TMT-based quantitative proteomic profiling of human monocyte-derived macrophages and foam cells

Yali Zhang,Liang Bai,Enqi Liu,Sihai Zhao,Rong Wang,Chenyang Zhang,Yu Fu,Weirong Wang,Wenbin Cao,Linying Jia,Naqash Alam

doi:10.1186/s12953-021-00183-x

Abstract

BackgroundCardiovascular diseases remain the leading cause of morbidity and mortality worldwide, most of which are caused by atherosclerosis. Discerning processes that participate in macrophage-to-foam cell formation are critical for understanding the basic mechanisms underlying atherosclerosis. To explore the molecular mechanisms of foam cell formation, differentially expressed proteins were identified.MethodsHuman peripheral blood mononuclear cells were stimulated with macrophage colony-stimulating factor, and obtained macrophages were transformed into foam cells by oxidized low-density lipoprotein. Tandem mass tag (TMT) labeling combined with mass spectrometry was performed to find associations between foam cell transformation and proteome profiles.ResultsTotally, 5146 quantifiable proteins were identified, among which 1515 and 182 differentially expressed proteins (DEPs) were found in macrophage/monocyte and foam cell/macrophage, respectively. Subcellular localization analysis revealed that downregulated DEPs of macrophages/monocytes were mostly located in the nucleus, whereas upregulated DEPs of foam cells/macrophages were mostly extracellular or located in the plasma membrane. Functional analysis of DEPs demonstrated that cholesterol metabolism-related proteins were upregulated in foam cells, whereas immune response-related proteins were downregulated in foam cells. The protein interaction network showed that the DEPs with the highest interaction scores between macrophages and foam cells were mainly concentrated in lysosomes and the endoplasmic reticulum.ConclusionsProteomics analysis suggested that cholesterol metabolism was upregulated, while the immune response was suppressed in foam cells. KEGG enrichment analysis and protein-protein interaction analysis indicated that DEPs located in the endoplasmic reticulum and lysosomes might be key drivers of foam cell formation. These data provide a basis for identifying the potential proteins associated with the molecular mechanism underlying macrophage transformation to foam cells.

Highlights

Cardiovascular diseases remain the leading cause of morbidity and mortality worldwide, most of which are caused by atherosclerosis
Before testing by mass spectrometry, monocytes were purified with CD14 microbeads as CD14 is a monocyte surface marker, and macrophages were identified by flow cytometry using CD11b antibodies
The results showed that oxidized low-density lipoproteins (oxLDL)-induced cells, stained with oil red O, presented red lipid droplets (Fig. 1C)

Summary

Introduction

Cardiovascular diseases remain the leading cause of morbidity and mortality worldwide, most of which are caused by atherosclerosis. Discerning processes that participate in macrophage-to-foam cell formation are critical for understanding the basic mechanisms underlying atherosclerosis. Despite extensive efforts to study its pathogenesis and develop effective drugs, atherosclerosis remains the leading cause of mortality and disability worldwide. Atherosclerosis is characterized by the accumulation of lipids and fibrous elements in the arterial intima [2]. Foam cells are key components of atherosclerotic plaque and play an important role in all atherosclerotic lesions, from the earliest fatty streak formations to the most advanced atheromas. In the early stages of atherosclerosis, foam cell accumulation in the arterial wall forms fatty streaks, the first sign of atherosclerosis, which is visible without magnification [3]. In the last phase of atherosclerosis, foam cells can become necrotic and are encapsulated by a thin fibrous cap that can rupture, resulting in thrombosis and vessel occlusion [4]

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