Pancreatic stellate cell activation by ethanol and acetaldehyde: is it mediated by the mitogen-activated protein kinase signaling pathway?

Pancreatic fibrosis is a characteristic feature of alcoholic chronic pancreatitis. Recent studies suggest that activated pancreatic stellate cells (PSCs) are the major cell-type involved in pancreatic fibrogenesis. Cultured PSCs become activated when exposed to ethanol or its metabolite acetaldehyde (as indicated by increased alpha-smooth muscle actin [alpha-SMA] expression and increased collagen synthesis). However the intracellular signaling mechanisms responsible for ethanol- or acetaldehyde-induced PSC activation remain to be fully elucidated. One of the major signaling pathways known to regulate protein synthesis in mammalian cells is the mitogen-activated protein kinase (MARK) pathway. To examine the effects of ethanol and acetaldehyde on the MAPK pathway (by assessing the activities of the 3 major subfamilies (extracellular-regulated kinases 1 and 2 [ERK 1/2], JNK and p38 kinase) in PSCs and to examine the role of p38 kinase in mediating the ethanol- and acetaldehyde-induced increase in alpha-SMA expression in activated rat PSCs. Rat PSCs were incubated with ethanol (50 mM) or acetaldehyde (200 microM) for 15 min, 30 min, 60 min, and 24 h; and activities of ERK 1/2, JNK, and p38 kinase were assessed in cell lysates using kinase assays and Western blot. In addition, rat PSCs were treated with the specific p38 MAPK inhibitor SB203580 in the presence or absence of ethanol or acetaldehyde for 24h, and activation of the downstream protein kinase MAPKAP kinase-2 (an indicator of p38 MAPK activity) was assessed by Western blot. Specific inhibitors were also used to inhibit the activity of ERK 1/2 and JNK. Following inhibition of the above signaling pathways, alpha-SMA expression by PSCs was assessed by Western blot. Ethanol and acetaldehyde increased the activation of all 3 subfamilies (ERK 1/2, JNK and p38 kinase) of the MAPK pathway in PSCs. Treatment of PSCs with SB203580 abolished the ethanol- and acetaldehyde-induced increase in p38 MAPK activity and also prevented the induction of alpha-SMA expression in PSCs. However, inhibition of ERK 1/2 and JNK had no effect on ethanoland acetaldehyde-induced alpha-SMA expression in PSCs. (1) The MAP kinase pathway is induced in PSCs after exposure to ethanol or acetaldehyde and this induction is sustained for at least 24h. (2) The p38 MAPK pathway mediates the activation (as indicated by increased alpha-SMA expression) of PSCs by ethanol or acetaldehyde.