Modification of cultured Madin-Darby canine kidney cells with dietary unsaturated fatty acids and regulation of arachidonate cascade reaction.

Abstract

Madin-Darby canine kidney (MDCK) cells were modified with dietary unsaturated fatty acids. The effects on the fatty acid composition in each phospholipid class and the formation of prostanoids upon stimulation were studied, from which the specificity of metabolism of individual unsaturated fatty acids and the regulation of arachidonate cascades in the modified cells were discussed. C18 unsaturated fatty acids were preferentially incorporated into phosphatidylcholine (PC) over phosphatidylethanolamine (PE), but arachidonic acid (20:4(n-6)) derived from gamma-linolenic acid (18:3(n-6)) was much more predominant in PE than PC. The fatty acid level in PE ranged from about 26-28% when the cells were modified with 20:4(n-6) or 5,8,11,14,17-eicosapentaenoic acid (20:5(n-3)), indicating the limitation of the storage of the eicosapolyenoic acids. The extra amounts appeared to be stored in PC. 18:3(n-6) was comparable to 20:4(n-6) to raise the level of 20:4(n-6) in PE, but not in PC which had half of 20:4(n-6) in PE. The supplementation of linoleic acid (18:2(n-6)). 18:3(n-6), and 20:4(n-6) caused significant increases in the synthesis of prostaglandin (PG)E2 up to almost the same levels when the modified cells were stimulated with 50 nM PMA and 100 nM A23187 for 24h. The cultured cells modified with eicosapolyenoic acids including 20:3(n-6), 20:4(n-6), and 20:5(n-3) were found to be inhibitory for the induction of PGE2 synthetic activity involving de nova synthesis of PG endoperoxide synthase, suggesting negative feedback regulation of the modified cells.