Application of Saturation Dye 2D-DIGE Proteomics to Characterize Proteins Modulated by Oxidized Low Density Lipoprotein Treatment of Human Macrophages

Abstract

Macrophages are believed to play a crucial role in atherogenesis and atherosclerotic plaque progression, mainly through their role in the accumulation of large amounts of cholesteryl ester and foam cell formation after the uptake into the arterial intima of oxidized LDL (oxLDL) particles known to be proatherogenic. The aim of this study was to use a differential proteomic approach to identify the response of human monocyte-derived macrophages after treatment with oxLDL for 24 h. Mass spectrometry analysis (MALDI-TOF) of 2D-DIGE gels made it possible to identify 9 intracellular and 3 secreted proteins that were up-regulated, 11 intracellular and 1 secreted proteins that were down-regulated, and 2 secreted proteins that were induced. This methodological approach not only confirmed the differential expression levels of proteins known to be regulated by oxLDL in macrophages, such as catalase and pyruvate kinase, but also identified oxLDL modulation of other proteins for the first time, including heat shock proteins (HSP) and Actin cytoskeletal proteins. Semiquantitative Western blot confirmed their role. The HSPs identified included heat shock cognate 71 kDa protein (Hsc70), 75 kDa glucose-regulated protein (GRP75), heat shock 70 kDa protein (Hsp70), and 60 kDa (Hsp60) proteins. These highly conserved intracellular protein chaperones, commonly seen in atherosclerotic plaques, appear to participate in protection against cellular stress. Interestingly, oxLDL also modulated several F-Actin capping proteins involved in Actin polymerization and motility: gelsolin, CapG, and CapZ. In conclusion, we have demonstrated the effects of oxLDL in the modulation of several proteins in human macrophages and established a functional profile of the human macrophage during the atherosclerotic process.

Nasal or Oral Immunization with GroEL Attenuates Porphyromonas gingivalis-Induced Atherosclerosis

“…The third, HSP60 was found to regulate some functions of oxidized LDL such as modulation of F-actin capping proteins involved in actin polymerization and macrophage motility (35). Since our previous studies also indicate that periodontal pathogens can accelerate atherosclerosis in Apoe shl mice by inducing HSP60 expression in aorta (36,37), it may say that circulating HSP 60 could play a role in chronic inflammation and could become target for therapeutic measures aiming at reducing the risk of atherogenic dyslipidemia and its complications.…”

P4‐276: Differential Expression Of Cell Surface Proteins In Human Bone Marrow Mesenchymal Stem Cells Cultured With Or Without Basic Fibroblast Growth Factor Containing Medium

“…Moreover, cytoskeletal proteins may serve as scaffolding for molecular signaling cascades, providing sites for localization, and anchoring of signaling molecules [62]. Actin-related proteins are particularly important in human macrophages, where they regulate actin filament length and concentration [63]. F-actin capping protein, also known as CapZ, is a heterodimer.…”

Transcriptomic and proteomic profiles of vascular cells involved in human abdominal aortic aneurysm

“…2D-DIGE was performed as recently described (Dupont et al, 2008 analysis was performed with Progenesis SameSpots v2.0 software. The differences in protein spots were then analyzed.…”

“…Four differentially expressed proteins were selected from 2D analysis of macrophages from the LILAS or an earlier study: HSP70, aconitase-1, GRP75 and beta-actin (Dupont et al, 2008). Four others were selected from microarray analysis, TIMP-3, ADAMTS10, fibronectin, and tenascin.…”