Yasushi Ikuno1, Andrius Kazlauskas. 1. Department of Ophthalmology, Schepens Eye Research Institute, Harvard Medical School, 20 Staniford Street, Boston, MA 02114, USA.
Abstract
PURPOSE: Proliferative vitreoretinopathy (PVR) is a serious problem in vitreoretinal surgeries. A report of a previous study has indicated that platelet-derived growth factor alpha receptor (alphaPDGFR) plays an important role in a rabbit model of this disease and that a dominant negative alphaPDGFR potently suppresses PVR in an ex vivo setting. Herein, the effect of in vivo gene delivery of a dominant negative alphaPDGFR on PVR was tested in a rabbit model of the disease. METHODS: The dominant negative alphaPDGFR (TalphaR) is a truncated version of the receptor, which does not have the intracellular domain. It was expressed by using a retrovirus. In vitro characterization of TalphaR was performed in primary cultures of rabbit conjunctival fibroblasts (RCFs). Western blot analysis was used to check the expression of TalphaR protein. A type I collagen gel contraction assay was performed to test the efficacy of TalphaR on PDGF-dependent cellular responses in vitro. The in vivo efficacy and specificity of the retrovirus was determined by injecting a green fluorescent protein (GFP) retrovirus into rabbits that had been preinjected with RCFs. The impact of the TalphaR retrovirus on PVR was tested by using the rabbit model in which PVR was induced by the injection of RCFs and platelet-rich plasma (PRP). RESULTS: TalphaR was expressed at more that 50 times the level of endogenous alphaPDGFR in RCFs and severely reduced PDGF-dependent contraction of collagen gels. Intravitreal injection of the GFP retrovirus resulted in expression of GFP primarily in the injected RCFs. Whereas injection of RCFs induced complete retinal detachment in 100% of the animals, co-injection of the TalphaR retrovirus substantially reduced the severity and incidence of retinal detachments. CONCLUSIONS: Gene therapy with a retrovirus used to express a dominant negative alphaPDGFR attenuates PVR in a rabbit model of the disease. This strategy may be a new approach to preventing PVR in humans.
PURPOSE: Proliferative vitreoretinopathy (PVR) is a serious problem in vitreoretinal surgeries. A report of a previous study has indicated that platelet-derived growth factor alpha receptor (alphaPDGFR) plays an important role in a rabbit model of this disease and that a dominant negative alphaPDGFR potently suppresses PVR in an ex vivo setting. Herein, the effect of in vivo gene delivery of a dominant negative alphaPDGFR on PVR was tested in a rabbit model of the disease. METHODS: The dominant negative alphaPDGFR (TalphaR) is a truncated version of the receptor, which does not have the intracellular domain. It was expressed by using a retrovirus. In vitro characterization of TalphaR was performed in primary cultures of rabbit conjunctival fibroblasts (RCFs). Western blot analysis was used to check the expression of TalphaR protein. A type I collagen gel contraction assay was performed to test the efficacy of TalphaR on PDGF-dependent cellular responses in vitro. The in vivo efficacy and specificity of the retrovirus was determined by injecting a green fluorescent protein (GFP) retrovirus into rabbits that had been preinjected with RCFs. The impact of the TalphaR retrovirus on PVR was tested by using the rabbit model in which PVR was induced by the injection of RCFs and platelet-rich plasma (PRP). RESULTS:TalphaR was expressed at more that 50 times the level of endogenous alphaPDGFR in RCFs and severely reduced PDGF-dependent contraction of collagen gels. Intravitreal injection of the GFP retrovirus resulted in expression of GFP primarily in the injected RCFs. Whereas injection of RCFs induced complete retinal detachment in 100% of the animals, co-injection of the TalphaR retrovirus substantially reduced the severity and incidence of retinal detachments. CONCLUSIONS: Gene therapy with a retrovirus used to express a dominant negative alphaPDGFR attenuates PVR in a rabbit model of the disease. This strategy may be a new approach to preventing PVR in humans.
Authors: Steven Pennock; David Kim; Shizuo Mukai; Matthew Kuhnle; Dal W Chun; Joanne Matsubara; Jing Cui; Patrick Ma; David Maberley; Arif Samad; Robert J Van Geest; Sarit L Oberstein; Reinier O Schlingemann; Andrius Kazlauskas Journal: Am J Pathol Date: 2013-04-09 Impact factor: 4.307