The protective effect of docosahexaenoic acid on human retinal microvascular endothelial cells exposed to oxidative stress

Abstract

Ischemic-driven ocular neovascularization is the most common cause of blindness in all age groups. Ischemia in retinopathy leads to several harmful events, including formation of reactive oxygen species (ROS). ROS play an important role in angiogenesis through various redox sensitive cascades and by upregulating VEGF expression, the key molecule in angiogenesis. Docosahexaenoic acid (DHA) is a major dietary ω-3 long-chain polyunsaturated fatty acid and a major structural lipid of sensory and vascular retina. In this study protective mechanisms of DHA in human retinal microvascular endothelial cells (HRMECs) exposed to oxidative stress were assessed.Primary cultures of HRMECs were stimulated by various concentrations of hydrogen peroxide (H2O2) to establish oxidative stress condition. Maximum HRMEC proliferation was observed at 20µM concentration. This condition was used for further experiments to show how physiologic concentration of DHA protects oxidative stress induced HRMEC changes. DHA (10µM) suppressed exogenous H2O2 stimulated proliferation of HRMECs. Anti-proliferative intracellular signaling mechanism of DHA involved PKC/MEK1,2/ERK cascade and coupled to the transcription of c-myc gene. DHA (10 µM) showed a potent scavenging ability of H2O2 stimulated intracellular ROS in HRMECs. NF-κB is a redox-sensitive transcription factor which activates inflammatory responses. DHA (10µM) inhibited H2O2 induced NF-κB activation (RT-PCR and western blot). DHA further suppressed inflammation by inhibition of H2O2 induced TNF-α, IL-1β, and IL-6 mRNA, and COX-2 and iNOS. DHA significantly decreased VEGF transcription and NADPH oxidase expression which is increased under oxidative stress condition. Finally H2O2 induced in vitro angiogenesis of HRMECs were significantly suppressed by DHA.Current treatment methods of retinal neovascularization such as laser photocoagulation, topical steroid, or intravitreal injection of anti-VEGF agent have side effects and work for limited period of time. We have documented that DHA protects HRMECs exposed to oxidative stress through various pathways. Therefore DHA consumption may have additive protective effects in retinal neovascular diseases such as diabetic retinopathy and retinopathy of prematurity