Oxidative interactions between haemoglobin and membrane lipid. A liposome model

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Abstract
The relationship between Hb and membrane oxidation was studied using [human] liposomes containing Hb (hemosomes) as a red cell model. Rapid oxidation occurred in hemosomes formed from purified Hb and unsaturated lipid (egg phosphatidylcholines). After 3 h at 37.degree. C most of the Hb was oxidized, predominantly to methemoglobin with some hemichrome formation. The oxidation of Hb was paralleled by membrane lipid peroxidation as measured by thiobarbituric acid reactivity. These changes were largely abolished by using freshly prepared hemolysate instead of purified Hb, or when hemosomes were prepared with saturated phophatidylcholines. In hemosomes consisting of fresh hemolysate and saturated phosphatidylcholine, the rate of Hb oxidation at 37.degree. C corresponded to that of non-encapsulated hemolysate, and after 4 mo. storage at 4.degree. C 45% of HbO2 was oxidized. In hemosomes prepared from purified Hb and egg lecithin, .alpha.-tocopherol, catalase and ascorbate each protected against both hemoglobin oxidation and lipid peroxidation. Superoxide dismutase or reduced glutathione had no effect. In unsaturated-lipid hemosomes containing hemolysate, the rate of Hb oxidation increased when catalase was inhibited or reduced glutathione was depleted, but after long term incubation only concurrent catalase-inhibition and glutathione depletion could increase thiobarbituric acid reactivity. A close interdependence between Hb oxidation and lipid peroxidation was demonstrated and it showed that constituents of hemolysate strongly protect against both processes. H2O2 appears to be an important mediator, with its removal by either catalase or the glutathione/glutathione peroxidase system protecting against both oxidative changes.