Nagata Mihoko, Takahashi Masafumi, Muramatsu Shin-Ichi, Ueda Yasuji, Hanazono Yutaka, Takeuchi Koichi, Okada Koji, Suzuki Yutaka, Kondo Yasushi, Suemori Masafumi, Ikeda Uichi, Nakano Imaharu, Kobayashi Eiji, Hasegawa Mamoru, Ozawa Keiya, Nakatsuji Norio, Shimada Kazuyuki
Department of Medicine, Jichi Medical School, Tochigi 329-0498, Japan.
J Gene Med. 2003 Nov;5(11):921-8. doi: 10.1002/jgm.431.
Embryonic stem (ES) cells continually proliferate and can generate large numbers of differentiated cells. Genetic manipulation of transplantable cells derived from primate ES cells offers considerable potential for development research and regenerative cell therapy. However, protocols for efficient gene transfer into primate ES-cell-derived cells have not yet been established.
Spontaneously contracting areas were derived from cynomolgus monkey ES cells. Features of cardiomyocytes in the area were analyzed according to gene expression (RT-PCR), morphology (immunostaining and electron microscopy), and function (intracellular calcium transience). Beating cells were transduced using a simian immunodeficiency virus (SIV) vector expressing enhanced green fluorescence protein (EGFP), then transplanted into ischemic rat myocardium.
Beating cells derived from monkey ES cells displayed gene expression, ultrastructural and functional properties of early-stage cardiomyocytes. Highly efficient (97% cardiac phenotype) and stable transduction of these ES-cell-derived cardiomyocytes was achieved using SIV vector without altering contractile function. In addition, transduced cardiomyocytes survived in the myocardium of a rat myocardial infarction model.
A lentiviral vector system based on SIV represents a useful vehicle for genetic modification of cardiomyocytes derived from primate ES cells, and can extend the application of primate ES cells to gene therapy.
胚胎干细胞持续增殖，能够产生大量分化细胞。对源自灵长类胚胎干细胞的可移植细胞进行基因操作，在发育研究和再生细胞治疗方面具有巨大潜力。然而，高效将基因导入灵长类胚胎干细胞衍生细胞的方案尚未建立。
从食蟹猴胚胎干细胞获得自发收缩区域。根据基因表达（逆转录聚合酶链反应）、形态（免疫染色和电子显微镜）及功能（细胞内钙瞬变）分析该区域心肌细胞的特征。使用表达增强型绿色荧光蛋白（EGFP）的猿猴免疫缺陷病毒（SIV）载体转导跳动细胞，然后将其移植到缺血大鼠心肌中。
源自猴胚胎干细胞的跳动细胞表现出早期心肌细胞的基因表达、超微结构和功能特性。使用SIV载体可实现对这些胚胎干细胞衍生心肌细胞的高效（97%心脏表型）且稳定的转导，同时不改变收缩功能。此外，转导的心肌细胞在大鼠心肌梗死模型的心肌中存活。
基于SIV的慢病毒载体系统是对源自灵长类胚胎干细胞的心肌细胞进行基因修饰的有效工具，可将灵长类胚胎干细胞的应用扩展到基因治疗。
