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Chen Y.,CAS Kunming Institute of Zoology | Chen Y.,University of Chinese Academy of Sciences | Ji S.,CAS Kunming Institute of Zoology | Ji S.,International Biomedical | And 9 more authors.
Journal of Animal and Veterinary Advances | Year: 2011

The objective of this study was to analysis the key factors which may be determinative to Superovulation (SO) in rhesus monkey. Total 361 rhesus monkeys were stimulated with recombinant human gonadotropins and Ova Pick Up (OPU) was conducted by laparoscopic follicular aspiration. Number of total ova and oocytes at Metaphase I (MI) and Metaphase II (Mil) (MI+MEI) from each retrieval was calculated, Mr+MII>10 was defined as effective SO-OPU while MI+MIK10 was defined as non-effective SO-OPU. Parameters including age, month in breeding season, stimulated times and stimulation interval were compared between effective and non-effective SO-OPU. The results showed that 5-10 years old rhesus monkeys were ideal oocytes donors while 3-4 years old and >10 years old monkeys included high rate of poor responders. SO conducted from late October to next February harvested similar fecundity while the rate of poor responders increased in March which imply proper SO breeding season should be from late October to next February. In addition, SO-OPU outcomes were gradually decreased along with the increased times of repeated stimulation. Further analysis showed that stimulation interval affected monkeys' response to recombinant human gonadotropins. When the interval was <2 menstrual cycles, the fecundity compromised significantly. In conclusion, the key determinative factors to SO-OPU should be animal's age and stimulation interval in rhesus monkey stimulated with recombinant human gonadotropins. © Medwell Journals, 2011.

Chen Y.,CAS Kunming Institute of Zoology | Chen Y.,University of Chinese Academy of Sciences | Niu Y.,CAS Kunming Institute of Zoology | Niu Y.,International Biomedical | And 9 more authors.
Journal of Animal and Veterinary Advances | Year: 2011

Current protocols for Superovulation (SO) of rhesus monkeys are not optimized and outcomes vary considerably. There has been little attempt to predict SO outcome according to the stimulation starting time point (Day of cycle) of the candidate oocyte donors. In the present study, researchers focus on the SO initiation timing and we hypothesized that this factor could be used to select optimal oocyte donor monkeys. Superovulation was initiated on menstrual cycle days 1-4 or 5 of 107 adult female rhesus monkeys then 81 animals were identified as good responders by ultrasonography after 6 days of treatment. Oocytes Pick Up (OPU) was performed by laparoscopic follicular aspiration. The numbers of total oocytes and metaphase II (Mil) oocytes from each retrieval were recorded. The results showed that SO begun on days 1-4 or 5 of the menstrual cycle produced similar outcomes indicating that SO initiation in rhesus monkeys can be on days 1-5 of menses. These data showed that SO initiation timing can not be used to predict the SO outcome in rhesus monkeys. © Medwell Journals, 2011.

Ji S.,CAS Kunming Institute of Zoology | Ji S.,Yunnan Key Laboratory of Animal Reproductive Biology | Ji S.,University of Chinese Academy of Sciences | Jin L.,CAS Kunming Institute of Zoology | And 6 more authors.
In Vitro Cellular and Developmental Biology - Animal | Year: 2010

Studies with hepatic progenitor cells from non-human primates would allow better understanding of their human counterparts. In this study, rhesus monkey liver epithelial progenitor cells (mLEPCs) were derived from a small piece of newborn livers in chemical defined serum-free medium. Digested hepatic cells were treated in Ca2+-containing medium to form cell aggregates. Two types of cell aggregates were generated: elongated spindle cells and polygonal epithelial cells. Elongated spindle cells were expressed as vimentin and brachyury, and they were disappeared within 5 d in our cultures. The remaining type consisted of small polygonal epithelial cells that expressed cytokeratin 7 (CK7), CK8, CK18, nestin, CD49f, and E-cad, the markers of hepatic stem cells, but were negative for α-fetoprotein, albumin, and CK19. They can proliferate and be passaged, if on laminin or rat tail collagen gel, to initiate colonies. When cultured with dexamethasone and oncostatin M, the expression of mature hepatocyte markers, such as α-1-antitrypsin, intracytoplasmic glycogen storage, indocyanine green uptake, and lipid droplet generation, were induced in differentiated cells. If transferred onto mouse embryonic fibroblasts feeders, they gave rise to CK19-positive cholangiocytes with formation of doughnut-like structure. Thus, mLEPCs with bipotency were derived from newborn monkey liver and may serve as a preclinical model for assessment of cell therapy in humans. © 2010 The Society for In Vitro Biology.

Tan T.,CAS Kunming Institute of Zoology | Tan T.,University of Chinese Academy of Sciences | Tan T.,Yunnan Key Laboratory of Animal Reproductive Biology | Tang X.,CAS Kunming Institute of Zoology | And 18 more authors.
PLoS ONE | Year: 2011

Trophoblast stem (TS) cells are ideal models to investigate trophectoderm differentiation and placental development. Herein, we describe the derivation of rabbit trophoblast stem cells from embryonic stem (ES) cells. Rabbit ES cells generated in our laboratory were induced to differentiate in the presence of BMP4 and TS-like cell colonies were isolated and expanded. These cells expressed the molecular markers of mouse TS cells, were able to invade, give rise to derivatives of TS cells, and chimerize placental tissues when injected into blastocysts. The rabbit TS-like cells maintained self-renewal in culture medium with serum but without growth factors or feeder cells, whilst their proliferation and identity were compromised by inhibitors of FGFs and TGF-β receptors. Taken together, our study demonstrated the derivation of rabbit TS cells and suggested the essential roles of FGF and TGF-β signalings in maintenance of rabbit TS cell self-renewal. © 2011 Tan et al.

Wang X.,CAS Kunming Institute of Zoology | Wang X.,University of Chinese Academy of Sciences | Wang X.,Yunnan Key Laboratory of Animal Reproductive Biology | Jin L.,CAS Kunming Institute of Zoology | And 11 more authors.
Cell Biology International | Year: 2011

rES (rhesus monkey embryonic stem) cells have similar characteristics to human ES (embryonic stem) cells, and might be useful as a substitute model for preclinical research. Before their clinical application, it is critical to understand the roles of factors that control the differentiation of ES cells into hepatocytes. Here, we analysed the effect of collagen gels on rES cells differentiation into hepatocytes by stepwise protocols. About 80% of DE (definitive endoderm) cells were generated from rES cells after being treated with activin A. The DE cells were then plated on to collagen gels or type I collagen-coated wells with growth factors to induce hepatocyte differentiation. In type I collagen systems, characteristics of immature hepatocytes were observed, including the expression of immature hepatic genes and the generation of 15¡3% AFP (alpha fetoprotein)/CK (cytokeratin)18 double-positive cells. In collagen gel culture, differentiated cells exhibited typical hepatocyte morphology and expressed adult liver-specific genes. The mRNA expression of AFP (immature hepatic gene) was detected at day 11 but decreased at day 18. In contrast, mRNA expression of albumin (mature hepatic gene) was detected at day 11 and increased at day 18. Compared with type I collagen systems, significantly higher AFP/CK18 double-positive cells (68¡7%) were produced in collagen gel culture. Furthermore, some differentiated cells acquired the hepatocytic function of glycogen storage. However, only immature hepatic genes were observed in collagen gel systems if growth factors were absent. Thus, collagen gels combined with hepatocyte-inducing growth factors efficiently promoted differentiation of hepatocytes from rES. © The Author(s) Journal compilation © 2011 Portland Press Limited.

Chen Y.,CAS Kunming Institute of Zoology | Chen Y.,Yunnan Key Laboratory of Animal Reproductive Biology | Chen Y.,International Biomedical | Chen Y.,University of Chinese Academy of Sciences | And 28 more authors.
American Journal of Primatology | Year: 2012

Much effort has been focused on improving assisted reproductive technology procedures in humans and nonhuman primates (NHPs). However, the pregnancy rate after embryo transfer (ET) has not been satisfactory, indicating that some barriers still need to be overcome in this important procedure. One of the key factors is embryo-uterine synchronicity, which is little known in NHPs. The objective of this study was to investigate the available ET time window in rhesus monkey (Macaca mulatta). Eighty-two adult female rhesus monkeys were superovulated with recombinant human FSH. Ovarian phases were identified according to estrogen (E 2) and progesterone (P 4) levels as well as ovarian examination by ultrasonography and laparoscopy. A total of 259 embryos were transferred by the laparoscopic approach into the oviducts of 63 adult female monkeys. Ovarian phases were divided into late follicular and early luteal phases. Similar pregnancy rates (30-36.4%) were obtained from recipients receiving ET either in their late follicular or early luteal phases, regardless of embryo developmental stages. This study indicates that the available time window for ET in rhesus monkeys is from the late follicular to early luteal phases. © 2011 Wiley-Liss, Inc.

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