叶酸缺乏的鼠胚成纤维细胞诱导为iPS细胞的实验研究

An Inducing System for Differentiation of Folate-deficient Mouse Embryo Fibroblasts to Pluripotent Stem Cells

目的研究降低诱导多潜能干(iPS)细胞致癌性的诱导体系,为iPS细胞的应用奠定基础。方法用不含叶酸的培养液对鼠胚成纤维细胞进行培养,3d后用分别含Oct4、Sox2和Klf4因子的3种逆转录病毒载体病毒颗粒进行感染,获得iPS细胞并对其进行鉴定。结果感染后第24天出现了具有胚胎干细胞形态的克隆,诱导效率为(0.010±0.005)%,且AKP染色,Oct4和SSEA-1免疫荧光染色均呈阳性。结论对靶细胞进行无叶酸培养,可以减少外源性转录因子的使用,在不降低诱导效率的前提下降低iPS细胞的致癌性。

Objective To examine the inducing system that can decrease the carcinogenicity of mouse induced pluripotent stem（iPS） cells. Methods Mouse embryonic fibroblasts were cultured in folate-deficient media for 3 days and then separately infected with 3 kinds of retroviruses that could express Oct4,Sox2 or Klf4. The obtained iPS cells were later identified. Results The clones which had the appearance of embryonic stem cells occurred on day 24 after the infection. The inducing efficiency was （0. 010±0. 005） % ,and the obtained iPS cells were positive for alkaline phosphatase,Oct4 or SSEA-1. Conclusion Folate-defi eient media help reduce the use of the external transcription factors and contribute to the differentiation into iPS cells with re- duced tumorigenicity while not reducing the inducing efficiency.