Carbamylated-Oxidized LDL: Proatherosclerotic Effects on Endothelial Cells and Macrophages

Abstract

Aim

Both oxidized LDL and carbamylated LDL are considered important for initiating atherosclerosis in patients with end-stage kidney disease through vascular endothelial cell dysfunction or injury. However their effects on each other and their relationship related to pro-atherosclerotic effects on endothelial cells and macrophages have not been investigated. In this study, we analyzed the competition between LDL carbamylation and oxidation, tested biological effects of carbamylated-oxidized LDL (coxLDL) toward the endothelial cells, assessed its ability to cause foam cell development, and determined the roles of scavenger receptors in this process.

Methods

Cross-competition between carbamylation and oxidation of LDL particles was tested using cell-free fluorescent ligand-receptor assay. Pro-atherogenic properties (cell proliferation, cytotoxicity, and foam cell formation) of all LDL isoforms were tested in vitro and ex vivo using endothelial cells and peritoneal macrophages. In addition, coxLDL was assessed in human sera and in vivo atherosclerotic plaques which were developed in mouse model of uremia-induced atherosclerosis.

Results

Our data suggest that there is potential competition between carbamylation and oxidation of LDL, and that oxidation is a much stronger inhibitor of carbamylation than vice versa. coxLDL is highly cytotoxic to endothelial cells and strongly induce their proliferation measured by DNA synthesis. All three tested LDL isoforms demonstrated strong ability for transformation of primary mouse peritoneal macrophages to foam cells using predominantly CD36 scavenger receptor. coxLDL was the most potent inducer of foam cell development and macrophages/foam cell injury assessed by cell count and TUNEL, respectively. Finally, LDL particles modified by oxidation and carbamylation were detected in blood and shown to co-localize in atherosclerotic plaques in mice.

Conclusion

Our study demonstrated that LDL particles can be simultaneously carbamylated and oxidized and modifications are likely coexisting in the same LDL particle. We also demonstrated pro-atherosclerotic properties of coxLDL and proposed its role in atherosclerosis.