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- Author: Hui Li x
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Department of Biology, Southern University of Science and Technology, Shenzhen, Guangdong, China
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Somatic cell nuclear transfer (SCNT) has been successfully used for cloning in a variety of mammalian species. However, SCNT reprogramming efficiency is relatively low, in part, due to incomplete DNA methylation reprogramming of donor cell nuclei. We previously showed that ten-eleven translocation 3 (TET3) is responsible for active DNA demethylation during preimplantation embryonic development in bovines. In this study, we constructed TET3-overexpressing cell lines in vitro and observed that the use of these fibroblasts as donor cells increased the blastocyst rate by approximately 18 percentage points compared to SCNT. The overexpression of TET3 in bovine SCNT embryos caused a decrease in the global DNA methylation level of the pluripotency genes Nanog and Oct-4, ultimately resulting in an increase in the transcriptional activity of these pluripotency genes. Moreover, the quality of bovine TET3-NT embryos at the blastocyst stage was significantly improved, and bovine TET3-NT blastocysts possessed more total number of cells and fewer apoptotic cells than the SCNT blastocysts, similar to in vitro fertilization (IVF) embryos. Nevertheless, DNA methylation of the imprinting control region (ICR) for the imprinted genes H19-IGF2 in SCNT embryos remained unaffected by TET3 overexpression, maintaining parent-specific activity for further development. Thus, the results of our study provide a promising approach to rectify incomplete epigenetic reprogramming and achieve higher cloning efficiency.
Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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Department of Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
Shanghai Key Laboratory of Female Reproductive Endocrine Related Diseases, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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Immune cells and cytokines have important roles in the pathogenesis of endometriosis. However, the production and role of cytokines of T helper type 1 (Th1) and Th2 cells in the progress of endometriosis have remained to be fully elucidated. The present study reported that the interferon (IFN)-γ levels and the percentage of IFN-γ+CD4+ cells were significantly increased in the peritoneal fluid (PF) at the early stage and maintained at a higher level at the advanced stage of endometriosis; furthermore, interleukin (IL)-10 and IL-10+CD4+ cells were elevated in the advanced stage of endometriosis. In addition, IL-2 levels in the PF at the advanced stage of endometriosis were elevated and negatively associated with IFN-γ expression. In a co-culture system of ectopic endometrial stromal cells (ESCs) and macrophages, elevated IL-2 was observed, and treatment with cytokines IL-2 and transforming growth factor-β led to upregulation of the ratio of IL-2+ macrophages. IL-27-overexpressing ESCs and macrophages were able to induce a higher ratio of IL-10+CD4+ T cells. Blocking of IL-2 with anti-IL-2 neutralizing antibody led to upregulation of the ratio of IFN-γ+CD4+ T cells in the co-culture system in vitro. Recombinant human IL-10 and IFN-γ promoted the viability, invasiveness and transcription levels of matrix metalloproteinase (MMP)2, MMP9, and prostaglandin-endoperoxide synthase 2 of ESCs, particularly combined treatment with IL-10 and IFN-γ. These results suggest that IL-2 and IL-27 synergistically promote the growth and invasion of ESCs by modulating the balance of IFN-γ and IL-10 and contribute to the progress of endometriosis.
The International Peace Maternity & Child Health Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
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The International Peace Maternity & Child Health Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
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The International Peace Maternity & Child Health Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
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The International Peace Maternity & Child Health Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
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The International Peace Maternity & Child Health Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
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The International Peace Maternity & Child Health Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
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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.
