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目的研究大鼠骨髓源性肝干细胞(BMDLSC)向肝细胞诱导分化过程中H3K27二甲基化变化情况,初步探讨H3K27二甲基化与BMDLSC向肝细胞分化过程的关系。方法分离收集大鼠骨髓细胞,通过免疫磁珠分选获得BMDLSC,而后体外诱导其分化为肝细胞,RT-PCR动态检测其白蛋白(ALB)及甲胎蛋白(AFP)的mRNA表达水平,通过Western blotting及免疫荧光染色检测第0、7、14天核小体组蛋白H3K27二甲基化的水平。结果分离纯化的BMDLSC在体外诱导分化过程中ALB的表达水平逐渐升高,而AFP的表达水平则先升高后下降;新鲜分离的BMDLSC H3K27无明显甲基化,诱导分化后第7天H3K27二甲基化水平较前升高,第14天进一步升高。结论 BMDLSC经体外诱导后定向分化为肝细胞,在分化过程中H3K27二甲基化水平逐渐升高,H3K27二甲基化修饰变化可能参与BMDLSC细胞向肝细胞分化的调控。
Objective To study the changes of H3K27 dimethylation during the differentiation of rat bone marrow-derived liver stem cells (BMDLSC) into hepatocytes and to explore the relationship between the methylation of H3K27 and the differentiation of BMDLSCs into hepatocytes. Methods BMDLSCs were isolated and sorted by immunomagnetic bead sorting. The cells were induced to differentiate into hepatocytes in vitro. The expression of albumin (ALB) and alpha-fetoprotein (AFP) mRNA was detected by RT-PCR. Western blotting and immunofluorescence staining were used to detect the level of nucleoside H3K27 dimethylation on day 0, 7 and 14. Results The expression level of ALB in the isolated and purified BMDLSC gradually increased while the expression level of AFP increased firstly and then decreased. The freshly isolated BMDLSC H3K27 had no obvious methylation. On the 7th day after induction, the expression of H3K27 Methylation level increased compared with the previous, further increased on the 14th day. Conclusion BMDLSCs are differentiated into hepatocytes after induced in vitro. The degree of methylation of H3K27 gradually increases during the differentiation process. The changes of H3K27 dimethylation may be involved in the regulation of BMDLSC differentiation into hepatocytes.