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Yan H.,Beijing Jiaotong University | Yan Z.,Beijing Jiaotong University | Ma Q.,Beijing Jiaotong University | Ma Q.,The Key Laboratory of Embryo Molecular Biology | And 8 more authors.
Journal of Genetics and Genomics | Year: 2011

Reconstructed embryos derived from intersubspecies somatic cell nuclear transfer (SCNT) have poorer developmental potential than those from intrasubspecies SCNT. Based on our previous study that Holstein dairy bovine (HD) mitochondrial DNA (mtDNA) haplotype compatibility between donor karyoplast and recipient cytoplast is crucial for SCNT embryo development, we performed intersubspecies SCNT using HD as donor karyoplast and Luxi yellow heifer (LY) as recipient cytoplast according to mtDNA haplotypes determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis. The results demonstrated that intersubspecies mtDNA homotype SCNT embryos had higher pre- and post-implantation developmental competence than intrasubspecies mtDNA heterotype embryos as well as improved blastocyst reprogramming status, including normal H3K9 dimethylation pattern and promoter hypomethylation of pluripotent genes such as Oct4 and Sox2, suggesting that intersubspecies SCNT using LY oocytes maintains HD cloning efficiency and may reprogram HD nuclei to develop into a normal cloned animal ultimately. Our results indicated that karyoplast-cytoplast interactions and mtDNA haplotype compatibility may affect bovine intersubspecies SCNT efficiency. This study on bovine intersubspecies SCNT is valuable for understanding the mechanisms of mtDNA haplotype compatibility between karyoplast and cytoplast impacting the bovine SCNT efficiency, and provides an alternative and economic resource for HD cloning. © 2011. Source


Moussa M.,Beijing Jiaotong University | Moussa M.,Shanghai JiaoTong University | Moussa M.,The Key Laboratory of Embryo Molecular Biology | Moussa M.,and Shanghai Laboratory of Embryo and Reproduction Engineering | And 11 more authors.
Animal Reproduction Science | Year: 2015

The oocyte is a central regulator of multiple aspects of female fertility, including ovarian follicular development and early embryogenesis. During its prolonged diplotene arrest, the oocyte is subjected to endogenous (i.e., reactive oxygen species from metabolism) and exogenous (i.e., heat stress, malnutrition) sources of damage-inducing factors, which may lead to a progressive deterioration of oocyte quality. A deficit in oocyte competence can lead not only to a failure of fertilization but also to a lower developmental rate after fertilization. Thus, an appropriate environment for growth and maturation of the oocyte, in vivo and in vitro, is critical to ensure optimal oocyte quality. The objectives of the current review are to give an overview of some maternal key factors that influence oocyte quality in cattle and describe some of the findings to date in the hope of obtaining competent oocytes that could be used for clinical and applied purposes. © 2015 Elsevier B.V. Source


Moussa M.,Shanghai JiaoTong University | Moussa M.,The Key Laboratory of Embryo Molecular Biology | Moussa M.,Shanghai Laboratory of Embryo and Reproduction Engineering | Moussa M.,Shanghai Tao Tao Transgenic Corporation | And 10 more authors.
Science China Life Sciences | Year: 2014

Cryopreservation techniques for mammalian oocytes and embryos have rapidly progressed during the past two decades, emphasizing their importance in various assisted reproductive technologies. Pregnancies and live births resulting from cryopreserved oocytes and embryos of several species including humans have provided proof of principle and led to the adoption of cryopreservation as an integral part of clinical in vitro fertilization. Considerable progress has been achieved in the development and application of the cryopreservation of mammalian oocytes and embryos, including preservation of the reproductive potential of patients who may become infertile, establishment of cryopreserved oocyte banks, and transport of oocytes and embryos internationally. However, the success rates are still far lower than those obtained with fresh oocytes and embryos, and there are still obstacles that need to be overcome. In this review, we address the major obstacles in the development of effective cryopreservation techniques. Such knowledge may help to eliminate these hurdles by revealing which aspects need improvement. Furthermore, this information may encourage further research by cryobiologists and increase the practical use of cryopreservation as a major part of assisted reproductive technologies for both humans and animal species. © 2014 The Author(s). Source


Yang C.,Shanghai JiaoTong University | Fan S.,Shanghai JiaoTong University | Tang H.,Shanghai JiaoTong University | Tang H.,The Key Laboratory of Embryo Molecular Biology | And 8 more authors.
Shengwu Gongcheng Xuebao/Chinese Journal of Biotechnology | Year: 2014

We established methods to isolate human amniotic fluid-derived progenitor cells (hAFPCs), and analyze the ability of hAFPCs to secrete human coagulation factor IX (hFIX) after gene modification. The hAFPCs were manually isolated by selection for attachment to gelatin coated culture dish. hFIX cDNA was transfected into hAPFCs by using a lentiviral vector. The hFIX protein concentration and activity produced from hAFPCs were determined by enzyme-linked immunosorbent assay (ELISA) and clotting assay. The isolated spindle-shaped cells showed fibroblastoid morphology after three culture passages. The doubling time in culture was 39.05 hours. Immunocytochemistry staining of the fibroblast-like cells from amniotic fluid detected expression of stem cell markers such as SSEA4 and TRA1-60. Quantitative PCR analysis demonstrated the expression of NANOG, OCT4 and SOX2 mRNAs. Transfected hAFPCs could produce and secrete hFIX into the culture medium. The observed concentration of secreted hFIX was 20.37%±2.77% two days after passage, with clotting activity of 16.42%±1.78%. The amount of hFIX:Ag reached a plateau of 50.35%±5.42%, with clotting activity 45.34%±4.67%. In conclusion, this study established method to isolate and culture amniotic fluid progenitor cells. Transfected hAFPCs can produce hFIX at stable levels in vitro, and clotting activity increases with higher hFIX concentration. Genetically engineered hAFPC are a potential method for prenatal treatment of hemophilia B. © 2014 by the Institute of Microbiology, the Chinese Academy of Sciences and the Chinese Society for Microbiology. Source

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