588. Effective Prevention of Liver Fibrosis by Liver-Targeted Hydrodynamic Gene Delivery of Matrix Metalloproteinase-13 in Rat Liver Fibrosis Model

Abstract

Liver fibrosis is final stage of chronic liver diseases which may cause liver failure and liver cancer. While various diagnostic methods, including serum marker have been established, no standard therapy has developed. The objective of this study is to assess the approach of overexpressing matrix metalloproteinase-13 gene (MMP13) in rat liver to block liver fibrosis. The rat liver fibrosis model was developed by the bile duct ligation (BDL). And the liver-targeted hydrodynamic gene delivery procedure involves catheter insertion via the inferior vena cava (IVC) to the junction of the IVC and the hepatic veins followed by the hydrodynamic injection of MMP13 expression vector, containing CAG promoter-MMP13-IRES-tdTomato-polyA cassette with temporal blood flow occlusions at the supra- and infra-hepatic IVC. Three groups (n=5 in each group) of animals, BDL alone (non-treated), BDL simultaneously followed by hydrodynamic delivery of pBGI-MMP13 (treated group), and hydrodynamic delivery of pBGI-MMP13 (control) were analyzed for: 1) MMP13 expression in serum and cells in the liver by ELISA, western blotting, immunohistochemical staining, and fluorescent detection; and 2) therapeutic effect on liver fibrosis by analysis of serum fibrotic markers of hyaluronic acid and collagen type IV, and histological analysis including silver staining. Serum concentration of MMP13 in pBGI-MMP13 treated group reached 71.±5.1 pg/ml (p<0.001) compared to ≈5 pg/ml in non-treated group in 2 weeks after the procedure which sustained for following 8 weeks and slowly declined to 37.±5.9 pg/ml in 20 weeks. The protein expression in the liver was also confirmed by western blotting and immunohistochemical staining of the liver samples. The expression of MMP13 contributed on the therapeutic effect which was evidenced by the statistically lower level of the hyaluronic acids of 50.±7.0 ng/ml in treated group than that of 20 ± 19 ng/ml in non-treated group (p<0.5) 2 weeks after the BDL, which sustained for 8 weeks. The serum level of collagen type IV showed similar pattern although no statistical difference was marked. Silver staining showed a significantly lower volume of the fibrotic tissue in treated group than that of non-treated. These results suggest that the liver-targeted hydrodynamic injection of pBGI-MMP13 could achieve long-term gene expression in rat liver and prevent the fibrotic development in the liver.