The broad long-term objectives are to establish the role of connective tissue growth factor (CTGF) in causing fibrotic disease. CTGF is a highly pro-fibrogenic molecule which is over- expressed in all fibrotic lesions examined to date. It is transcriptionally activated by transforming growth factor-beta (TGF-beta) and mediates many of the matrix-inducing properties that have previously been attributed to TGF-beta. The studies described in this proposal focus on the role of CTGF in liver fibrosis, including that related to alcohol abuse. Preliminary data show that CTGF is over-expressed in fibrotic livers and is produced by hepatic stellate cells (HSCs), the principal fibrogenic cell type, both in response to TGF-beta and as a function of activation. HSCs show enhanced adhesion and levels of alpha smooth muscle actin in response to CTGF. In addition, ethanol and its fibrogenic metabolite, acetaldehyde, stimulate CTGF transcription in fibroblasts. Our hypothesis is that local up-regulation of CTGF in the liver drives the fibrogenic response, including that initiated by alcohol.
Our Specific Aims are (1) To determine mechanisms of CTGF regulation in HSCs, including the role played by TGF-beta and acetaldehyde (which stimulate HSCs and CTGF production) as well as retinoic acid and TNF-alpha (which inhibit HSC function and CTGF production); (2) To determine the effects of CTGF on HSC function by examining HSC DNA synthesis, division, matrix metabolism, vitamin A content, and adhesion in HSCs treated with or over-expressing various mass forms (1OkDa, 16-20kDa, 38kDa) of CTGF which occur naturally in vivo and which are a product of HSCs maintained in vitro, and to determine the role of CTGF-stimulated kinases in these processes; and (3) To produce recombinant adeno-associated viruses for the delivery of the CTGF gene into the liver in vivo to directly establish the ability of 10 kDa and 38kDa CTGF to stimulate liver fibrosis. These studies will define the fibrogenic properties of CTGF in terms of its regulation, biological properties, signaling mechanisms and protein structure. In addition, these studies will help establish whether CTGF is a therapeutic target for treating fibrosis, which is a contributing factor in 45 percent of deaths in the USA.
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