RESUMO
OBJECTIVE: To determine the effects of Smad7 gene transfer in the prevention of fibrogenic responses by the retinal pigment epithelium, a major cause of proliferative vitreoretinopathy after retinal detachment, in mice. METHODS: Retinal detachment-induced proliferative vitreoretinopathy in a mouse model. Forty-eight eyes received either an adenoviral gene transfer of Smad7 or Cre recombinase gene only. The eyes were histologically analyzed. A retinal pigment epithelial cell line, ARPE-19, was used to determine whether Smad7 gene transfection suppresses the fibrogenic response to transforming growth factor (TGF) beta2 exposure. RESULTS: The Smad7 gene transfer inhibited TGF-beta2/Smad signaling in ARPE-19 cells and expression of collagen type I and TGF-beta1 but had no effect on their basal levels. In vivo Smad7 overexpression resulted in suppression of Smad2/3 signals and of the fibrogenic response to epithelial-mesenchymal transition by the retinal pigment epithelium. CONCLUSION: Smad7 gene transfer suppresses fibrogenic responses to TGF-beta2 by retinal pigment epithelial cells in vitro and in vivo. Clinical Relevance Smad7 gene transfer might be a new strategy to prevent and treat proliferative vitreoretinopathy.
Assuntos
Modelos Animais de Doenças , Expressão Gênica/fisiologia , Epitélio Pigmentado Ocular/metabolismo , Proteína Smad7/genética , Fator de Crescimento Transformador beta/farmacologia , Vitreorretinopatia Proliferativa/prevenção & controle , Adenoviridae/genética , Animais , Western Blotting , Linhagem Celular , Colágeno Tipo I/metabolismo , Fibronectinas/metabolismo , Fibrose , Técnica Indireta de Fluorescência para Anticorpo , Vetores Genéticos , Camundongos , Epitélio Pigmentado Ocular/efeitos dos fármacos , Epitélio Pigmentado Ocular/patologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Transfecção , Fator de Crescimento Transformador beta1/metabolismo , Regulação para Cima , Vitreorretinopatia Proliferativa/metabolismo , Vitreorretinopatia Proliferativa/patologiaRESUMO
Proliferative vitreoretinopathy (PVR) is one of the major causes of the failure of retinal detachment surgery. Its pathogenesis includes a fibrotic reaction by the retinal pigment epithelium and other retina-derived non-neural cells, leading to fixation of the detached retina. We examined the role of p38 mitogen-activated protein kinase (MAPK) in transforming growth factor (TGF)-beta2-dependent enhancement of the fibrogenic reaction in a human retinal pigment epithelial cell line, ARPE-19, and also evaluated the therapeutic efficacy of inhibiting p38MAPK by adenoviral gene transfer of dominant-negative (DN) p38MAPK in a mouse model of PVR. Exogenous TGF-beta2 activates p38MAPK in ARPE-19 cells. It also suppresses cell proliferation, but this was unaffected by addition of the p38MAPK inhibitor, SB202190. SB202190 interfered with TGF-beta2-dependent cell migration and production of collagen type I and fibronectin, but had no effect on basal levels of these activities. While SB202190 did not affect phosphorylation of the C-terminus of Smads2/3, it did suppress the transcriptional activity of Smads3/4 as indicated by a reporter gene, CAGA12-Luc. Gene transfer of DN-p38MAPK attenuated the post-retinal detachment fibrotic reaction of the retinal pigment epithelium in vivo in mice, supporting its effectiveness in preventing/treating PVR.
