INHIBITION OF NFKB IN ACTIVATED RAT HEPATIC STELLATE CELLS BY PROTEASOME INHIBITORS AND AN IKB SUPER-REPRESSOR

The hepatic stellate cell (HSC), following a fibrogenic stimulus, is t ransformed from a quiescent to an activated cell. Cytokines induce NF kappa B activity in activated but not in quiescent HSCs with subsequen t expression of NF kappa B-responsive genes, such as intercellular adh esion molecule (ICAM)-1 and interleukin (IL)-6. We investigated the ef fect of proteasome inhibitors and an I kappa B super-repressor on the cytokine mediated activation of NF kappa B, ICAM-1, and IL-6 in activa ted HSCs. Culture-activated HSCs were stimulated with IL-1 beta or tum or necrosis factor alpha (TNF alpha) in the presence or absence of pro teasome inhibitors, ALLN or MG-132, or after infection with an adenovi rus expressing the I kappa B super-repressor (Ad5I kappa B) or beta-ga lactosidase (Ad5LacZ) as a control. NF kappa B activity was evaluated by immunofluorescence and by electrophoretic mobility shift assay. The steady state level of cytoplasmic I kappa B protein was measured by W estern Blot, ICAM-1 and IL-6 expression was measured by reverse transc riptase-polymerase chain reaction and enzyme-linked immunosorbant assa y. Proteasome inhibitors, which block the degradation of I kappa B, an d the Ad5I kappa B, which provides an exogenous nondegradable I kappa B, block the stimulation of NF kappa B activity by TNF alpha and IL-1 beta in activated HSCs. These reagents block the subsequent nuclear tr anslocation of p65 NF kappa B and induction of ICAM-1 and IL-6 by cyto kines. The specificities of the proteasome inhibitors and the I kappa B super-repressor are demonstrated by their failure to block c-Jun N-t erminal kinase induction by cytokines. Cytokine-induced stimulation of NF kappa B, ICAM-1, and IL-6 is blocked by proteasome inhibitors and Ad5I kappa B in activated HSCs. Inhibition of I kappa B alpha degradat ion is a potential target for anti-inflammatory therapy in the liver a nd might influence the activation process of HSCs following fibrotic s timuli.