巴马香猪诱导多能干细胞系的建立

Generation of Bama Mini-pig Induced Pluripotent Stem Cells with a Drug-inducible System

猪在农业和生物医学领域应用非常广泛,尤其是基因修饰的小型猪在器官移植和建立人类疾病模型中有重要的意义。然而,真正的猪胚胎干细胞(embryonic stem cells,ES cells)尚未建立,这极大地限制了转基因猪的产生。目前,诱导多能干细胞(induced pluripotent stem cells,iPS cells)技术是产生多功能干细胞的有效策略。虽然猪iPS cells建系已有数篇报道,但许多有价值的小型猪的iPS cells仍有待建系。利用可诱导基因表达慢病毒系统(Tet-on系统)在小型猪贵州巴马香猪骨髓基质细胞(bone marrow stromal cells,BMSCs)内过表达外源基因Oct4、Sox2、c-Myc和Klf4,获得两株巴马香猪iPS细胞系(biPS4-1,biPS4-2)。获得的巴马香猪iPS cells与人ES cells(hES cells)形态相似;呈碱性磷酸酶(alkaline phosphatase,AP)阳性;表达干细胞特异的标志物SSEA4、Tra-1-81、Nanog、Rex1和CDH1等,但不表达SSEA1;核型检测正常。进一步的研究显示,它在体外和体内均能分化为外、中、内三胚层,具有多向分化的潜力。可诱导的巴马香猪iPS细胞的建立将为建立转基因猪提供便捷的工具,而基于该细胞系建立的转基因猪也有望服务于医学基础研究和器官移植等临床研究。

Pigs have been widely used in the fields of agriculture and biomedicine. In particular, genetically modified mini-pigs play important roles in organ transplantation therapies and human disease models. However, authentic pig embryonic stem cells（ES cells） have not yet been generated, which greatly limits the generation of genetically modified pigs. Induced pluripotent stem（i PS） cell technology appears to be a useful strat-egy to generate pluripotent stem cells. Although pig iPS cells have been generated, iPS cells derived from many valuable mini-pig breeds remain unavailable. Two Bama mini-pig iPS cell lines（biPS4-1 and bi PS4-2）were established by overexpression of Oct4, Sox2, c-Myc and Klf4 in bone marrow stromal cells（BMSCs） via a drug-inducible system（Tet-on system）. Bama mini-pig iPS cells closely resembled human ES cells and ex-pressed ES cell markers, including alkaline phosphatase（AP）, SSEA4, Tra-1-81, Nanog, Rex1 and CDH1,but did not express ES cell marker SSEA1. biPS cells also showed normal karyotypes. These cells were able to differentiate into all three germ layers both in vitro and in vivo. The Bama mini-pig iPS cells reported here may be used to efficiently generate genetically modified mini-pigs and may promote the application of mini-pigs in organ transplantation.