Wnt3a诱导小鼠肝前体细胞上皮-间质转化

发布时间：2019-03-11 11:02

【摘要】：目的探讨Wnt3a对小鼠肝前体细胞发生上皮-间质转化的影响。方法将表达Wnt3a的腺病毒Ad-GFPWnt3a转入小鼠肝前体细胞中,与空白对照组相比,观察其形态变化。细胞划痕实验和细胞迁移实验观察其对小鼠肝前体细胞迁移能力的影响。实时荧光定量Real-time PCR和Western blot分别检测小鼠肝前体细胞中上皮标志物和间质标志物的表达改变。结果镜下观察,高表达Wnt3a的小鼠肝前体细胞由不规则多边形变为长梭形。细胞划痕实验结果显示,Ad-Wnt3a组细胞48h迁移距离为(0.53±0.05)mm,Ad-GFP组细胞48 h迁移距离为(0.33±0.02)mm,相对对照组,其迁移距离增加,差异具有统计学意义(P0.05)。细胞迁移实验结果显示,Ad-GFP组24 h穿过小孔的细胞为(10.33±2.08)个,而Ad-Wnt3a组穿过的细胞为(62.00±3.60)个,相对对照组,其穿过小孔的细胞数量增加,差异具有统计学意义(P0.01)。RT-PCR和Western blot结果显示,间质标志物N-cadherin、vimentin和snail的mRNA和蛋白水平表达上调,相反上皮标志物E-cadherin和CK-18的mRNA水平和蛋白水平表达下调,差异具有统计学意义(P0.05)。结论 Wnt3a能够促使小鼠肝前体细胞发生上皮-间质转化,提示其可能参与了肝纤维化的发展进程。
[Abstract]:Objective to investigate the effect of Wnt3a on epithelial-interstitial transformation of mouse hepatic precursor cells. Methods adenovirus Ad-GFPWnt3a expressing Wnt3a was transferred into mouse liver precursor cells and compared with the blank control group, the morphological changes were observed. Cell scratch test and cell migration test were used to observe their effects on the migration ability of mouse liver precursor cells. Real-time fluorescent quantitative Real-time PCR and Western blot were used to detect the expression of epithelial markers and interstitial markers in mouse liver precursor cells. Results under microscope, the mouse liver precursor cells with high expression of Wnt3a changed from irregular polygons to fusiform. The results of cell scratch test showed that the 48-hour migration distance of cells in Ad-Wnt3a group was (0.53 卤0.05) mm, and the migration distance of cells in mm,Ad-GFP group was (0.33 卤0.02) mm, which was higher than that in control group. The difference was statistically significant (P0.05). The results of cell migration test showed that the number of cells passing through the pores in Ad-GFP group was (10.33 卤2.08), and that in Ad-Wnt3a group was (62.00 卤3.60). Compared with the control group, the number of cells traversing through micropores increased, and the number of cells passing through micropores in Ad-Wnt3a group was significantly higher than that in control group (P < 0.05). The results of RT-PCR and Western blot showed that the expression of mRNA and protein were up-regulated in stroma markers such as N-cadherin, vimentin and snail. On the contrary, the levels of mRNA and protein of epithelial markers E-cadherin and CK-18 were down-regulated, the difference was statistically significant (P0.05). Conclusion Wnt3a can promote epithelial-interstitial transformation of hepatic precursor cells in mice, suggesting that it may be involved in the development of hepatic fibrosis.
【作者单位】：重庆医科大学分子医学与肿瘤研究中心;重庆医科大学生物化学与分子生物学教研室;重庆医科大学病理生理学教研室;
【基金】：国家自然科学基金(81071770,81201679)~~
【分类号】：R329

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