Osteopontin mediates hypoxia-induced proliferation of cultured mesangial cells: Role of PKC and p38 MAPK

Background. We previously reported that hypoxia induces the proliferation o f cultured mesangial cells mediated by the stimulation of intracellular cal cium and the activation of protein kinase C (PKC). In the present study, we examined the roles of mesangial cell specific growth factors (platelet-der ived growth factor and endothelin-1) and osteopontin (OPN) in hypoxia-induc ed proliferation of mesangial cells. In addition, we determined the effect of hypoxia on p38 mitogen-activated protein (MAP) kinase activity and the r oles of both PKC and p38 MAP kinase in hypoxia-induced alterations in OPN a nd mesangial cell growth. Methods. Quiescent cultures of mesangial cells were exposed to hypoxia (3% O-2) or normoxia (18% O-2) in a serum-free medium, and [H-3]-thymidine inco rporation, OPN protein and mRNA expression, and p38 MAP kinase activity wer e assessed. Results. Hypoxic-conditioned medium mimicked the effect of hypoxia on thymi dine incorporation, suggesting the release of diffusable growth promoting f actor(s) by hypoxia. Neither anti-endothelin-1 nor anti-platelet-derived gr owth factor-neutralizing antibodies had an effect on increased thymidine in corporation induced by hypoxia. However, blocking the effects of OPN either with anti-OPN antibody or its beta 3 integrin receptor antibody completely prevented the hypoxia-induced increase in thymidine incorporation. Hypoxia also stimulated OPN protein and mRNA levels. Hypoxia caused an acute activ ation of p38 MAP kinase, which was inhibited by both verapamil and an inhib itor of PKC (calph C). PKC inhibitor and an inhibitor of p38 MAP kinase (SB 203580) reduced the hypoxia-induced stimulation of both OPN and cell growth . Conclusions. These studies provide, to our knowledge, the first evidence de monstrating the role of OPN in hypoxia-induced proliferation of mesangial c ells. In addition, hypoxia causes an activation of p38 MAP kinase in a calc ium- and PKC-dependent manner, and the activation of PKC and p38 MAP kinase appears to be involved in the stimulation of both OPN and mesangial cell p roliferation induced by hypoxia.