携带rIL-10基因的大鼠肝细胞及肝星状细胞对TGF-β1表达的影响

Effects of hepatocyte and hepatic stellate cell transfected rat interleukin-10 gene on the expression of transforming growth factor-β1

目的:观察携带大鼠白介素10(rIL-10)基因的正常大鼠肝细胞(BRL)及大鼠肝星状细胞(HSC-T6)是否可通过旁分泌作用或自分泌作用减少肝星状细胞分泌转化生长因子β1(TGF-β1),进一步探讨rIL-10抗纤维化的作用机制。方法:IL-10基因通过尾静脉高压注射干预猪血清诱导肝纤维化大鼠,造模结束时收集各实验组大鼠血浆;通过半乳糖受体介导的脂质体转染试剂及阳离子脂质体转染试剂将携带rIL-10基因的pCDNA3-rIL-10质粒与pCDNA3空质粒转染大鼠BRL细胞及HSC-T6细胞,收集转染后24及48小时细胞培养上清;另将携带pCDNA3-rIL-10质粒与pCDNA3空质粒的大鼠BRL细胞与HSC-T6细胞共培养,收集共培养24及48小时后细胞培养上清液;ELISA法检测各实验组细胞培养上清液中TGF-β1的表达。结果:rIL-10基因干预后的肝纤维化大鼠血浆中TGF-β1表达下降,携带rIL-10基因的BRL细胞及HSC-T6细胞可表达高浓度细胞因子rIL-10,HSC-T6分泌的rIL-10能减少自身表达TGF-β1水平,BRL细胞表达rIL-10通过旁分泌作用可抑制与之共培养HSC-T6上清中TGF-β1的表达。结论:rIL-10基因可通过旁分泌或自分泌作用抑制肝星状细胞分泌转化生长因子β1,进一步阐明rIL-10基因的抗纤维化作用机理。

Objective：To explore the anti-fibrotic effect of rat interleukin-10.Methods：Rat interleukin-10 eukaryotic expression vector（pcDNA3-rIL-10） was transferred to rat＇s liver by hydrodynamic-based transfection.At the end of treatment,all rats were put to death and then collected plasma.The plasmid of pcDNA3-rIL-10 and pcDNA3 were transfected into BRL cells and hepatic stellate cells by asialoglycoprotein receptor mediated liposome PEIjet-gal and cationic lipid-containing liposomes respectively.The cell culture supernatant was harvested 24 h or 48 h after transfection,and then BRL cells transfected pcDNA3-rIL-10 and pcDNA3 and HSCT6 cells were co-cultured in the 6-well plate,cell culture supernatant was harvested 24 h or 48 h after co-culture.The expression of rIL-10 gene and TGF-β1 were detected by ELISA.Results：ELISA analysis showed that the expression of TGF-β1 in fibrotic rat＇s plasma was significantly decreased after rIL-10 gene treatment,and that the high concentration rIL-10 was detected in BRL and HSCT6 cell culture supernatant after rIL-10 gene transfection,and that autocrine rat interleukin-10 of HSC-T6 could decrease dramatically the expression of TGF-β1 in cell culture supernatant,and that paracrine rat interleukin-10 of BRL could decrease dramatically the expression of TGF-β1 in cell co-culture supernatant.Conclusion：Paracrine or autocrine rat interleukin-10 suppresses the expression of transforming growth factor-β1 in rat hepatic stellate cells.