RESUMEN
Osteopontin (OPN), an arginine-glycine-aspartate (RGD)-containing adhesive glycoprotein, is constitutively expressed in rat aorta and carotid arteries and is markedly elevated in response to vascular injury. OPN is chemotactic for vascular smooth muscle cells (SMCs), suggesting a role in vascular remodeling. However, the mechanism for the regulation of OPN expression is poorly understood. In the present study, the effect of platelet-derived growth factor (PDGF) on OPN mRNA expression was investigated in cultured rat aortic SMCs (RASMCs). When RASMCs were stimulated with 1 nmol/L PDGF, a 2.4-fold increase in OPN mRNA expression was observed at 3 hours (P < .05) that peaked at 14 hours with a 6.7-fold increase (P < .001). This induction was blocked by a monoclonal anti PDGF antibody. Further studies revealed that OPN mRNA expression was induced by PDGF-AB or PDGF-BB but not by PDGF-AA, indicating that only the beta-type PDGF receptor mediates this response. Compared with basic fibroblast growth factor, epidermal growth factor, transforming growth factor-beta, and interleukin-1 beta, PDGF was the most potent factor studied to induce OPN mRNA expression in RASMCs. Immunohistochemical studies demonstrated the elevation of OPN protein in PDGF-stimulated RASMCs. The temporal expression of OPN mRNA after rat carotid artery balloon angioplasty as assessed by both reverse transcription-polymerase chain reaction and Northern blot analysis revealed a 1.5-fold increase at 6 hours (P < .01) that peaked at 1 and 3 days with a 3.1-fold increase (P < .001). Immunohistochemical studies of carotid artery after angioplasty localized OPN expression in the medical SMCs at 1 day, ie. at a time of significant platelet adherence to the injured vessel, and thereafter to the intimal lesion during neointimal formation. These data suggest that OPN expression in vascular SMCs is regulated by PDGF through the beta-type PDGF receptor in vitro, and possibly in vivo in situations that involve PDGF released from platelets or other cellular sources, such as blood vessels after angioplasty injury.
Asunto(s)
Arterias Carótidas/metabolismo , Músculo Liso Vascular/metabolismo , Factor de Crecimiento Derivado de Plaquetas/farmacología , Sialoglicoproteínas/biosíntesis , Angioplastia de Balón , Animales , Arterias Carótidas/patología , Células Cultivadas , Inmunohistoquímica , Masculino , Músculo Liso Vascular/patología , Osteopontina , Reacción en Cadena de la Polimerasa , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND AND PURPOSE: Free radical generation mediates part of the ischemic neuronal damage caused by the excitatory amino acid glutamate. Carvedilol, a novel multiple-action antihypertensive agent, has been shown to scavenge free radicals and inhibit lipid peroxidation in swine heart and rat brain homogenates. Therefore, we studied the neuroprotective effect of carvedilol on cultured cerebellar neurons and on CA1 hippocampal neurons of gerbils exposed to brain ischemia. METHODS: Neuroprotective mechanisms were studied using an in vitro ischemia model of cultured rat cerebellar granule cell neurons exposed to either glutamate or oxygen free radical-generating systems. Prevention of lipid peroxidation by carvedilol was studied by measuring the formation of thiobarbituric acid-reactive substance. Gerbil CA1 neuron survival was examined by direct neuronal count 7 days after 6 minutes of global ischemia with reperfusion. RESULTS: Carvedilol protected cultured neurons in a dose-dependent manner against glutamate-mediated excitotoxicity (inhibitory concentration [IC50] = 1.1 microM) as well as against a 20-minute oxidative challenge (IC50 = 5 microM). The IC50 against the oxidative challenge was lowered to 1.3 microM by growing neurons for 24 hours in the presence of carvedilol. At 10 microM carvedilol inhibited lipid peroxidation 50% and 73% (n = 4, p < 0.001) in neurons exposed to two different free radical-generating systems. Neuroprotection of 52% (n = 22, p = 0.009 versus vehicle) of gerbil CA1 hippocampal neurons was achieved by pretreatment and posttreatment with subcutaneous injection of 3 mg/kg carvedilol twice a day for 4 and 3 days, respectively. CONCLUSIONS: Carvedilol provided neuroprotection in both in vitro and in vivo models of neuroinjury, where oxygen radicals are likely to play an important role. Therefore, carvedilol may reduce the risk of cerebral ischemia and stroke by virtue of both its antihypertensive action and its antioxidative properties.