly, the apoptotic effect in CLDN1 knockdown cells was rescued after BIRC3 overexpression. Overall, these results indicate that a decrease in CLDN1 inhibits BIRC3 expression and increases cleaved PARP levels thus participating in the pathogenesis of preeclampsia.

Abnormal growth and migration of trophoblast cells is one of the main causes of spontaneous abortion. Eukaryotic translation initiation factor 5A (eIF5A) plays an important role in trophoblast cell growth and migration; however, its underlying mechanism remains largely unknown. Here, we first confirmed that eIF5A knockdown reduced human chorionic trophoblast HTR8 cells viability, proliferation, and migration. Next, we sought to systematically identify the genes regulated by eIF5A and observed changes in the transcriptome profile of eIF5A-knockdown HTR8 cells by RNA-seq analysis. Especially, we found that inhibition of eIF5A reduced both the mRNA and protein levels of methyltransferase-like protein 14 (METTL14). Furthermore, inhibition of METTL14 expression resulted in the reduction of viability, proliferation, and migration of HTR8 cells. In addition, we showed that overexpression of METTL14 rescued the effects of eIF5A knockdown in HTR8 cells. Finally, we revealed that eIF5A and METTL14 expression was decreased in spontaneous abortion samples compared to that in elective-induced abortion samples. Collectively, our study demonstrated that eIF5A plays a crucial role in HTR8 cells via modulation of METTL14 expression and may serve as a novel potential target for spontaneous abortion diagnosis and treatment.

Protein disulfide isomerase 3 (PDIA3) is a chaperone protein that modulates the folding of newly synthesized glycoproteins, has isomerase and redox activity, and has been implicated in the pathogenesis of many diseases. However, the role of PDIA3 in pregnancy-associated diseases remains largely unknown. Our present study reveals a key role for PDIA3 in the biology of placental trophoblasts from women with preeclampsia (PE). Immunohistochemistry and Western blot analysis revealed that PDIA3 expression was decreased in villous trophoblasts from women with PE compared to normotensive pregnancies. Further, using a Cell Counting Kit-8 assay, flow cytometry, and 5-ethynyl-2’-deoxyuridine (EdU) staining, we found that siRNA-mediated PDIA3 knockdown significantly promoted apoptosis and inhibited proliferation in the HTR8/SVneo cell line, while overexpression of PDIA3 reversed these effects. Furthermore, RNA sequencing and Western blot analysis demonstrated that knockdown of PDIA3 inhibited MDM2 protein expression in HTR8 cells, concurrent with marked elevation of p53 and p21 expression. Conversely, overexpression of PDIA3 had the opposite effects. Immunohistochemistry and Western blot further revealed that MDM2 protein expression was downregulated and p21 was increased in trophoblasts of women with PE compared to women with normotensive pregnancies. Our findings indicate that PDIA3 expression is decreased in the trophoblasts of women with PE, and decreased PDIA3 induces trophoblast apoptosis and represses trophoblast proliferation through regulating the MDM2/p53/p21 pathway.

The survival and development of a semi-allogeneic fetus during pregnancy require the involvement of decidual stromal cells (DSCs), a series of cytokines and immune cells. Insulin-like growth factor 1 (IGF1) is a low molecular weight peptide hormone with similar metabolic activity and structural characteristics of proinsulin, which exerts its biological effects by binding with its receptor. Emerging evidence has shown that IGF1 is expressed at the maternal–fetal interface, but its special role in establishment and maintenance of pregnancy is largely unknown. Here, we found that the expression of IGF1 in the decidua was significantly higher than that in the endometrium. Additionally, decidua from women with normal pregnancy had high levels of IGF1 compared with that from women with unexplained recurrent spontaneous miscarriage. Estrogen and progesterone led to the increase of IGF1 in DSCs through upregulating the expression of WISP2. Recombinant IGF1 or DSCs-derived IGF1 increased the survival, reduced the apoptosis of DSCs, and downregulated the cytotoxicity of decidual NK cells (dNK) through interaction with IGF1R. These data suggest that estrogen and progesterone stimulate the growth of DSCs and impair the cytotoxicity of dNK possibly by the WISP2/IGF1 signaling pathway.

Decline in successful conception decreases more rapidly after 38 years of age owing to follicular depletion and decreased oocyte quality. However, limited information is available regarding the underlying mechanism and the useful treatment. This study aimed to evaluate the effects of growth hormone supplementation on oocyte maturation in vivo in aged and young mice and to determine its effect on mitochondrial function. The influence of three different doses of recombinant human growth hormone (rhGH) (0.4, 0.8 and 1.6 mg/kg/day) for 8 weeks before ovarian stimulation was analyzed. Superovulated oocytes were released from the oviduct of 12-week-old and 40-week-old female C57BL/6J mice 14–16 h after administration of human chorionic gonadotropin. Ovarian follicle and morphological analysis and oocyte maturation parameters were then evaluated. This study is the first, to our knowledge, to report that medium- and high-dose rhGH significantly increases antral follicles in aged mice but anti-Müllerian hormone (AMH) levels. Furthermore, derived oocytes, MII-stage oocyte rate, ATP levels, mitochondrial membrane potential and frequencies of homogeneous mitochondrial distribution increased. In contrast, in both aged and young mice, the mtDNA copy numbers per oocyte were similar before rhGH administration, and upon saline administration, they did not differ significantly. We conclude that medium-dose rhGH supplementation before standard ovarian stimulation regimens improves oocyte quality in aged mice, probably by enhancing mitochondrial functionality.

The number of children born after assisted reproductive technology (ART) is accumulating rapidly, and the health problems of the children are extensively concerned. This study aims to evaluate whether ART procedures alter behaviours in male offspring. Mouse models were utilized to establish three groups of offspring conceived by natural conception (NC), in vitro fertilization and embryo transfer (IVF-ET), and frozen-thawed embryo transfer (IVF-FET), respectively. A battery of behaviour experiments for evaluating anxiety and depression levels, including the open field test (OFT), elevated plus maze (EPM) test, light/dark transition test (L/DTT), tail suspension test (TST), forced swimming test (FST), and sucrose preference test (SPT) was carried out. Aged (18 months old), but not young (3 months old), male offspring in the IVF-ET and IVF-FET groups, compared with those in the NC group, exhibited increased anxiety and depression-like behaviours. The protein expression levels of three neurotrophins in PFC or hippocampus in aged male offspring from the IVF-ET and IVF-FET groups reduced at different extent, in comparison to NC group. RNA sequencing (RNA-Seq) was performed in the hippocampus of 18 months old offspring to further explore the gene expression profile changes in the three groups. KEGG analyses revealed the coexisted pathways, such as PI3K-Akt signalling pathway, which potentially reflected the similarity and divergence in anxiety and depression between the offspring conceived by IVF-ET and IVF-FET. Our research suggested the adverse effects of advanced age on the psychological health of children born after ART should be highlighted in the future.