Induction of cell death in activated hepatic stellate cells by targeted gene expression of the thymidine kinase/ganciclovir system

Biochem Biophys Res Commun. 2004 Apr 16;316(4):1107-15. doi: 10.1016/j.bbrc.2004.02.147.

Abstract

Liver fibrosis is the result from a relative imbalance between synthesis and degradation of matrix proteins. Following liver injury of any etiology, hepatic stellate cells undergo a response known as activation, which is the transition of quiescent cells into proliferative, fibrogenic, and contractile myofibroblasts. Upon this cellular transdifferentiation the effector cell becomes the major source of fibrillar and non-fibrillar matrix proteins resulting in excessive scar formation and cirrhosis, the end stage of fibrosis. Concomitant with progressive liver fibrosis, the tissue inhibitor of metalloproteinases-1 (TIMP-1) is strongly activated in hepatic stellate cells. We have developed a recombinant replication-defective adenovirus in which the TIMP-1 promoter is coupled to the herpes simplex virus thymidine kinase gene rendering activated hepatic stellate cells susceptible to ganciclovir. This novel targeted suicide gene approach was validated in a culture model considered to reflect an accelerated time course of the cellular and molecular events that occur during liver fibrosis. We demonstrate that transfer of the suicide gene to culture-activated hepatic stellate cells results in a strong expression of the respective transgene as assessed by Northern blot and Western blot analyses. The enzyme catalyzed the proper conversion of its prodrug subsequently initiating programmed cell death as estimated by caspase-3 assay and Annexin V-Fluos staining. Altogether, these results indicate that induction of programmed cell death is a promising approach to eliminate fibrogenic HSC.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Apoptosis / genetics*
  • Cell Differentiation
  • Cells, Cultured
  • Ganciclovir / metabolism
  • Gene Expression Regulation / genetics*
  • Gene Silencing
  • Gene Targeting / methods*
  • Hepatocytes / classification
  • Hepatocytes / metabolism*
  • Hepatocytes / pathology
  • Liver Cirrhosis / genetics*
  • Liver Cirrhosis / metabolism*
  • Liver Cirrhosis / pathology
  • Male
  • Rats
  • Rats, Sprague-Dawley
  • Thymidine Kinase / genetics
  • Thymidine Kinase / metabolism
  • Tissue Inhibitor of Metalloproteinase-1 / genetics*
  • Tissue Inhibitor of Metalloproteinase-1 / metabolism*

Substances

  • Tissue Inhibitor of Metalloproteinase-1
  • Thymidine Kinase
  • Ganciclovir