Lipoproteomic Methodology And Its Application In The Analysis Of High Density Lipoproteins From Type 2 Diabetes And Coronary Heart Disease

The lipoproteins are a set of spherical particles in blood, which containing a hydrophobic core of neutral lipid (cholesteryl ester and triglycerides) and a surface rich in amphipathic proteins, phospholipids, and free cholesterol. Based on the relative contents of lipid and protein, the lipoprotein particles can be divided into five major classes, including high density lipoprotein (HDL), low density lipoprotein (LDL), intermediate density lipoprotein (IDL), very low density lipoprotein (VLDL) and Chylomicrons(CM). Lipoproteins play vital roles in extracellular lipid package, storage, transport, and metabolism. Therefore, dysfunction of lipoproteins metabolism has been implicated in numerous human disease, such as cancer, neurological disorders, diabetes and others. High density lipoprotein, the smallest but heaviest particles, exhibit anti-atherogenic properties for its function in reverse cholesterol transport, antioxidant, anti-inflammation, anti-thrombosis and protection of vascular endothelial cells. However, accumulating evidences have revealed that HDL levels are not positively related with their functions in the patients with atherosclerosis, metabolic syndrome, chronic inflammation or immune system disease. Moreover, HDL could be remodeled into dysfunctional HDL or pro-inflammatory HDL, which may lose its beneficial role against atherosclerosis, even exhibit pro-atherogenic effect, such as pro-inflammatory and pro-oxidation. The dynamic changes of protein composition in HDL particles are believed to be associated to such process. However, the precise mechanism remained to be elucidated. In current study, we have developed a novel method for the HDL particles isolation. By determining the diameter and homogeneity of the HDL particle isolated, we found that the isolation of purified and homogenous HDL particles can be achieved by our method. Combined with proteomic approaches, our methods present a powerful tool for the research of the relationship between the dynamic changes of protein composition in HDL with its function in different diseases.To explore the relationship between the dynamic changes of protein composition in HDL with the development of diabetes and coronary artery disease, HDL particles from four groups of populations, including normal control, type 2 diabetes, coronary artery disease and type 2 diabetes coupled with coronary artery disease, were isolated by our method and subjected to LC-MS/MS based label-free proteomic analysis. In the present study, we used label-free based quantitative proteomic approach to identify and quantify 105 unique proteins in HDL from four groups with high confidence. To the end,105 proteins were identified and quantified with high confidence from the HDL particles isolated from four groups mentioned above.33 out Of these proteins were found to be changed significantly, which could made up eight differentially changing clusters. The GO analysis showed that these proteins function in multiple anti- or pro-atherosclerosis processes, such as vasoconstriction, endothelium injury, thrombosis, and pro-inflammation. These results suggest that the dynamic changes of proteins that exhibit anti-or pro-atherogenic properties are associated with the dysfunction of the HDL during the development of type 2 diabetes or coronary artery diseases. Our results could provide an important knowledge to better understand the proteome remodeling in HDL dysfunction and may improve the diagnosis and treatment evaluation of type 2 diabetes and coronary artery diseases.