Ethanol activates NF kappa B DNA binding and p56(lck) protein tyrosine kinase in human osteoblast-like cells

Alcoholics frequently suffer from moderate to severe bone loss that results in bone fractures. Both decreased bone production and increased bone resor ption have been postulated to contribute to ethanol (ETOH)-mediated bone lo ss. Bone resorption is induced by several proinflammatory cytokines such as interleukin-1 and -6, The expression of these cytokines is induced by the transcription factor NF kappaB, which, in turn, is activated by several kin ases. It follows that protein kinase and NF kappaB activation may contribut e to ETOH-induced bone loss, Accordingly, we sought to determine if ETOH ac tivates protein tyrosine kinases (PTK) and NF kappaB DNA binding in a human osteoblast-like cell line (HOBIT), Ethanol at 50 and 100 mmol/L (reflectiv e of blood ethanol levels reached in chronic alcoholics) for 24 h did not a lter HOBIT cell viability. In contrast, 200 mmol/L ethanol decreased cell v iability by 40%. Treatment of HOBIT cells with 100 mmol/L ETOH induced nucl ear NF kappaB:DNA complex formation and NF kappaB activity. Incubation of H OBIT cells with ETOH at 50 and 100 mmol/L for 30 min induced a 2.5- and 4.2 -fold increase in PTK activity, respectively, Preincubation of HOBIT cells with damnacanthal (DAM), which inhibits p56(lck), blocked ETOH-mediated PTK activity; whereas, preincubation with herbimycin A, which inhibits pp60(sr c), did not. DAM inhibited both ethanol-induced NF kappaB activation in HOB IT cells and interleukin-6 expression in primary human osteoblasts. Finally , preincubation with the protein kinase C inhibitor, bisindolylmaleimide I HCl (BIS), diminished ETOH-mediated PTK activity; whereas, preincubation wi th the protein kinase A inhibitor, H89, did not. These data demonstrate tha t ETOH induces NF kappaB nuclear translocation through p56(lck) in HOBIT ce lls. BIS' inhibition of PTK activation suggests that ETOH activates PTK thr ough a protein kinase C-dependent pathway. These data suggest that ETOH may contribute to bone loss through activation of signal transduction that res ults in production of an osteoclastogenic cytokine interleukin-6) in osteob lasts. (Bone 28:167-173; 2001) (C) 2001 by Elsevier Science Inc, All rights reserved.