Over-expression Of CNTF In Bone Marrow Mesenchymal Stem Cells Protects The Retina From Blue-light Injury

Background: Blue-light is everywhere in daily life.Moderate intensity of blue-light exposure plays essential roles in maintaining visual function and the circadian rhythm.However,intensive blue light exposure induces oxidative stress in retinal pigment epithelial(RPE)cells,which ultimately leads to the retinal damage.The development and progression of optical damage is similar to that of age-related macular degeneration(AMD).But laser and anti-neovascular treatment were proved not so ideal.Therefore,the optical damage model has significant value to the study of AMD.Bone marrow mesenchymal stem cells(BMSCs)have been widely used for the investigation of retinopathies,due to their neuroprotective and immunoregulative effects,and properties such as high plasticity.CNTF protects photoreceptor cells,RPE cells and Müller cells in the retina,and therefore facilitates the survival of retinal cells.A single injection of CNTF protein produces a short duration of action,while repeated injections easily cause endophthalmitis and may disrupt the augenstructure.In our study,CNTF-modified BMSCs were used to investigate their effects on blue-light injured retina.Objective: Cell or animal blue light injury model was established by exposing RPE cells or SD rat retina to blue light.The influences of CNTF-modified BMSCs on RPE cells as well as SD rat retina were examined.In addition,the effects of gene modification on the functions of BMSCs were investigated.This study provides novel strategies for the treatment of retinopathies.Methods: Primary culture of BMSCs and RPE cells was prepared in vitro followed by identification.Adenoviral and lentiviral system was applied to establish BMSCs overexpressing CNTF.RPE cells subjected to blue-light injury were co-cultured with CNTF-BMSCs and GFP-BMSCs in vitro.In vivo study was carried out by transplanting CNTF-BMSCs and GFP-BMSCs in subretinal space of SD rats after blue-light injury.The MDA and SOD levels in RPE cells or retina were measured by a microplate reader.The concentrations of secreted VEGF in cells or retina were examined by ELISA.Retinal vessel samples were used for the determination of retinal vascular leakage.After pretreatment with Baf A1,the changes in autophagic flux in RPE cells from different experimental groups were assessed by Western blot.In order to investigate the differentiation of stem cells,the co-localization between CK18 and GFP in CNTF-BMSCs was evaluated by confocal laser microscope.The relationship between autophagy and apoptosis was determined by evaluating the fluorescence of LC3 and TUNEL.F-ERG assay was used to investigate the influences of CNTF-BMSCs and GFP-BMSCs on visual function after blue light injury.Results: BMSCs and RPE primary culture and identification were achieved.Intensive blue light exposure resulted in the oxidative injury in RPE cells and retina.Co-culture of RPE cells with CNTF-BMSCs or transplantation of CNTF-BMSCs in subretinal space of SD rats could reduce MDA level,increase SOD activity and reduce the secretion of VEGF.The transplanted CNTF-BMSCs were CK18 positive and increased the amplitude in Max F-ERG assay of SD rats.Conclusions: CNTF modification enhances the resistance against oxidative stress in BMSCs.CNTFmodified BMSCs inhibits the VEGF secretion from retina,prevents retinal vascular leakage,promotes autophagy and mitigates apoptosis in retina.After transplantation,these cells have the ability to differentiate into RPE cells and alleviate retinopathy induced by blue-light exposure.