DOCOSAHEXAENOIC ACID IS A MAJOR N-3 POLYUNSATURATED FATTY-ACID IN BOVINE RETINAL MICROVESSELS

The aim of this study was to purify microvessels from bovine retina an d also to cultivate bovine retinal endothelial cells (BRECs) or intram ural pericytes, to determine their fatty acid composition, Microvessel s were obtained after Dounce homogenization of the retina followed by centrifugation on albumin cushion and finally microvessels in the pell et were trapped on a 100-mu m nylon filter. Contamination of microvess el preparations by neuronal tissue, assessed after both microscopic ex amination and western blotting with a monoclonal antibody raised again st rhodopsin, was minor. In the entire bovine retina, docosahexaenoic acid (DHA) represented 23.3% of the total fatty acids and there was ab out three limes less arachidonic acid (AA) (8.2%) than DHA. In contras t, DHA and AA levels were almost equivalent in the retinal microvessel s with similar to 10% of total fatty acids. When compared with intact microvessels, the DHA proportion of confluent monolayers of both BRECs or pericytes in primary cultures dropped to similar to 2% of the tota l fatty acids, whereas AA was unchanged, Culture medium supplementatio n with unesterified DHA (10 mu M) restored the DHA proportion of BRECs close to the microvascular value at the expense of linoleic acid with out affecting AA very much. In contrast, DHA supplementation in pericy tes increased the DHA proportion of these cells at the expense of AA. In conclusion, DHA of intact microvessels represented 10% of the total fatty acids, which was close to the AA proportion. Mild DHA supplemen tation of BRECs or pericytes in primary cultures restored their DHA pr oportion to the original microvessel value. This high percentage of po lyunsaturated fatty acids in retinal microvessels should allow us to t est the hypothesis that oxidation products derived from these fatty ac ids may be involved in the pathogenic process leading to diabetic reti nopathy.