论文部分内容阅读
观察血管内皮细胞生长因子(vascular endothelial growth factor,VEGF)和维生素C对小鼠诱导多能干细胞(induced pluripotent stem cells,iPSCs)体外分化为心肌细胞的诱导作用。采用直接悬浮培养法使iPSCs形成拟胚体(embryoid bodies,EBs),分别以VEGF和维生素C作为诱导剂,自然分化作为阴性对照组,加入1%二甲亚砜诱导剂为阳性对照组。倒置显微镜下观察细胞生长情况,记录跳动的拟胚体出现的时间和数目,计算心肌细胞分化率,细胞免疫荧光检测心肌特异性蛋白cTnT的表达,RT-PCR检测心肌特异性基因β-MHC mRNA的表达。在LIF条件下,iPSCs在饲养层上成集落状生长;未分化的iPSCs中Oct-4及AKP呈阳性表达;与自然分化组相比,二甲亚砜、VEGF和维生素C均能提高iPSCs的心肌细胞分化效率(P<0.05);分化的心肌细胞可自发搏动,同时分化细胞中表达心肌特异性蛋白cTnT以及心肌特异性基因β-MHC。VEGF和维生素C可以促进iPSCs向心肌细胞分化。
To observe the induction of vascular endothelial growth factor (VEGF) and vitamin C on cardiomyocytes induced by mouse induced pluripotent stem cells (iPSCs) in vitro. The direct suspension culture method was used to make iPSCs into embryoid bodies (EBs). VEGF and Vitamin C were used as inducers, respectively, and their differentiation was used as a negative control group. 1% DMSO was used as a positive control. The cell growth was observed under an inverted microscope. The time and number of beating embryoid bodies were recorded. The differentiation rate of cardiomyocytes was calculated. The expression of myocardial specific protein cTnT was detected by immunofluorescence. The expression of β-MHC mRNA expression. Under LIF conditions, iPSCs grew in a colony-like pattern on feeder layers; Oct-4 and AKP were positive in undifferentiated iPSCs; dimethyl sulfoxide, VEGF, and vitamin C increased iPSCs compared with naturally differentiated Cardiomyocytes differentiation efficiency (P <0.05). Differentiated cardiomyocytes spontaneously beat and expressed cardiac specific protein cTnT and cardiac specific gene β-MHC in differentiated cells. VEGF and Vitamin C promote the differentiation of iPSCs into cardiomyocytes.