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目的诱导人胚胎生殖细胞(hEGC)向心肌细胞分化,初步探讨其定向分化过程及调控机制。方法组织块培养法体外培养hEGC,采用抗坏血酸诱导hEGC向心肌细胞分化,通过CCK-8法测定诱导后心肌细胞增殖率,免疫荧光法检测心肌连接蛋白Cx43的表达;利用半定量RT-PCR的方法鉴定hEGC和诱导后心肌细胞GATA-4和ACTC1等基因的表达;结合生物信息学方法推断分化过程中各蛋白的相互作用关系。结果诱导后细胞阳性表达Cx43,且表现出一定的增殖能力;其表达GATA-4、ACTC1等基因的强度与未经诱导的hEGC均有所不同;GATA-4、Nkx2.5与各类心肌肌钙蛋白有着密切的联系。结论 hEGC能诱导分化为心肌细胞;蛋白质组学方法可以协助探讨其分化过程及调控机制。
Objective To induce human embryonic germ cells (hEGC) to differentiate into cardiomyocytes, and to investigate their differentiation and regulation mechanism. Methods hEGC was cultured in vitro and cultured in vitro. Ascorbic acid was used to induce hEGC to differentiate into cardiomyocytes. The proliferation of cardiomyocytes was measured by CCK-8 assay. The expression of Cx43 was detected by immunofluorescence. The expression of Cx43 was detected by semi-quantitative RT-PCR Identification of genes such as GATA-4 and ACTC1 in hEGC and induced cardiomyocytes, and bioinformatics methods were used to deduce the relationship between the proteins in the process of differentiation. Results The positive expression of Cx43 was observed after induction, and showed some proliferative capacity. The expression of GATA-4, ACTC1 and other genes were different from those of non-induced hEGC; GATA-4, Nkx2.5 and various types of cardiac muscle Calprotectin is closely linked. Conclusion hEGC can induce the differentiation into cardiomyocytes. The proteomics method can help to explore the differentiation process and regulation mechanism.