Modification of red cell membrane lipids by hypochlorous acid and haemolysis by preformed lipid chlorohydrins

Summary

Hypochlorous acid (HOCI), a strong oxidant generated by the myeloperoxidase system of neutrophils and monocytes, has been implicated in inflammatory tissue damage by these cells. Reaction of HOCI with the double bonds of unsaturated lipids produces α, β-chlorohydrin isomers. We have expose red cell membranes to HOCI and used thin layer chromatography (TLC) of the extracted lipids and enzyme-linked immunosorbent assay (ELISA), using an antichlorohydrin monoclonal antibody, to show that fatty acyl chlorohydrins are formed. The ELISA was approximately 25 fold more sensitive than TLC, and chlorohydrins were detected when membranes from 10⁶ cells were treated with ≥ 0.16 nmoles HOCI. Lipid chlorohydrins are more polar and bulky than their parent lipids and as such could affect membrane stability and function. To determine the effect of incorporation of lipid chlorohydrins into cell membranes, preformed fatty acid and cholesterol chlorohydrins were incubated with red cells. Lysis was measured as release of haemoglobin and incorporation of lipids was determined by ¹⁴C scintillation counting. Addition of HOCI-treated oleic acid to red cells resulted in rapid lysis ofa fraction of the cells in a concentration dependent manner. HOCI-treated cholesterol also caused a small amount of cell lysis that was predominantly due to chlorohydrin 3, one of the three major cholesterol chlorohydrin products. Chlorohydrin 3, which has a decreased planarity and polarity, was also primarily responsible for altering the critical micelle concentration of HOCI-treated cholesterol-containing liposomes.