RESUMEN
Vascular endothelial growth factor (VEGF) plays an essential role in the initiation and regulation of angiogenesis-a crucial component of wound healing and cancer growth. Prostaglandins (PGs) stimulate angiogenesis but the precise mechanisms of their pro-angiogenic actions remain unexplained. We investigated whether prostaglandin E(2) (PGE(2)) can induce VEGF expression in rat gastric microvascular endothelial cells (RGMEC) and the signaling pathway(s) involved. We demonstrated that PGE(2) significantly increased ERK2 and JNK1 activation and VEGF mRNA and protein expression. Incubation of RGMEC with PD 98059 (MEK kinase inhibitor) significantly reduced PGE(2)-induced ERK2 activity, VEGF mRNA and protein expression. Furthermore, PD 98059 treatment almost completely abolished JNK1 activation. Our data suggest that PGE(2)-stimulates VEGF expression in RGMEC via transactivation of JNK1 by ERK2. One potential implication of this finding is that increased PG levels in cancers could facilitate tumor growth by stimulating VEGF synthesis and angiogenesis.
Asunto(s)
Dinoprostona/metabolismo , Factores de Crecimiento Endotelial/metabolismo , Endotelio Vascular/citología , Endotelio Vascular/metabolismo , Linfocinas/metabolismo , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Animales , Western Blotting , Células Cultivadas , Medio de Cultivo Libre de Suero/farmacología , Relación Dosis-Respuesta a Droga , Activación Enzimática , Inhibidores Enzimáticos/farmacología , Flavonoides/farmacología , Microcirculación , Proteína Quinasa 8 Activada por Mitógenos , Músculo Liso/citología , Neovascularización Patológica , ARN/metabolismo , ARN Mensajero/metabolismo , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal , Factores de Tiempo , Activación Transcripcional , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial VascularRESUMEN
Gastric mucosal injury triggers angiogenesis and activation of VEGF expression, but the mechanism(s) of VEGF gene activation are not known. In some tissues (e.g. myocardium), hypoxia triggers activation of hypoxia-inducible factor-1alpha (HIF-1alpha), a transcription factor known to activate VEGF gene expression. This study was aimed to determine whether hypoxia and/or alcohol injury may induces HIF-1alpha in gastric mucosa. Normal rat gastric tissue was incubated in organ culture under either hypoxic or normoxic conditions for 6hrs. Rats received, intragastrically, either saline or alcohol and gastric mucosa bordering necrosis was obtained at 1-24hrs. HIF-1alpha mRNA and protein were determined by RT-PCR and Western-blot analysis. HIF-1alpha and VEGF proteins were localized by immunostaining. Incubation of normal gastric mucosa under hypoxia caused a significant elevation of HIF-1alpha mRNA (20+/-2%, p<0.05) and protein (262+/-15%, p<0.005) vs. normoxia. Following alcohol injury, gastric mucosa bordering necrosis demonstrated a significant increase in HIF-1alpha mRNA at 3 and 6hrs (40+/-4%, 19+/-2%; p<0.05), and protein (>300+/-16%; p<0.02 at all time points; highest at 1-3hrs). HIF-1alpha signal was detected in regenerating mucosal microvessels, where it co-localized with VEGF. Since HIF-1alpha initiates transcription of VEGF mRNA, HIF-1alpha activation by ethanol-induced injury is likely responsible for activation of VEGF gene and induction of angiogenesis.
Asunto(s)
Etanol/toxicidad , Mucosa Gástrica/efectos de los fármacos , Neovascularización Patológica/metabolismo , Factores de Transcripción/metabolismo , Animales , Western Blotting , Hipoxia de la Célula , Factores de Crecimiento Endotelial/análisis , Factores de Crecimiento Endotelial/biosíntesis , Factores de Crecimiento Endotelial/genética , Mucosa Gástrica/irrigación sanguínea , Mucosa Gástrica/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia , Inmunohistoquímica , Linfocinas/análisis , Linfocinas/biosíntesis , Linfocinas/genética , Masculino , Necrosis , Técnicas de Cultivo de Órganos , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Regeneración , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Transcripción/análisis , Factores de Transcripción/genética , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial VascularRESUMEN
Nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit angiogenesis in vivo and in vitro, but the mechanism of this action is unclear. Angiogenesis-formation of new capillary vessels-requires endothelial proliferation, migration, and tube formation. It is stimulated by basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF). The cell cycle is regulated positively by cyclins and negatively by cyclin-dependent kinase inhibitors (CKI) and the retinoblastoma protein (pRb). Since the effects of NSAIDs on cell cycle-regulatory proteins in endothelial cells remain unknown, we examined the effect of indomethacin on bFGF-stimulated endothelial cell proliferation and on cell cycle regulatory proteins in rat primary aortic endothelial cells (RAEC). Indomethacin significantly inhibited basal and bFGF-stimulated endothelial cell proliferation. This inhibition correlated significantly with reduced cyclin D1 and increased p21 protein expression. Furthermore, indomethacin reduced pRb phosphorylation. These findings suggest that indomethacin arrests endothelial cell proliferation essential for angiogenesis by modulating cell cycle protein levels.
