The Role of Histone Deacetylase 4 during Hepatic Stellate Cell activation

Abstract

RATIONALE: The activation of hepatic stellate cells (HSCs) is a major feature of fibrotic liver disease during which HSCs lose their high vitamin A content and undergo transdifferentiation to myofibroblast-like cells. Knowledge of the signal pathways that are involved in the activation process has provided insight into targets for treating and preventing hepatic fibrosis. However, no effective treatment for human liver fibrosis has been established, emphasizing the need to explore other treatments. The ability of various histone deacetylase (HDAC) inhibitors to inhibit different phenotypic changes associated with HSCs activation indicated that some HDACs might play a role during HSC activation and liver fibrosis. Thus identification and specific targeting of HDACs that are involved in liver fibrosis might prove a good treatment strategy.

AIM: The aim of this study is to determine whether HDAC4 is expressed in HSCs and to identify its possible role in these cells.

METHODS: We used immunofluorescence and Western blotting to show the presence of HDAC4 in HSCs. Immunofluoresence and co-immunoprecipitation were used to study the possible co-localization of HDAC4 with cytoskeletal proteins. siRNA was used to silence HDAC4 in order to study its role in the regulation of genes whose expression is influenced by HSC activation. Real time (quantitative) PCR was used to quantify the relative mRNA level of the genes while their protein level was quantified by Western blotting. For further study on this project, we produced lentiviral vectors encoding HDAC4 shRNA using the Gateway technology, and used the vectors to produce viruses.

RESULTS: We showed that HDAC4 was expressed in HSCs. It was partially localized in the nucleus as well as cytoplasm, but also in filaments reminiscent of cytoskeletal fibres. However, it did not co-localize with any of the cytoskeletal proteins studied. Our data also showed that HDAC4 is important for proper expression of lysyl oxidase (LOX), an important enzyme that is upregulated during HSC activation, and plays an important role in the cross-linking of collagen and elastin molecules.

CONCLUSION: The expression of HDAC4 in the HSCs and its regulatory role on LOX shows that HDAC4 is important in HSC activation and could be a target for the treatment of liver fibrosis after proper elucidation of its role.

Date of Award	4 Sep 2008
Original language	English
Supervisor	Leo van Grunsven (Promotor)