Wang J Y, Li Z H, Ye M, Feng Q, Chen Z M, Ye X S, Wu Z G, Wang B, Liu L, Yao J
Department of General Surgery, Shenzhen Children's Hospital, Shenzhen, Guangdong Province, China.
Department of Radiation Oncology, Jinan University of Medical Sciences, Shenzhen Municipal People's Hospital, Shenzhen, Guangdong Province, China.
Genet Mol Res. 2016 Sep 2;15(3):gmr7753. doi: 10.4238/gmr.15037753.
Biliary atresia (BA) is a destructive bile duct disease occurring in newborn children within a few weeks after birth. In this study, the effect of miR-29c and miR-129-5p on epithelial-mesenchymal transition (EMT) in experimental BA was explored by constructing BA mouse models via Rhesus rotavirus vaccine infection. miR-29c and miR-129-5p expression was analyzed by real-time quantitative polymerase chain reaction. EMT was established by induction with transforming growth factor (TGF)-β1. miR-29c and miR-129-5p were overexpressed and inhibited, respectively, by Lipofectamine transfection. EMT-related protein (formin-like 2, FMNL2; E-cadherin; vimentin; and cytokeratin-19, CK-19) expression was analyzed by western blot and immunofluorescent assay. The results indicated that miR-29c and miR-129-5p were downregulated and upregulated in BA mice. TGF-β1 induction caused a time-dependent decrease and increase in miR-29c and miR-129-5p, respectively. Additionally, TGF-β1 induced an increase in FMNL2 and vimentin expression and a decrease in E-cadherin and CK-19 expression (P < 0.05). Overexpression or suppression of miRNA-29c or miR-129-5p, respectively, induced the inhibition of FMNL2 and vimentin, and promotion of E-cadherin and CK-19 expression, in the test groups compared to the non-intervention group (P < 0.05). However, the FMNL2, vimentin, E-cadherin, and CK- 19 expression did not differ between the control and non-intervention groups (P > 0.05). Thus, miR-29c upregulation or miR-129-5p downregulation effectively prevented EMT in BA by regulating the expression of EMT pathway-related proteins. Therefore, miR-29c and miR-129-5p could be utilized as therapeutic targets for BA in the future.
胆道闭锁（BA）是一种发生在新生儿出生后几周内的破坏性胆管疾病。在本研究中，通过恒河猴轮状病毒疫苗感染构建BA小鼠模型，探讨了miR-29c和miR-129-5p对实验性BA上皮-间质转化（EMT）的影响。通过实时定量聚合酶链反应分析miR-29c和miR-129-5p的表达。用转化生长因子（TGF）-β1诱导建立EMT。通过脂质体转染分别过表达和抑制miR-29c和miR-129-5p。通过蛋白质印迹和免疫荧光分析EMT相关蛋白（formin样蛋白2，FMNL2；E-钙黏蛋白；波形蛋白；细胞角蛋白-19，CK-19）的表达。结果表明，miR-29c和miR-129-5p在BA小鼠中表达下调和上调。TGF-β1诱导分别导致miR-29c和miR-129-5p随时间下降和增加。此外，TGF-β1诱导FMNL2和波形蛋白表达增加，E-钙黏蛋白和CK-19表达减少（P<0.05）。与非干预组相比，分别过表达或抑制miRNA-29c或miR-129-5p可诱导测试组中FMNL2和波形蛋白的抑制以及E-钙黏蛋白和CK-19表达的促进（P<0.05）。然而，对照组和非干预组之间FMNL2、波形蛋白、E-钙黏蛋白和CK-19的表达没有差异（P>0.05）。因此，上调miR-29c或下调miR-129-5p可通过调节EMT途径相关蛋白的表达有效预防BA中的EMT。因此，miR-29c和miR-129-5p未来可作为BA的治疗靶点。
