Hypoxia stimulates osteopontin expression and proliferation of cultured vascular smooth muscle cells - Potentiation by high glucose

We examined the effect of hypoxia on proliferation and osteopontin (OPN) ex pression in cultured rat aortic vascular smooth muscle (VSM) cells. In addi tion, we determined whether hypoxia-induced increases in OPN and cell proli feration are altered under hyperglycemic conditions. Quiescent cultures of VSM cells were exposed to hypoxia (3% O-2) or normoxia (18% O-2) in a serum -free medium, and cell proliferation as well as the expression of OPN was a ssessed. Cells exposed to hypoxia for 24 h exhibited a significant increase in [H-3]thymidine incorporation followed by a significant increase in cell number at 48 h in comparison with respective normoxic controls. Exposure t o hypoxia produced significant; increases in OPN protein and mRNA expressio n at 2 h followed by a gradual decline at 6 and 12 h, with subsequent signi ficant increases at 24 h. Neutralizing antibodies to either OPN or its rece ptor beta3 integrin but not neutralizing antibodies to beta5 integrin preve nted the hypoxia-induced increase in [H-3]thymidine incorporation. Inhibito rs of protein kinase C (PKC) and p38 mitogen-activated protein (MAP) kinase also reduced the hypoxia-induced stimulation of proliferation and OPN synt hesis. Exposure to high-glucose (HG) (25 mmol/l) medium under normoxic cond itions also resulted in significant increases in OPN protein and mRNA level s as well as the proliferation of VSM cells. Under hypoxic conditions, HG f urther stimulated OPN synthesis and cell proliferation in an additive fashi on. In conclusion, hypoxia-induced proliferation of cultured VSM cells is m ediated by the stimulation of OPN synthesis involving PKC and p38 MAP kinas e. In addition, hypoxia also enhances the effect of HG conditions on both O PN and proliferation of cultured VSM cells, which may have important implic ations in the development of diabetic atherosclerosis associated with arter ial wall hypoxia.