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) expression in cultured rat aortic vascular smooth muscle (VSM) cells. In addition, we determined whether hypoxia-induced increases in OPN and cell proliferation 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 assessed. Cells exposed to hypoxia for 24 h exhibited a significant increase in [(3)H]thymidine incorporation followed by a significant increase in cell number at 48 h in comparison with respective normoxic controls. Exposure to hypoxia produced significant increases in OPN protein and mRNA expression at 2 h followed by a gradual decline at 6 and 12 h, with subsequent significant increases at 24 h. Neutralizing antibodies to either OPN or its receptor beta3 integrin but not neutralizing antibodies to beta5 integrin prevented the hypoxia-induced increase in [(3)H]thymidine incorporation. Inhibitors of protein kinase C (PKC) and p38 mitogen-activated protein (MAP) kinase also reduced the hypoxia-induced stimulation of proliferation and OPN synthesis. Exposure to high-glucose (HG) (25 mmol/l) medium under normoxic conditions also resulted in significant increases in OPN protein and mRNA levels as well as the proliferation of VSM cells. Under hypoxic conditions, HG further stimulated OPN synthesis and cell proliferation in an additive fashion. In conclusion, hypoxia-induced proliferation of cultured VSM cells is mediated by the stimulation of OPN synthesis involving PKC and p38 MAP kinase. In addition, hypoxia also enhances the effect of HG conditions on both OPN and proliferation of cultured VSM cells, which may have important implications in the development of diabetic atherosclerosis associated with arterial wall hypoxia.

Hypoxia and high glucose cause exaggerated mesangial cell growth and collagen synthesis: role of osteopontin.

The effect of hypoxia on the proliferation and collagen synthesis of cultured rat mesangial cells was examined under normal-glucose (NG, 5 mM) and high-glucose (HG, 25 mM)-media conditions. In addition, a role for osteopontin (OPN) in mediating these processes was assessed. Quiescent cultures were exposed to hypoxia (3% O(2)) and normoxia (18% O(2)) in a serum-free medium with NG or HG, and cell proliferation, collagen synthesis, and OPN expression were assessed. Cells exposed to hypoxia in NG medium resulted in significant increases in [(3)H]thymidine incorporation, cell number, and [(3)H]proline incorporation, respectively. HG incubations also produced significant stimulation of these parameters under normoxic conditions, which were markedly enhanced in cells exposed to hypoxia in HG medium. In addition, hypoxia and HG stimulated the mRNA levels of type IV collagen, and the combination of hypoxia and HG resulted in additive increases in type IV collagen expression. Hypoxia and HG also stimulated OPN mRNA and protein levels in an additive fashion. A neutralizing antibody to OPN or its beta(3)-integrin receptor significantly blocked the effect of hypoxia and HG on proliferation and collagen synthesis. In conclusion, these results demonstrate for the first time that hypoxia in HG medium produces exaggerated mesangial cell growth and type IV collagen synthesis. In addition, OPN appears to play a role in mediating the accelerated mesangial cell growth and collagen synthesis found in a hyperglycemic and hypoxic environment.

Screening of in vitro antibacterial activity of Terminalia chebula, Eclapta alba and Ocimum sanctum.

Study of in vitro antibacterial activity of extracts from the plants T. chebula, E. alba and O. sanctum was carried out by the disk diffusion technique. All showed such activity against human pathogenic Gram positive and Gram negative bacteria. The activity against Salmonella organisms was shown only by T. chebula; against Shigella organisms by T. chebula and E. alha; but not by O. sanctum. The widest spectrum of antibacterial activity was shown by T. chebula. It was also most potent. The antibacterial spectrum of E. alba was in between that of T. chebula and O. sanctum. The narrowest spectrum of antibacterial activity was also most potent. The antibacterial spectrum of E. alba was in between that of T. chebula and O. sanctum. The narrowest spectrum of antibacterial activity was observed in O. sanctum.

Assessment of burn wound sepsis by swab, full thickness biopsy culture and blood culture--a comparative study.

Fifty patients with burns ranging from 30 to 50 per cent of their body surface area were monitored for sepsis throughout their hospital stay using swab, blood and full thickness biopsy culture techniques. The relative merits of these techniques in the diagnosis of burn wound sepsis were evaluated. Only 62.5 per cent of the patients with a positive surface culture showed signs of clinical sepsis, while 87.5 per cent of the patients with significant bacterial count on biopsy culture showed signs of clinical sepsis. A decrease in bacterial count on follow up correlated with clinical improvement while a count of 10(8) orgs/gm indicated a bad prognosis. Wound surface cultures, though the simplest method gave poor indication of the organisms invading into the burn wound. Blood cultures were of only prognostic value. Full thickness biopsy culture and quantification of the number of bacteria in the burn wound was felt to be the best method for rapid diagnosis and for assessing the progress of burn wound infection.