关键词: 肝祖细胞, Yes相关蛋白, 胆管反应, 细胞增殖, 小鼠

Abstract: Objective The aim of this experiment was to investigate the impact of yes-associated protein (YAP)signal on hepatic ductular reaction in mice with choline-deficient and ethionine-supplemented (CDE) diet-induced chronic liver injury (CLI) and YAP's effect on liver progenitor cell proliferation in vitro. Methods The male wild-type C57BL/6J mice were fed with CDE diet for 3 weeks. The Western blot and qPCR were applied to detect YAP protein and gene levels in liver tissue. The immunohistochemistry were used to detect hepatic ductular reaction, the immunofluorescence to evaluate the YAP expression in ductular reaction cells, and the two-step perfusion method was used to isolate primary liver progenitor cells. The liver progenitor cells were infected by lentivirus and were divided into YAP overexpression (YAP-OE group),overexpression empty vector(OE-control group),YAP knockdown(shYAP group), and knockdown empty vector(sh-control group). The infection efficacy was evaluated by Western blot. CCK8 and EdU experiments were applied evaluate the effects of YAP on the liver progenitor cell proliferation. Results The expression of YAP protein inliver tissues in the CDE-induced model increased greatly compared to in the control group and the YAP mRNA level increased by 2.45 times than that in the control group (P＜0.01); the degree of hepatic ductular response in the CDE model and the YAP mRNA level in liver progenitor cells were higher than in the control group; the EdU test found that under the same culture conditions, the percentage of EdU positive cells in the YAP-OE group was significantly increased compared to in the OE-control group [(0.41±0.05) vs. (0.25±0.06), respectively, P＜0.01]; the CCK8 detection showed that the proliferation activity of cells in the YAP-OE group was 1.78 times higher than that in the OE-control group at 72 hours of culture (P＜0.01); the EdU test found that the proportion of EdU positive cells in the shYAP group was significantly lower than that in the sh-control group [(0.25±0.04) vs. (0.13±0.02), respectively, P＜0.01]; the CCK8 detection indicated that the proliferation activity of cells in the shYAP group was 1.92 times lower than that in the sh-control group after 72 hours of culture (P＜0.01). Conclusion The CDE diet could induce chronic liver injury in mice, which results in the increase of YAP expression in progenitor cells with active ductual response. The YAP might promote the proliferation of liver progenitor cells in vitro.

Key words: Liver progenitor cells, Yes-associated protein, Ductular reaction, Proliferation, Mice