Melatonin inhibits oxidative modification of human low-density lipoprotein.

An important property of melatonin is that it is a free-radical scavenger or antioxidant. Since free radicals can induce oxidative modification of low-density lipoprotein (LDL), a process believed to be involved in atherogenesis, we were prompted to evaluate the capacity of melatonin to prevent oxidative modification of LDL. To induce oxidation, human LDL (0.4 mg protein/ml) was incubated at 37 degrees C with either 10 microM cupric chloride or 10 mM 2,2'-azo-bis-(2-amidinopropane) dihydrochloride (AAPH) for 3 hr or 24 hr, respectively. Several assays were then performed to unequivocally determine the extent of LDL oxidation. Compared to native LDL, oxidized LDL had increased agarose gel electrophoretic mobility and weaker immunoreactivity with a murine monoclonal antibody to human apolipoprotein B-100. Measurement of thiobarbituric acid-reactive substances (TBARS) revealed that native LDL contained 1.8 +/- 0.6 nmoles TBARS/mg protein, whereas copper-oxidized LDL contained 53 +/- 4 nmoles TBARS/mg protein. However, when present during incubation, melatonin (0.125-4 mM) inhibited in a concentration-dependent manner the increase in electrophoretic mobility, decrease in immunoreactivity of LDL, and increase in formation of TBARS caused by either copper or AAPH. In a fourth assay, phospholipid analysis of LDL was performed. Native LDL contained 420 +/- 9 nmoles phosphatidylcholine (PC)/mg LDL protein and 30 +/- 20 nmoles lysophosphatidylcholine (LysPC)/mg LDL protein. LDL incubated with copper had a decreased PC content (276 +/- 48 nmoles PC/mg LDL protein) and increased LysPC content (76 +/- 22 nmoles LysPC/mg LDL protein). But when present during the incubation of LDL with copper, melatonin attenuated in a concentration-dependent manner the degradation of PC to LysPC. Therefore, we conclude that melatonin can inhibit oxidative modification of LDL in vitro.