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Zhen-Yu Zheng State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100080, People’s Republic of China, College of Life Sciences, Northeast Agricultural University, Harbin 150030, People’s Republic of China and Department of Veterinary Pathobiology, University of Missouri-Columbia, Columbia, MO 65211, USA

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Qing-Zhang Li State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100080, People’s Republic of China, College of Life Sciences, Northeast Agricultural University, Harbin 150030, People’s Republic of China and Department of Veterinary Pathobiology, University of Missouri-Columbia, Columbia, MO 65211, USA

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Da-Yuan Chen State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100080, People’s Republic of China, College of Life Sciences, Northeast Agricultural University, Harbin 150030, People’s Republic of China and Department of Veterinary Pathobiology, University of Missouri-Columbia, Columbia, MO 65211, USA

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Heide Schatten State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100080, People’s Republic of China, College of Life Sciences, Northeast Agricultural University, Harbin 150030, People’s Republic of China and Department of Veterinary Pathobiology, University of Missouri-Columbia, Columbia, MO 65211, USA

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Qing-Yuan Sun State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100080, People’s Republic of China, College of Life Sciences, Northeast Agricultural University, Harbin 150030, People’s Republic of China and Department of Veterinary Pathobiology, University of Missouri-Columbia, Columbia, MO 65211, USA

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The protein kinase Cs (PKCs) are a family of Ser/Thr protein kinases categorized into three subfamilies: classical, novel, and atypical. The phosphorylation of PKC in germ cells is not well defined. In this study, we described the subcellular localization of phopho-PKC in the process of mouse oocyte maturation, fertilization, and early embryonic mitosis. Confocal microscopy revealed that phospho-PKC (pan) was distributed abundantly in the nucleus at the germinal vesicle stage. After germinal vesicle breakdown, phospho-PKC was localized in the vicinity of the condensed chromosomes, distributed in the whole meiotic spindle, and concentrated at the spindle poles. After metaphase I, phospho-PKC was translocated gradually to the spindle mid-zone during emission of the first polar body. After sperm penetration and electrical activation, the distribution of phospho-PKC was moved from the spindle poles to the spindle mid-zone. After the extrusion of the second polar body (PB2) phospho-PKC was localized in the area between the oocyte and the PB2. In fertilized eggs, phospho-PKC was concentrated in the pronuclei except for the nucleolus. Phospho-PKC was dispersed after pronuclear envelope breakdown, but distributed on the entire spindle at mitotic metaphase. The results suggest that PKC activation may play important roles in regulating spindle organization and stabilization, polar-body extrusion, and nuclear activity during mouse oocyte meiosis, fertilization, and early embryonic mitosis.

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Zhengkai Wei College of Life Sciences and Engineering, Foshan University, Foshan, Guangdong Province, People’s Republic of China

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Tingting Yu Jilin Provincial Key Laboratory of Animal Embryo Engineering, Institute of Zoonosis, College of Animal Sciences, Jilin University, Changchun, Jilin Province, People’s Republic of China

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Jingjing Wang Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, Changchun, Jilin Province, People’s Republic of China

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Chaoqun Wang Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, Changchun, Jilin Province, People’s Republic of China

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Xiao Liu Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, Changchun, Jilin Province, People’s Republic of China

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Zhen Han Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, Changchun, Jilin Province, People’s Republic of China

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Xu Zhang Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, Changchun, Jilin Province, People’s Republic of China

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Yong Zhang Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine, Jilin University, Changchun, Jilin Province, People’s Republic of China

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Hongsheng Ouyang Jilin Provincial Key Laboratory of Animal Embryo Engineering, Institute of Zoonosis, College of Animal Sciences, Jilin University, Changchun, Jilin Province, People’s Republic of China

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Zhengtao Yang College of Life Sciences and Engineering, Foshan University, Foshan, Guangdong Province, People’s Republic of China

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Sperm motility, fertilization and embryo implantation are several important factors in reproduction. Except healthy state of sperm and embryo themselves, successful pregnancy is closely related to the status of female reproductive tract immune system. Increased immune cells in reproductive tract often leads to low sperm motility and low chance of embryo implantation, but the mechanisms remain not well clarified. The aim of this study is to investigate the direct effects of swine polymorphonuclear neutrophils (PMNs) on sperm or embryo in vitro and then try to clarify the molecular mechanisms undergoing the phenomenon. Swine sperm-triggered neutrophil extracellular traps (NETs) were observed by scanning electron microscopy (SEM). PMNs phagocytosis of sperms was examined by transmission electron microscopy (TEM). Sperm-triggered NETs were quantitated by Pico Green®. Vital staining of the interaction between PMNs and embryo were observed by using confocal microscope. It was showed that PMNs were directly activated by sperm in the form of phagocytosis or casting NETs and that sperm-triggered-NETs formation was made up with DNA co-located with citrullinated histone 3 (citH3) and myeloperoxidase (MPO). In addition, the potential mechanism of NETs release was relevant to NADPH oxidase, ERK1/2 or p38 MAPK signaling pathways. Of great interest was that swine embryo was first found entangled in NETs in vitro, but the function and mechanism of this action in vivo fertilization still needed further investigation. In conclusion, this is the first report about swine sperm-induced NETs that entangle sperm and embryo, which might provide an entirely understanding of swine reproductive physiology and immunology.

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Xue-Min Qiu NHC Key Lab of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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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Zhen-Zhen Lai NHC Key Lab of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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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Si-Yao Ha NHC Key Lab of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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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Hui-Li Yang NHC Key Lab of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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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Li-Bing Liu Department of Gynecology, Changzhou No. 2 People’s Hospital, affiliated with Nanjing Medical University, Changzhou, Jiangsu Province, People’s Republic of China

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Yan Wang 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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Jia-Wei Shi NHC Key Lab of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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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Lu-Yu Ruan NHC Key Lab of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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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Jiang-Feng Ye Clinical Epidemiology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China

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Jiang-Nan Wu Clinical Epidemiology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China

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Qiang Fu Department of Immunology, Binzhou Medical College, Yantai, Shandong Province, People’s Republic of China

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Xiao-Fang Yi Department of 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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Kai-Kai Chang Laboratory for Reproductive Immunology, Institute of Obstetrics and Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
Department of Gynecology, Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China

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Ming-Qing Li NHC Key Lab of Reproduction Regulation (Shanghai Institute of Planned Parenthood Research), Hospital of Obstetrics and Gynecology, Fudan University, Shanghai, People’s Republic of China
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.

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Li-Ying Yan State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Jun-Cheng Huang State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Zi-Yu Zhu State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Zi-Li Lei State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Li-Hong Shi State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Chang-Long Nan State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Zhen-Jun Zhao State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Ying-Chun OuYang State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Xiang-Fen Song State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Qing-Yuan Sun State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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Da-Yuan Chen State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, 25 Beisihuanxi Road, Haidian, Beijing 100080, China, Graduate School, Chinese Academy of Sciences, Beijing 100080, China and College of Life Science, Soochow University, Suzhou 215006, China

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The assembly of microtubules and the distribution of NuMA were analyzed in rabbit oocytes and early cloned embryos. α-Tubulin was localized around the periphery of the germinal vesicle (GV). After germinal vesicle breakdown (GVBD), multi-arrayed microtubules were found tightly associated with the condensed chromosomes and assembled into spindles. After the enucleated oocyte was fused with a fibroblast, microtubules were observed around the introduced nucleus in most reconstructed embryos and formed a transient spindle 2–4 h post-fusion (hpf). A mass of microtubules surrounded the swollen pseudo-pronucleus 5 hpf and a normal spindle was formed 13 hpf in cloned embryos. NuMAwas detected in the nucleus in germinal vesicle-stage oocytes, and it was concentrated at the spindle poles in both meiotic and mitotic metaphase. In both donor cell nucleus and enucleated oocyte cytoplasm, NuMA was not detected, while NuMA reappeared in pseudo-pronucleus as reconstructed embryo development proceeded. However, no evident NuMA staining was observed in the poles of transient spindle and first mitotic spindle in nuclear transfer eggs. These results indicate that NuMA localization and its spindle pole tethering function are different during rabbit oocyte meiosis and cloned embryo mitosis.

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Xiao-yu Yang Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Hua Li Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Qing-wen Ma Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Jing-bin Yan Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Jiang-guo Zhao Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Hua-wei Li Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Hai-qing Shen Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Hai-feng Liu Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Ying Huang Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Shu-Zhen Huang Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Yi-Tao Zeng Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Fanyi Zeng Shanghai Institute of Medical Genetics, Shanghai Children’s Hospital, Shanghai Jiao Tong University, 24/1400 West Beijing Road, Shanghai 200040, People’s Republic of China

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Somatic cell nuclear transfer (SCNT) has been used for the cloning of various mammals. However, the rates of successful, healthy birth are generally poor. To improve cloning efficiency, we report the utilization of an ‘autologous SCNT’ cloning technique in which the somatic nucleus of a female bovine donor is transferred to its own enucleated oocyte recovered by ovum pick up, in contrast to the routine ‘allogeneic SCNT’ procedure using oocytes from unrelated females. Our results showed that embryos derived from autologous SCNThave significantly higher developmental competence than those derived from allogeneic SCNT, especiallyat the eight-cell (60 vs 44%), morula (45 vs 36%), and blastocyst (38 vs 23%) stages. The pregnancy and birth rates were also higher for the autologous (39 and 23%), compared to the allogeneic (22 and 6%) SCNT groups. Genome-wide histone3-lysine9 methylation profiles reveal that autologous SCNTembryos have less epigenetic defects than the allogeneic SCNTembryos. This study indicates that autologous SCNT can improve the efficiency of bovine cloning with less reprogramming deficiency.

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Fenfen Xie Department of Obstetrics and Gynecology, Reproductive Medicine Center, The First Affiliated Hospital of Anhui Medical University, Hefei, China
NHC Key Laboratory of Study on Abnormal Gametes and Reproductive Tract, Anhui Medical University, Hefei, China
Key Laboratory of Population Health Across Life Cycle, Anhui Medical University, Ministry of Education of the People’s Republic of China, Hefei, China
Anhui Province Key Laboratory of Reproductive Health and Genetics, Hefei, China
Biopreservation and Artificial Organs, Anhui Provincial Engineering Research Center, Anhui Medical University, Hefei, China
Department of Histology and Embryology, Anhui Medical University, Hefei, China

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Junhui Zhang Department of Obstetrics and Gynecology, Reproductive Medicine Center, The First Affiliated Hospital of Anhui Medical University, Hefei, China
NHC Key Laboratory of Study on Abnormal Gametes and Reproductive Tract, Anhui Medical University, Hefei, China
Key Laboratory of Population Health Across Life Cycle, Anhui Medical University, Ministry of Education of the People’s Republic of China, Hefei, China
Anhui Province Key Laboratory of Reproductive Health and Genetics, Hefei, China
Biopreservation and Artificial Organs, Anhui Provincial Engineering Research Center, Anhui Medical University, Hefei, China

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Muxin Zhai First Clinical Medical College, Anhui Medical University, Hefei, China

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Yajing Liu Department of Obstetrics and Gynecology, Reproductive Medicine Center, The First Affiliated Hospital of Anhui Medical University, Hefei, China
NHC Key Laboratory of Study on Abnormal Gametes and Reproductive Tract, Anhui Medical University, Hefei, China
Key Laboratory of Population Health Across Life Cycle, Anhui Medical University, Ministry of Education of the People’s Republic of China, Hefei, China
Anhui Province Key Laboratory of Reproductive Health and Genetics, Hefei, China
Biopreservation and Artificial Organs, Anhui Provincial Engineering Research Center, Anhui Medical University, Hefei, China

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Hui Hu First Clinical Medical College, Anhui Medical University, Hefei, China

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Zhen Yu Department of Obstetrics and Gynecology, Reproductive Medicine Center, The First Affiliated Hospital of Anhui Medical University, Hefei, China
NHC Key Laboratory of Study on Abnormal Gametes and Reproductive Tract, Anhui Medical University, Hefei, China
Key Laboratory of Population Health Across Life Cycle, Anhui Medical University, Ministry of Education of the People’s Republic of China, Hefei, China

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Junqiang Zhang Department of Obstetrics and Gynecology, Reproductive Medicine Center, The First Affiliated Hospital of Anhui Medical University, Hefei, China
NHC Key Laboratory of Study on Abnormal Gametes and Reproductive Tract, Anhui Medical University, Hefei, China
Key Laboratory of Population Health Across Life Cycle, Anhui Medical University, Ministry of Education of the People’s Republic of China, Hefei, China

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Shuai Lin Department of Histology and Embryology, Anhui Medical University, Hefei, China

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Dan Liang Department of Obstetrics and Gynecology, Reproductive Medicine Center, The First Affiliated Hospital of Anhui Medical University, Hefei, China
NHC Key Laboratory of Study on Abnormal Gametes and Reproductive Tract, Anhui Medical University, Hefei, China
Key Laboratory of Population Health Across Life Cycle, Anhui Medical University, Ministry of Education of the People’s Republic of China, Hefei, China
Anhui Province Key Laboratory of Reproductive Health and Genetics, Hefei, China
Biopreservation and Artificial Organs, Anhui Provincial Engineering Research Center, Anhui Medical University, Hefei, China

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Yunxia Cao Department of Obstetrics and Gynecology, Reproductive Medicine Center, The First Affiliated Hospital of Anhui Medical University, Hefei, China
NHC Key Laboratory of Study on Abnormal Gametes and Reproductive Tract, Anhui Medical University, Hefei, China
Key Laboratory of Population Health Across Life Cycle, Anhui Medical University, Ministry of Education of the People’s Republic of China, Hefei, China
Anhui Province Key Laboratory of Reproductive Health and Genetics, Hefei, China
Biopreservation and Artificial Organs, Anhui Provincial Engineering Research Center, Anhui Medical University, Hefei, China

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Emerging evidence has demonstrated that melatonin (MT) plays a crucial role in regulating mammalian reproductive functions. It has been reported that MT has a protective effect on polycystic ovary syndrome (PCOS). However, the protective mechanisms of MT remain poorly understood. This study aims to explore the effect of MT on ovarian function in PCOS and to elucidate the relevant molecular mechanisms in vivo and in vitro. We first analysed MT expression levels in the follicular fluid of PCOS patients. A significant reduction in MT expression levels was noted in PCOS patients. Intriguingly, reduced MT levels correlated with serum testosterone and inflammatory cytokine levels in follicular fluid. Moreover, we confirmed the protective function of MT through regulating autophagy in a DHEA-induced PCOS rat model. Autophagy was activated in the ovarian tissue of the PCOS rat model, whereas additional MT inhibited autophagy by increasing PI3K−-Akt pathway expression. In addition, serum-free testosterone, inflammatory and apoptosis indexes were reduced after MT supplementation. Furthermore, we also found that MT suppressed autophagy and apoptosis by activating the PI3K-Akt pathway in the DHEA-exposed human granulosa cell line KGN. Our study showed that MT ameliorated ovarian dysfunction by regulating autophagy in DHEA-induced PCOS via the PI3K-Akt pathway, revealing a potential therapeutic drug target for PCOS.

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Xue-Ying Zhang The Key Laboratory of Reproductive Genetics (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, China

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Yi-Meng Xiong The Key Laboratory of Reproductive Genetics (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, China

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Ya-Jing Tan The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Li Wang The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Rong Li The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Yong Zhang The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Xin-Mei Liu The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Xian-Hua Lin The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Li Jin The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Yu-Ting Hu The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Zhen-Hua Tang The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Zheng-Mu Wu The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Feng-Hua Yin The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Zheng-Quan Wang The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Ye Xiao The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China

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Jian-Zhong Sheng The Key Laboratory of Reproductive Genetics (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, China
Department of Pathology and Pathophysiology, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, China

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He-Feng Huang The Key Laboratory of Reproductive Genetics (Zhejiang University), Ministry of Education, Hangzhou, Zhejiang, China
The International Peace Maternity and Child Health Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai, China
Shanghai Key Laboratory of Embryo Original Diseases, Shanghai, China

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Fertilization failure often occurs during in vitro fertilization (IVF) cycles despite apparently normal sperm and oocytes. Accumulating evidence suggests that mitochondria play crucial roles in the regulation of sperm function and male fertility. 3-Nitrophthalic acid (3-NPA) can induce oxidative stress in mitochondria, and melatonin, as an antioxidant, can improve mitochondrial function by reducing mitochondrial oxidative stress. The role of sperm mitochondrial dysfunction in fertilization failure during IVF is unclear. The present study revealed that spermatozoa with low, or poor, fertilization rates had swollen mitochondria, increased mitochondria-derived ROS, and attenuated mitochondrial respiratory capacity. 3-NPA treatment enhanced mitochondrial dysfunction in sperm. Spermatozoa with poor fertilization rates, and spermatozoa treated with 3-NPA, had reduced penetration ability. The concentration of melatonin was decreased in semen samples with low and poor fertilization rates. Melatonin, not only decreased excessive mitochondria-derived ROS, but also ‘rescued’ the reduced penetration capacity of spermatozoa treated with 3-NPA. Taken together, the study suggested that mitochondria-derived ROS and mitochondrial respiratory capacity are independent bio-markers for sperm dysfunction, and melatonin may be useful in improving sperm quality and overall male fertility.

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