Influence of cumene hydroperoxide and 4-hydroxynonenal on the glutathione metabolism during in vitro ageing of human skin fibroblasts

Abstract

Cumene hydroperoxide (Chp) and 4-hydroxynonenal (HNE) were used to investigate the effect of peroxidative challenge upon the glutathione (GSH) metabolism of human skin fibroblasts. Cellular GSH contents decreased during short-term incubations with Chp and oxidised glutathione (GSSG) was formed concomitantly. During longer incubations the GSH level was restored and the substrate flux through the pentose phosphate shunt increased. So in the presence of hydroperoxides the GSH level is maintained by reduction of GSSG.HNE caused a strong decrease in cellular GSH contents. Prolonged incubation with HNE lead to a rise in GSH contents above the basal level. The flux through the pentose phosphate shunt did not change during exposure to HNE. Hence, during incubation with HNE the cell maintains its GSH content by de novo synthesis of GSH. This conclusion is further substantiated by the findings with a cell strain deficient in GSH synthetase. These cells survived if incubated with Chp but not if exposed to HNE.GSH contents of normal cells from phase 11 (young) cultures and from phase I11 (aged) cultures responded similarly to Chp during short-term incubations and during a week of culture with the test compound. The flux through the pentose phosphate shunt rose much more in phase 111 than in phase I1 cells when incubated with the same concentration series of Chp. We conclude that during in vitro ageing the amount of NADPH needed to maintain cellular GSH levels in the presence of hydroperoxides increases, while the capacity to respond to such a challenge is not affected.The phenomenon of cells ageing in culture was first described by Hayflick and Moorhead [l] and has been reproduced in many different systems [2 -41. To explain this phenomenon two types of theory have been put forward: one invokes an underlying genetic programme [5] while the other presumes stochastically arising damage to be the basic cause. One of the main hypotheses of the second type is the freeradical theory of ageing [6]. Free-radical reactions, for instance those arising upon the one-electron reduction of oxygen 171, are presumed to damage the cell. One of the major effects of free-radical reactions in the cell is lipid peroxidation [8]. Cellular aerobic metabolism gives rise to several species of reactive oxygen, including superoxide, hydroperoxides and hydroxyl radicals [9, 101, against which the cell has defence mechanisms such as the glutathione system [ll]. During in vitro ageing the level of GSH remains almost constant [12, 131. While GSH is equally available in all cell organelles, glutathione reductase and glutathione peroxidase activities decrease considerably during in vitro ageing [13]. On the other Correspondence to M.

Diabetes Accelerates Smooth Muscle Accumulation in Lesions of Atherosclerosis

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Cytotoxicity of diatom-derived oxylipins in organisms belonging to different phyla

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Glucose Flux through the Hexose Monophosphate Shunt and NADP(H) Levels during in vitro Ageing of Human Skin Fibroblasts

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