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. 2021 Oct 4;129(11):1039–1053. doi: 10.1161/CIRCRESAHA.121.319272

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© 2021 The Authors.

Circulation Research is published on behalf of the American Heart Association, Inc., by Wolters Kluwer Health, Inc. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution, and reproduction in any medium, provided that the original work is properly cited.

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Figure 5. — Integrated proteomics and lipidomics analysis of HDL (high-density lipoprotein) in patients with coronary artery disease (CAD). A, PCSK9 (proprotein convertase subtilisin/kexin type 9) distribution across lipoprotein fractions within a cohort of 172 patients with varying CAD-related phenotypes was assessed using a modified sandwich ELISA. Significance was determined using the Kruskal-Wallis test across the groups. B, ucHDL (ultracentrifuge-isolated HDL) from these patients with CAD (n=191, including samples from patients with 6 mo follow-up) was analyzed by quantitative proteomics. The coefficients of variation in HDL protein abundances, as measured by label-free mass spectrometry (MS), were calculated across the whole cohort. C, PCSK9 protein correlations against the core HDL proteome are represented as a volcano plot. D, Quantitation by label-free and tandem-mass tag (TMT) proteomics upon ucHDL revealed proteins that were altered by sex (males, n=98. females n=66). Significant proteins in at least one method are labeled; fold changes across methodologies were compared using a linear regression analysis. E, Three hundred sixty-five lipid species were quantified in HDL by targeted MS with reference standards. A Spearman correlation matrix was generated between the sum of each lipid species in a respective class and the HDL apolipoprotein profile, as well as PCSK9, PLTP, and CLU. A hierarchical cluster analysis is represented as a heat map. F, Crosslinking mass spectrometry (XLMS) analysis of immunoHDL (immuno-isolated HDL) and ucHDL revealed a strong protein-protein interaction between apoA2 and apoC1, green lines represent crosslinks (xiview.org). AC indicates acylcarnitines; ANTXR1, anthrax toxin receptor 1; C4B, complement factor 4B; CE, cholesterol esters; CER, ceramides; CLU, clusterin; DG, diacylglycerides; LBP, lipopolysaccharide-binding protein; LPC, lyso-phosphatidylcholine; PC, phosphatidylcholine; PCYOX, prenylcysteine oxidase; PLTP, phospholipid transfer protein; PON, paraoxonase; RLU, relative light units; SAA, serum amyloid A; SFTPB, surfactant protein B; SM, sphingomyelins; and TG, triglycerides.