关键词: 肝纤维化 , 结缔组织生长因子, 短发夹RNA, 转化生长因子β, RNA干扰

Abstract: Objective：To investigate the effect of CTGF short hairpin RNA(shRNA) on the expression of TGF-β1, CTGF, type Ⅰ procollagen, type Ⅲ procollagen mRNA and extracellular matrix secretion in HSC-T6. Methods： The CTGFshRNA plasmids containing EGFP successfully constructed and screened, were transfected to HSC-T6 with the transfection reagent Metafectene. There were another three HSC-T6 groups: one with plasmids and without CTGFshRNA, one with only Metafectene, and one untreated. The expression of TGF-β1, CTGF, type Ⅰ procollagen and type Ⅲ procollagen mRNA levels were determined by reverse transcriptionpolymerase chain reaction (RTPCR) after 24 and 48 hours. The content of precollagen type Ⅲ, Ⅳtype collagen, and hyaluronate (HA), laminin (LN) in the supernatants were determined by radioimmunoassay. Cell transfection efficiency was analyzed by flow cytometry. Results： The transfection efficiency of CTGFshRNA plasmids in HSC were (60±5)％, (42±3)％ at 24 and 48 hours respectively.Compared with those of the blank control, there was no difference in the expression of genes and extracellular matrix secretion in HSC-T6 in the other groups, the group with plasmids and without CTGF shRNA and the group with only Metafectene. Expression of CTGF, type Ⅰ collagen and type Ⅲ collagen mRNA were markedly downregulated in CTGFshRNA transfected HSC-T6(P<0.01 or P<0.05). But there was no difference in the expression of TGF-β1 mRNA. The contents of precollagen type Ⅲ, Ⅳtype collagen, hyaluronate (HA) , and laminin (LN) in the supernatants decreased significantly in CTGFshRNA transfected HSCT6(P<0.01 or P<0.05). Conclusion： CTGFshRNA can regulate the gene expression of the growth factor and the extracellular matrix molecule and inhibits the extracellular matrix secretion in HSCT6.

Key words: RNA interference, Short hairpin RNA, Transforming growth factor β, Connective tissue growth factor, Liver fibrosis