摘要
No efficient therapy for liver fibrosis has been available. This study wasaimed to provide evidence that the introduction of a plasmid expressing antisense tissue inhibitorof metalloproteinase-1 (TIMP-1) into a rat model of immunologically induced liver fibrosis canresult in the increased activity of interstitial collagenase, thus enhancing the degradation ofcollagen. Real-time nested polymerase chain reaction ( RT-Nested-PCR) and gene recombinationtechniques were used to construct a rat antisense TIMP-1 recombinant plasmid that can be expressedin eukaryotic cells. Both the recombinant plasmid and an empty vector ( _(pc)DNA3 ) wereencapsulated with glycosyl-poly-L-lysine and injected into rats suffering from pig serum-inducedliver fibrosis. The expression of ex'ogenous transfected plasmid was assessed by Northern blot,RT-PCR, and Western blot. Hepatic interstitial collagenase activity was detected usingfluorescinisothiocyanate ( FITC)-labeledtype Ⅰ collagen. In addition to hepatic hydroxyprolinecontent, hepatic collagen types Ⅰ and Ⅲ were detected by immunohistochemical staining, and thestages of liver fibrosis by Van Gieson staining. Exogenous antisense TIMP-1 was successfullyexpressed in vivo and could block the gene and protein expression of TIMP-1. Active and latenthepatic interstitial collagenase activities were elevated (P<0. 01), hepatic hydroxyproline contentand the accumulation of collagen types Ⅰ and Ⅲ were lowered, and liver fibrosis was alleviated inthe antisense TIMP-1 group (P <0. 01) as compared with the model group. The results demonstratethat antisense TIMP-1 recombinant plasmids have some inhibitory effect on liver fibrosis.
No efficient therapy for liver fibrosis has been available. This study wasaimed to provide evidence that the introduction of a plasmid expressing antisense tissue inhibitorof metalloproteinase-1 (TIMP-1) into a rat model of immunologically induced liver fibrosis canresult in the increased activity of interstitial collagenase, thus enhancing the degradation ofcollagen. Real-time nested polymerase chain reaction ( RT-Nested-PCR) and gene recombinationtechniques were used to construct a rat antisense TIMP-1 recombinant plasmid that can be expressedin eukaryotic cells. Both the recombinant plasmid and an empty vector ( _(pc)DNA3 ) wereencapsulated with glycosyl-poly-L-lysine and injected into rats suffering from pig serum-inducedliver fibrosis. The expression of ex'ogenous transfected plasmid was assessed by Northern blot,RT-PCR, and Western blot. Hepatic interstitial collagenase activity was detected usingfluorescinisothiocyanate ( FITC)-labeledtype Ⅰ collagen. In addition to hepatic hydroxyprolinecontent, hepatic collagen types Ⅰ and Ⅲ were detected by immunohistochemical staining, and thestages of liver fibrosis by Van Gieson staining. Exogenous antisense TIMP-1 was successfullyexpressed in vivo and could block the gene and protein expression of TIMP-1. Active and latenthepatic interstitial collagenase activities were elevated (P<0. 01), hepatic hydroxyproline contentand the accumulation of collagen types Ⅰ and Ⅲ were lowered, and liver fibrosis was alleviated inthe antisense TIMP-1 group (P <0. 01) as compared with the model group. The results demonstratethat antisense TIMP-1 recombinant plasmids have some inhibitory effect on liver fibrosis.
基金
ThisstudywassupportedbytheNationalNaturalScienceFoundationofChina(No 39970339)
