TGF-beta(1) modulates NOS expression and phosphorylation of Akt/PKB in rat myocytes exposed to hypoxia-reoxygenation

Myocardial hypoxia-reoxygenation (H-R) is associated with upregulation of inducible nitric oxide synthase (iNOS), decrease in endothelial NOS (eNOS), and increase in protein kinase B (PKB). Previous work also shows that transforming growth factor-beta(1) (TGF-beta(1)) can attenuate myocardial injury induced by H-R. We examined the modulation of NOS and PKB expression in response to H-R by TGF- beta(1). Myocytes from Sprague-Dawley rat hearts were cultured and exposed to hypoxia (95% N(2)-5% CO(2), PO(2) ~30 mmHg) for 24 h and reoxygenation (95% air-5% CO(2)) for 3 h. Myocytes were then examined for lactate dehydrogenase (LDH) release, iNOS activity (conversion of L-[(3)H]arginine to L-[(3)H]citrulline), iNOS and eNOS expression, and PKB phosphorylation. H-R alone resulted in myocyte injury, upregulation of iNOS activity and expression, decrease in eNOS expression, and increase in PKB phosphorylation (all P < 0.05 vs. cells cultured in normoxic conditions). Treatment of myocytes with TGF-beta(1) (1 ng/ml) resulted in a reduction in LDH release, attenuation of the alterations in NOS expression (both iNOS and eNOS), and PKB phosphorylation in response to H-R (all P < 0.05 vs. H-R alone). These observations suggest that TGF-beta(1) decreases H-R injury and attenuates alterations in NOS and PKB phosphorylation in myocytes exposed to H-R.