ABSTRACT
BACKGROUND:The hepatic stellate cells(HSCs)plays a key role in the development of liver fibrosis.Studies have shown that bone marrow-derived mesenchymal stem cell(BMSCs)transplantation can be used to treat liver fibrosis,but the mechanism for reversal of liver fibrosis remains unknown.OBJECTIVE:To explore the mechanism of bone marrow mesenchymal stem cells to regulate the proliferation of HSCs under co-culture in vitro.METHODS:Rat BMSCs and HSCs in the experimental group were cultured in the plastic culture plate(6 holes)to establish the upper and lower double-cell co-culture system.Rat normal fibroblast cell lines were seeded as control group;HSCs were cultured alone as blank group.Cell proliferation was determined by WST8 and cell cycle was determined by flow cytometry.The Cyclin D1 and P27 mRNA expression in HSC was determined by reverse transcription-polymerase chain reaction(RT-PCR)and the level of Cyclin D1 and P27 protein by Western blot.RESULTS AND CONCLUSION:HSCs co-cultured with BMSCs significantly inhibited HSC proliferation compared with the blank and control groups at 24,48,and 72 hours(P ＜ 0.01);Flow cytometry showed that the percentage of G_0/G_1 phase cells of co-culture group was increased but the S phase cells reduced(P ＜ 0.01)compared with the other groups at 72 hours,and BMSCs blocked HSC to convert from G_0/G_1 period to S phase.After HSCs co-cultured with BMSCs for 24 hours,the expression of Cyclin D1 mRNA and protein was reduced,and significantly less than the blank and control groups at 72 hours(P ＜ 0.01);no differences were detected in P27 mRNA expression in each group during the co-culture(P ＞ 0.05).After co-culture of 24 hours,the p27 protein expression was significantly increased compared with the blank and control groups(P ＜ 0.01).BMSCs inhibited the proliferation of HSCs,possibly through inhibiting CydinDI expression,increasing the p27 protein expression to cause cell cyde arresting in G_0/G_1 phase.