全反式维甲酸诱导肝前体细胞向肝细胞分化的研究

Role of all-trans retinoic acid in the differentiation of hepatic progenitor cells

摘要:

目的：探讨全反式维甲酸（All-trans retinoic acid，ATRA）对肝前体细胞分化的影响。方法：取孕14.5 d胎鼠制备肝前体细胞，瞬时转染带白蛋白启动子（Albumin promoter）的荧光素酶报告质粒pALB-Luc，以便实时检测细胞的分化状态。优化ATRA诱导条件，分别于诱导后3、6、9 d收集细胞总RNA，半定量RT-PCR检测分化相关指标的变化。并于诱导后11 d用苏木精-伊红染色法观察细胞形态的改变，PAS染色观察细胞的分化成熟程度。结果：ATRA浓度为1 μmol/L时对肝前体细胞分化诱导作用最强。与对照组相比，随培养时间的延长，ATRA组分化早期指标膜蛋白（Delta like，DLK）、细胞角蛋白（Cytokeratin19，CK19）表达降低更为明显，而甲胎蛋白（Alpha fetal protein，AFP）、白蛋白（Albumin，ALB）表达更为升高。形态学观察细胞体积增大，呈多边形，核浆比率降低，相嵌紧密排列，可见双核细胞。PAS染色阳性率上升。结论：浓度1 μmol/L的ATRA对肝前体细胞分化诱导作用最强，ATRA能诱导肝前体细胞分化为类肝细胞。

Abstract:

Objective：To investigate the effect of all-trans retinoic acid （ＡＴＲＡ） in the differentiation of hepatic progenitor cell. Methods：The hepatic progenitor cell was isolated from E14.5 mouse liver and transfected with pALB-Luc，in which the ex－pression of luciferase was driven by （Albumin，ALB） promoter. The concentration of ATRA based on the expression of ALB was optimized. The change of differentiation markers of （Alpha fetal protein，AFP），（Delta like，DLK），（Cytokeratin19，CK19） and ALB in the hepatic progenitor cell treated with ATRA was evalu－ated by RT-PCR. The morphological change was observed by HE staining and PAS staining. Results：The expression of ALB was the highest when the cell was treated with ATRA a concentration of 1 μmol/L. Compared with control group，the expressions of early markers of DLK and CK19 more clearly decreased while AFP and ALB were increased in experimental group. The morphological changes were mainly enlargement of cellular size，decrease of karyoplasms ratio and increase of glycogenesis. Conclusion：ATRA can induce the differentiation of hepatic progenitor cell，especially at the concentration of 1 μmol/L.