Carbon Monoxide Generation, Heme Turnover, and Lipid Peroxidation During Heme-Mediated LDL Oxidation: Association of HO-1, Hemopexin, and A1M with LDL

Heme promotes oxidative modification of low-density lipoprotein (LDL), a process implicated in atherosclerosis. However, the fate of heme during LDL oxidation and the mechanisms governing its degradation remain poorly understood. We investigated heme catabolism during heme/H2O2-induced LDL oxidation and identified LDL-associated proteins potentially involved in this process. Lipid peroxidation, heme Soret absorbance, and carbon monoxide (CO) production were assessed. Heme induced lipid peroxidation, loss of Soret signal, and CO generation, whereas CuSO4-mediated oxidation induced lipid peroxidation without detectable CO, supporting the association of CO production with heme-dependent oxidation. Targeted and data-independent proteomics and immunological analyses identified heme oxygenase-1 (HO-1), hemopexin (HPX), and alpha-1-microglobulin (A1M) in LDL. HO-1 inhibition reduced CO production and delayed early heme loss and lipid peroxidation, although these effects diminished over time. DTPA similarly delayed early heme loss and lipid peroxidation and reduced CO production, whereas BHT strongly suppressed lipid peroxidation, preserved the heme Soret signal, and reduced CO production throughout oxidation. HPX attenuated heme-induced LDL oxidation and CO generation. These findings indicate that HO-1-associated iron release contributes to early phase heme-mediated LDL oxidation along with CO generation, and heme-associated iron without liberation also contributes the oxidative modification of LDL.