NSP4与整联蛋白α2β1在轮状病毒致肝外胆管上皮细胞损伤中的作用

The role of NSP4 and integrin α2β1 in the damage of extrahepatic biliary epithelial cells

目的探讨NSP4与整联蛋白α2β1在轮状病毒致肝外胆管上皮细胞损伤中的作用。方法取Balb／c小鼠的肝外胆管进行肝外胆管上皮细胞培养至第三天，用激光共聚焦方法检测整联蛋白α2、β1亚基及轮状病毒NSP4。用siRNA干扰技术沉默培养至第三天的肝外胆管上皮细胞的整联蛋白以后加入轮状病毒，用Real TimeRT-PCR方法检测以亚基及轮状病毒NSP4的mRNA），并进行统计学分析。结果用激光共聚焦方法检测到了整联蛋白α2、β1亚基存在于肝外胆管上皮细胞膜表面；轮状病毒NSP4从感染细胞向紧邻的非感染细胞进行扩散；siRNA干扰技术表明沉默整联蛋白以后轮状病毒NSP4 mRNA含量1±0．01较对照组0．56±0．09减低（P〈0．05）。结论肝外胆管上皮细胞表面表达整联蛋白α2、β1且整联蛋白α2β1与轮状病毒NSP4相互作用并进一步导致肝外胆管上皮细胞损伤。

Objective To investigate the role of rotavirus non structural protein 4 （NSP4） and integrin α2β1 in the damage of extrahepatic biliary epithelial cells. Methods Extrahepatic hilt duct of Balb/c mice was harvested for ex vivo culture of biliary epithelial cells （EHBE）. The primary cultured epithelial cells were fixed. Immunofluorescent （IF） assay was performed to detect integrin subunits α2β1 and rotavirus nonstructural protein 4 （NSP4）. Small interfering RNA targeting integrin α2 was transfected into EHBE before rotavirus infection. Real time RT-PCR was used to measure the mRNAs of integrin α2 and NSP4. Results Integrin α2 and β1 were detected at the surface of EHBE. Further- more, the expression of NSP4 was colocalized with integrin a2 on the surface of rotavirus-infected ex- trahepatic biliary epithelial cells, indicating that NSP4 released from rotavirus infected EHBE binded with the integrin α2 on the adjacent uninfected cells. Silencing of integrin α2 decreased the mRNA of rotavirus NSP4. Conclusions The integrin subunits α2, β1 are expressed and locate on the surface of primary cultured extrahepatic biliary epithelial cells. The integrin α2β1 and the rotavirus NSP4 play importnat roles in the injury of extrahepatic biliary epithelial cells.