ACTIVATION OF HEPATIC STELLATE CELLS BY TGF-ALPHA AND COLLAGEN TYPE-IIS MEDIATED BY OXIDATIVE STRESS THROUGH C-MYB EXPRESSION

Excessive production of collagen type I is a major contributor to hepa tic fibrosis, Activated (myofibroblastic), but not quiescent, hepatic stellate cells (lipocytes) have a high level of collagen type I and al pha-smooth muscle actin expression. Therefore, stellate cell activatio n is a critical step in hepatic fibrosis, Here we show that quiescent stellate cells were activated by the generation of free radicals with ascorbate/FeSO4 and by malondialdehyde, a product of lipid peroxidatio n, In addition, stellate cell activation by collagen type I matrix and TGF alpha was blocked by antioxidants, such as d-alpha-tocopherol and butylated hydroxytoluene. Moreover, oxidative stress, TGF alpha and c ollagen type I markedly stimulated stellate cell entry into S-phase, N FkB activity and c-myb expression, which were prevented by antioxidant s. c-myb antisense oligonucleotide blocked the activation and prolifer ation of stellate cells induced by TGF alpha. Nuclear extracts from ac tivated, but not from quiescent, stellate cells formed a complex with the critical promoter E box of the alpha-smooth muscle actin gene, whi ch was disrupted by c-myb and NFkB65 antibodies, and competed by c-myb and NFkB cognate DNA. c-Myb expression was also stimulated in activat ed stellate cells in carbon tetrachloride-induced hepatic injury and f ibrogenesis. This study indicates that oxidative stress plays an essen tial role, through the induction of c-myb and NFkB, on stellate cell a ctivation.