Lipid peroxidation contributes to hydrogen peroxide induced cytotoxicity in renal epithelial cells

Abstract

We have examined the role of lipid peroxidation in the cytotoxicity of H2O2 in OK cells containing markedly differing amounts of cell membrane polyunsaturated fatty acids (PUFA). In OK cells grown in a serum free medium, PUFA were undetectable. The membranes of these cells contained predominantly oleic, stearic and palmitic acids. When cultured in medium containing 10% calf serum, OK cells contained measurable amounts of PUFA [linoleic (5 +/- 1%) and arachidonic acids (8 +/- 1%)]. When the serum containing medium was supplemented with 60 mM linoleic acid, the membrane content of both linoleic (21 +/- 1%) as well as arachidonic acid (15 +/- 1%) as substantially increased. The severity of injury induced by H2O2 in OK cells was substantially altered by the PUFA content of the cell membrane. Exposure of OK cells to 1.25 mM H2O2 for one hour resulted in more cell death (determined by a trypan blue assay) in cells grown in serum supplemented with linoleic acid with "normal" PUFA content (90 +/- 2%) than in cells with "reduced" levels of PUFA grown in unsupplemented calf serum (81 +/- 3%). Cells gown in defined, serum free medium with undetectable levels of PUFA suffered the least H2O2-induced lethal cell injury (47 +/- 8%). Comparable differences in the cytotoxicity of H2O2 among cells with differing PUFA content were found using a clonogenic assay of cell viability. Malondialdehyde (MDA) accumulation induced by 1.25 mM H2O2 was greater in cells with "normal" PUFA content (702 +/- 103 pM/microgram cell DNA/hr) than in cells with "reduced" PUFA (328 +/- 112 pM/100 microgram DNA/hr) and was undetectable in cells grown in defined, serum free medium. In summary, the content of PUFA of cells in culture is profoundly influenced by culture conditions. Our data provide novel and direct evidence that peroxidation of cell membranes contributes directly to the severity of cell injury and death induced by H2O2.