Mediation effect of pregnancy-induced hypertension on the association between assisted reproductive technology and adverse neonatal outcomes: a population-based study.

BACKGROUND: Assisted reproductive technology (ART) has been widely used in the treatment of infertility, and is associated with adverse maternal and neonatal outcomes. However, the potential pathways by which ART affects adverse neonatal outcomes are unclear. We aimed to investigate the role of pregnancy-induced hypertension (PIH) in the association between ART and adverse neonatal outcomes. METHODS: Adult women (aged ≥ 18 years) with a singleton pregnancy in the National Vital Statistics System (NVSS) 2020 were enrolled in this retrospective cohort study. Study outcomes were adverse neonatal outcomes, including premature birth, low birth weight, and admission to the neonatal intensive care unit (NICU). Logistic regression models were utilized to investigate the association between ART, PIH, and adverse neonatal outcomes, expressed as odds ratio (OR) and 95% confidence interval (CI). The distribution-of-the-product method was used to explore whether there was a mediating effect of PIH between ART and adverse neonatal outcomes, and the 95% CI of the distribution-of-the-product did not contain 0 indicating a mediating effect. RESULTS: This study included 2,824,418 women, of whom 35,020 (1.24%) women used ART, 239,588 (8.48%) women had PIH, and 424,741 (15.04%) neonates had any adverse neonatal outcomes. The use of ART was associated with higher odds of PIH (OR = 1.42; 95%CI: 1.37-1.46) and any adverse neonatal outcomes (OR = 1.47; 95%CI: 1.43-1.51). The distribution-of-the-product was 0.31 (95%CI: 0.28-0.34), and 8.51% of the association between ART and adverse neonatal outcomes was mediated through PIH. Among different adverse neonatal outcomes, PIH mediated 29.17% of the association between ART and low birth weight, 9.37% of the association between ART and premature birth, and 12.20% of the association between ART and NICU admission. The mediating effect of PIH was found in women of different ages (< 35 years and ≥ 35 years) and parities (primipara and multipara). CONCLUSION: This study supports a mediating role for PIH in the association between ART and adverse neonatal outcomes. Further studies are needed to determine the mechanisms by which AR affects PIH so that interventions to reduce PIH can be developed to reduce adverse neonatal outcomes associated with ART.

Identification and validation of immune and prognosis-related genes in hepatocellular carcinoma: A review.

PURPOSE: Bioinformatics methods were used to identify the key genes associated with the immune microenvironment of hepatocellular carcinoma (HCC) to construct an immune risk prognostic model (IRPM) and to study the correlation between IRPM's risk groups and immune characteristics of patients with HCC. METHODS: HCC transcriptome sequencing information was searched for immune-related genes (IRGs) that were regularly expressed in cancer tissues. The IRGs, which were strongly linked to overall survival were screened; the prognostic characteristics model was constructed using Cox regression analysis. IRPM's independent prognostic value was explored; Kaplan-Meier survival and receiver-operating characteristic curves were used to determine the model prediction ability in the led-to queue. RESULTS: Patients in the high-risk group (HRG) showed significantly poor outcomes. Gene Set Enrichment Analysis revealed factors involved in both the HRG and low risk group. Immune-related hub genes (IRHGs) and drug sensitivity expression levels revealed that all IRHGs were correlated with drug sensitivity for certain chemotherapy drugs. CONCLUSION: The study results may serve as a reference for improving prognosis, early screening, and immunotherapy in patients with HCC.

Electroacupuncture Delays Cartilage Degeneration by Modulating Nuclear Factor-κB Signaling Pathway.

OBJECTIVE: To illustrate the molecular mechanisms underlying the therapeutic effects of electroacupuncture (EA) on knee osteoarthritis (OA). METHODS: Twenty-seven six-month-old New Zealand white rabbits were allocated into three groups in accordance with a random number table: normal group (no surgery-induced OA; without treatment), model group (surgery-induced OA; without treatment) and EA group [surgery-induced OA; received treatment with EA at acupoints Dubi (ST 35) and Neixiyan (EX-LE 5), 30 min twice a day]. After eight consecutive weeks of treatment, the histopathological alterations in cartilage were observed using optical microscopy and transmission electron microscopy, cartilage degeneration was evaluated by modified Mankin's score principles, the synovial fluid concentration of interleukin-1ß (IL-1ß), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and matrix metalloproteinase-3 (MMP-3) were evaluated by enzyme-linked immunosorbent assay, and the protein expression levels of IL-1ß, IL-6, TNF-α, MMP-3, IκB kinase-ß (IKK-ß), nuclear factor of α light polypeptide gene enhancer in B-cells inhibitor α (IκB-α) and nuclear factor-κB (NF-κB) p65 were quantified by Western blot analysis. RESULTS: EA treatment significantly improved cartilage structure arrangement and reduced cellular degeneration. The IL-1ß, IL-6, TNF-α and MMP-3 of synovial fluid in the EA-treated group were significantly decreased compared with the model group (all P<0.01). Compared with the model group, the IL-1ß, IL-6, TNF-α, MMP-3, IKK-ß and NF-κB p65 protein expressions in cartilage of EA-treated group were significantly decreased (all P<0.01), whereas IκB-α expression was significantly up-regulated (P<0.01). CONCLUSION: EA treatment may delay cartilage degeneration by down-regulating inflammatory factors through NF-κB signaling pathway, which may, in part, explain its clinical efficacy in the treatment of knee OA.

Beneficial effects of Heqi san on rat model of polycystic ovary syndrome through the PI3K/AKT pathway.

BACKGROUND: Heqi San, a traditional Chinese medicine (TCM) has been reported to regulate hormone levels in patients with metabolic disease, suggesting a potential clinical application. In the current study, we aimed to elucidate the effect of Heqi San on rat model of polycystic ovary syndrome (PCOS). METHOD: PCOS model was established in female SD rats. Rats were randomly divided into four groups: the control, untreated PCOS model, Heqi San treated PCOS model (8.1 g/kg) and metformin (MET) treated PCOS model (135 mg/kg) groups. All animals were subcutaneously injected with 6 mg/100 g dehydroepiandrosterone (DHEA) in the neck once a day for 20 consecutive days. The serum hormone levels were measured by ELISA. The ovarian tissues were stained with hematoxylin and eosin (HE) to undergo pathological examination. The expression levels of GLTU4 and PTEN mRNA were examined by real time PCR. The crucial proteins in the PI3K/APT pathway were analyzed by western blotting. Then, the functions of the target genes were analyzed using bioinformatics approaches. RESULTS: We found that Heqi San was able to recover the serum hormone levels and improve insulin resistance in PCOS rat model. A morphological lesion of the ovary was also restored with the Heqi San treatment. More importantly, we discovered a correlation between the PI3K/AKT signaling pathway and the beneficial effects of Heqi San, demonstrating that its application could alter the expression levels of p-ERK, p-AKT, p-GSK3ß, IRS-1, PTEN and GLTU4, all key factors in the PI3K/APT pathway. Through a bioinformatical analysis, we predicted the related gene function and pathway of the pathological mechanism of PCOS and found miRNAs that are likely to be critical in PCOS occurrence, including rno-miR-144-3p, rno-miR-30c-2-3p, rno-miR-486, rno-miR-3586-3p and rno-miR-146b-5p. CONCLUSION: The beneficial effects of Heqi on PCOS, including alter serum hormone levels, recover ovary morphological lesions and improve insulin resistance, which is mediated through the PI3K/AKT pathway. The potential role of miRNA-144-3p in PCOS pathogenesis.

The Changes of Cytoskeletal Proteins Induced by the Fast Effect of Estrogen in Mouse Blastocysts and Its Roles in Implantation.

It is necessary for estrogen to activate mouse blastocysts, so that they can attach to endometrial epithelium in implantation and in our previous research, we have proved estrogen can induce a fast increase in intracellular calcium of mouse blastocysts through acting on G protein-coupled receptor 30 (GPR30), which further promotes their implantation. Moreover, there has been evidence that cytoskeletal proteins are involved in integrin-mediated adhesion of many kinds of cells, which also plays an important role in implantation. To prove estrogen induces rapidly the changes of cytoskeletal proteins in mouse blastocysts and its roles in implantation, we first used immunofluorescence staining and laser confocal microscopy to investigate the fast effect of estrogen on the expression and localization of cytoskeletal proteins in mouse blastocysts. Second, we used electroporation associated with RNA interference to knock down one of the important cytoskeletal proteins, talin, in the mouse blastocyst cells to investigate the fast effect of estrogen on the localization of integrins and the binding activity of integrins with their ligand fibronectin (FN). At last, mouse blastocysts with different treatments were cultured with FN or uterine epithelial cell line Ishikawa in vitro, respectively, and transferred into the bilateral uterine horns of recipient mice, to study the role of the fast effect of estrogen on cytoskeletal proteins in blastocysts adhesion and implantation. Our results indicated that estradiol (E2), E2 conjugated with bovine serum album (E2-BSA) and G-1 (a GPR30-specific agonist) could induce cytoskeletal protein talin, vinculin, and actin to cluster in the mouse blastocysts, while G15 (a GPR30-specific antagonist) and BAPTA (a calcium chelator) may block this effect induced by E2-BSA. Furthermore, E2-BSA could induce the clustering and relocalization of integrin ß1 and ß3 and increase the FN-binding activity of integrins in blastocyst cells, while E2-BSA could not induce these effects in the blastocysts pretreated with talin-small interfering RNA (siRNA). Meanwhile, the adhesion rate and implantation rate of blastocysts pretreated with talin-siRNA were significantly lower than those pretreated with control-siRNA. We provided the first evidence that the fast effect of estrogen might cause the clustering of the cytoskeletal proteins in mouse blastocyst cells and further induce the changes of localization and functional activity of integrins in the blastocyst cells, which play important roles in blastocyst implantation.

[Progress of non-genomic action of estrogen and its impact on female reproduction].

Estrogen is one of the steroid hormones. Besides the genomic action mediated by its intracellular receptor on target cells, there is now increasing body of evidence indicating that estrogen also has non-genomic action. For the non-genomic action, estrogen binds to its receptor on cell membrane, subsequently rapidly activates various intracellular signaling pathways, such as PLC/Ca(2+), ERK/MAPK, cAMP-PKA, PI3K-AKT-NOS, and finally induces biological effects. The non-genomic effects of estrogen on physiologic and pathologic processes have been found in many tissues within the reproductive, nervous and cardiovascular systems and bone etc. In reproductive system, it has been demonstrated that estrogen plays important roles in follicle development, fertilization and embryo implantation, and it is involved in the genesis and development of genital tract tumors and breast cancer. In this review, we focus on the general characteristics of non-genomic action of estrogen, its main nonnuclear signaling pathways and physiological and pathological significance, especially its influences in female reproductive functions.

Relocalisation and activation of integrins induced rapidly by oestrogen via G-protein-coupled receptor 30 in mouse blastocysts.

Integrins are the dominant and final adhesion molecules in the attachment process between the blastocysts and endometrium. It is necessary for oestrogen to rapidly activate mouse blastocysts so that they attach to the endometrial epithelium. Our previous study suggested that oestrogen can rapidly induce an increase in intracellular calcium in mouse blastocysts via G-protein-coupled receptor 30 (GPR30). Thus, we deduced that integrins may be involved in GPR30 mediation of the fast effect of oestrogen on mouse blastocysts in implantation. To prove our hypothesis, we used immunofluorescence staining and in vitro coculture of mouse blastocysts and endometrial epithelial cell line (EECs), Ishikawa cells, in the present study. We found that αv and ß1 integrin clustered in mouse blastocysts, and that ß3 integrin was relocalised to the apical membrane of blastocyst cells when embryos were treated with 1 µM 17ß-estradiol (E2), 1 µM E2 conjugated to bovine serum albumin (E2-BSA) and 1 µM G-1, a specific GPR30 agonist, for 30 min respectively, whereas pretreatment with 1 µM G15, a specific GPR30 antagonist, and 5 µM 1,2-Bis(2-aminophenoxy)ethane-N,N,N'',N''-tetraacetic acid tetrakis (acetoxymethyl ester)(BAPTA/AM), a cellular Ca2+ chelator, blocked the localisation of integrins induced by oestrogen via GPR30 in mouse blastocyst cells. E2, E2-BSA and G-1 increased the fibronectin (FN)-binding activity of integrins in blastocysts, whereas G15 and BAPTA/AM blocked the activation of integrins induced by oestrogen via GPR30 in mouse blastocysts. Inhibition of integrins by Arg-Gly-Asp peptide in blastocysts resulted in their failure to adhere to EECs in vitro, even if oestrogen or G-1 was provided. Together, the results indicate the fast effect of oestrogen via the GPR30 membrane receptor further induces relocalisation and activation of integrins in mouse blastocysts, which play important roles in the adhesion of blastocysts to EECs.

[Estrogen induced rapid increase of intracellular calcium plays important role in the implantation of mouse blastocysts].

OBJECTIVE: To study the roles of the increased intracellular calcium induced rapidly by estrogen in the implantation of mouse blastocysts. METHODS: The mouse blastocysts were collected from the female mice on the pregnant day 4, divided into 3 groups: control, E2-BSA and BAPTA +E2-BSA. Immunofluorescence staining, confocal microscopy, embryo and endometrial epithenial cells co-culture and embryo transfer were used to investigate the effect of increased intracellular calcium induced by E2-BSA on the expression and localization of integrins in blastocysts and their adhesion to endometrial epithenial calls (EECs) and implantation into the endometrium. RESULTS: The increase of intracellular calcium induced rapidly by estrogen could cause the cluster and relocation of integrin av and beta3, and BAPTA might block this effect, the adhesion rate of blastocysts in contol group was 35.5%, BAPTA +E2-BSA group was 26.7% and significantly lower than 65.6% of E2-BSA group (P<0.05), and the implantation rate in BAPTA+E2-BSA group was 11.8%, which was significantly lower than 52.9% of E2-BSA group (P<0.05). CONCLUSION: The rapid increase of intracellular calcium induced by estrogen may cause the relocalization of integrin in blastocysts and their adhesion to ECCs, which is important in the process of implantation.

GPR30 Mediates the Fast Effect of Estrogen on Mouse Blastocyst and its Role in Implantation.

Our previous work demonstrated that estrogen could rapidly increase intracellular Ca(2+) in dormant mouse blastocysts. The purpose of the present study is to investigate the physiological relevance of G protein-coupled receptor 30 (GPR30) in the fast effect of estrogen on mouse blastocyst and in embryo implantation. We used reverse transcription-polymerase chain reaction, immunofluorescence, embryo coculture with Ishikawa uterine epithelial cell line, and embryo transfer technology to detect the expression of GPR30 in mouse embryos and the nongenomic effects of estrogen via GPR30 on blastocyst. We found that GPR30 is expressed in the mouse blastocyst, and its location is mostly consistent with the binding site of estrogen. Both estrogen and GPR30-specific agonist G-1 rapidly increase the intracellular Ca(2+) and phospholipase C activation in blastocyst cells, while GPR30-specific antagonist G-15 blocked this effect of estrogen. The pretreatment of G-15 on blastocysts lead to a lower attachment rate and implantation rate. Our data collectively suggested that GPR30 can mediate the fast effect of estrogen on blastocysts and play an important role in embryo implantation.

Identification and characterization of progesterone- and estrogen-regulated MicroRNAs in mouse endometrial epithelial cells.

In endometrial epithelial cells, progesterone (P4) functions in regulating the cell structure and opposing the effects of estrogen. However, the mechanisms of P4 that oppose the effects of estrogen remain unclear. MicroRNAs (miRNAs) are important posttranscriptional regulators that are involved in various physiological and pathological processes. Whether P4 directly induces miRNA expression to antagonize estrogen in endometrial epithelium is unclear. In this study, total RNAs were extracted from endometrial epithelium of ovariectomized mice, which were treated with estrogen alone or a combination of estrogen and P4. MicroRNA high-throughput sequencing with bioinformatics analysis was used to identify P4-induced miRNAs, predict their potential target genes, and analyze their possible biological functions. We observed that 146 mature miRNAs in endometrial epithelial cells were significantly upregulated by P4. These miRNAs were extensively involved in multiple biological processes. The miRNA-145a demonstrated a possible function in the antiproliferative action of P4 on endometrial epithelial cells.