oa Docosahexaenoic Acid (DHA) Remedies Hyperglycemia-Induced Retinal Endothelial Dysfunction: A Potential Therapeutic Intervention For Diabetic Retinopathy

Diabetic retinopathy (DR) is a leading cause of blindness among working diabetic patients worldwide. The current therapeutic strategies are limited by several side effects. Thus, it is important to investigate novel approaches for therapeutic intervention. Early inflammatory response and late neovascularization (NV) are cardinal clinical signs of the DR. We have previously demonstrated the involvement of bioactive lipids especially the products of omega-6 polyunsaturated fatty acids (PUFAs) such as arachidonic acids in the development of retinal hyperpermeability and NV during DR. Hyperglycemia is a major risk factor for endothelial cell activation and development of microvascular dysfunction during DR. Overwhelming evidences suggest that omega-3 PUFAs like DHA (C22:6n-3) are protective in proliferative DR. In the present study, we investigate the effect of exogenous supplementation of DHA on human retinal microvascular endothelial cells (HREC) under hyperglycemic conditions. Cells were treated with or without high glucose (HG, 25 mM) for 36 hrs in the presence or absence of the DHA (30 µM). To study the early inflammatory response, cells were analyzed for cell barrier function by measuring the changes in transcellular electrical resistance (TER) using the Electrical Cell-substrate Impedance Sensing (ECIS). Hyperglycemia resulted in a significant decline in TER while it was significantly improved by DHA. Furthermore, to investigate the effect of DHA on the proangiogenc properties of hyperglycemia in HREC a scratch-wound repair assay was carried out in cultured HREC cells treated with or without HG in the presence or absence of DHA. The migration and proliferation of cells induced by HG was reduced on supplementation of cells with DHA. Levels of apoptosis and oxidative stress were also monitored under the effect of hyperglycemia. While HG treatment did not result in any change in level of apoptosis, it resulted in sharp induction of oxidative stress in cells, which however was ameliorated under the effect of DHA treatment to hyperglycemic cells. Real-time PCR revealed significantly enhanced transcripts for the vascular endothelial growth factor (vegf) in hyperglycemic cells while reduced expression of the pigment epithelium derived factor (pedf) and the peroxisome proliferator activated receptor gamma (PPAR) was observed. These changes were prevented by DHA. Anti-inflammatory properties of DHA were further evinced by monitoring the levels of several inflammatory cytokines and angiogenic factors by multiplex enzyme-linked immunosorbent assay. Glucose treatment resulted in upregulation of various pro-angiogenic factors like angiogenin, angiopoietin, pdgf, pigf and this effect of HG was prevented by DHA treatment. Similarly, inflammatory cytokines like TNF-alpha and MCP1, which are involved in promotion of angiogenesis and inflammation were induced by HG and was attenuated in the presence of DHA. Interestingly, anti-inflammatory cytokines like IL-1 beta and IL-6 were markedly increased by DHA as compared to HG alone. These results suggest that DHA protects HREC against the pro-angiogenic, -inflammatory and -oxidative effects of HG and thus it can be proposed as alternative or supplementary treatment to the current therapeutic strategies of DR.