Modification with homocysteine does not increase susceptibility of human low-density lipoprotein to iron-mediated oxidation

Abstract

Oxidation of human low-density lipoprotein (LDL) is centrally involved in the development of cardiovascular diseases. This study investigated whether homocysteine-mediated thiolation of LDL rendered it more susceptible to oxidation by iron. After in vitro exposure to homocysteine thiolactone for 60 minutes, LDL's thiol content increased from 26 +/- 5 (control) to 224 +/- 20 nmol/mg of protein (thiolated; P < .0001). Control and thiolated LDL (0.2 mg of protein per milliliter) were incubated with either redox active iron (Fe(3+); 10 micromol/L) or, as a positive control, copper (Cu(2+); 10 micromol/L). Consistent with the observation of others, thiolation decreased Cu(2+)-dependent formation of lipid oxidation products in LDL (17 +/- 16 nmol/mg of protein formed in thiolated LDL, compared with 81 +/- 21 nmol/mg of protein in control, during 6 hours of incubation; P < .01). Thiolation had no effect, however, on Fe(3+)-mediated oxidation of LDL with lipid oxidation products remaining essentially nondetectable during prolonged incubation (up to 48 hours). Thiolation similarly had no effect on oxidation of LDL (0.2 mg of protein per milliliter) by heme-complexed iron (hemin; 10 micromol/L), with lipid oxidation products increasing to 24 +/- 1 and 27 +/- 4 nmol/mg of protein for control and thiolated LDL, respectively, during 6 hours of incubation (P > .05). Similar results were observed using LDL with varying degrees of thiolation (29 +/- 5, 85 +/- 14, 130 +/- 15, and 213 +/- 19 nmol of thiol per milligram of protein). In conclusion, these results demonstrate that thiolation has no effect on LDL's susceptibility to iron-mediated oxidation.