Proteomic analysis of the secretome of human umbilical vein endothelial cells using a combination of free‐flow electrophoresis and nanoflow LC‐MS/MS

Abstract

Human umbilical vein endothelial cells are the most widely used in vitro model for endothelial cells. Their secreted proteins, however, have not been comprehensively analysed so far. In this study, we accomplished to map the secretome of human umbilical vein endothelial cells by combining free-flow electrophoresis with nanoflow LC-MS/MS. This comprehensive analysis provides a basis for future comparative studies of protein secretion by endothelial cells in response to cardiovascular risk factors and is available on our website http://www.vascular-proteomics.com.

Plasma MicroRNA Profiling Reveals Loss of Endothelial MiR-126 and Other MicroRNAs in Type 2 Diabetes

“…29,30 HUVECs exposed to high glucose (25 mmol/L) were cultured in complete medium for 6 days. As a control, HUVECs were cultured in complete medium (5 mmol/L glucose) supplemented with mannitol, to exclude effects of osmotic stress.…”

Proteomics

“…In a shotgun proteomics analysis of the secretome from human umbilical vein endothelial cells, 12% to 15% of all identified spectra corresponded to albumin peptides. 42 In contrast, SMCs tolerate serum starvation well and can be kept in serum-free medium for longer. In this setting, a shotgun proteomics strategy was able to mine deeper into the secretome and detected all cytokines present at concentrations Ͼ10 ng/mL (of 27 tested in a multiplex assay).…”

Overview of Current Proteomic Approaches for Discovery of Vascular Biomarkers of Atherosclerosis

“…In this respect, cell culture systems could represent a useful tool to partially overcome drawbacks of tissue analyses, allowing researchers to study single aspects of the atherosclerotic process in very controlled conditions. In the last years, studies on proteome (the intracellular proteins) and secretome (the proteins released into the cell culture medium) by 2DE and MS of ECs (Bruneel et al, 2003;Chen et al, 2007;Tunica et al, 2009), VSMCs (McGregor et al, 2001Dupont et al, 2005;Lee et al, 2006), and monocytes/macrophages (Dupont et al, 2004;Fach et al, 2004;Slomianny et al, 2006;Zhang et al, 2007;Zhao et al, 2009) have been performed. Moreover, in the attempt to help in elucidating the mechanisms of atherogenesis, several proteomic studies have been carried out on vascular cells cultured in different experimental conditions (table 3).…”

“…Very few studies applied gel free proteomic approaches such as LC MS/MS (Fach et al, 2004;X.L. Wang et al, 2007;Zhao et al, 2009;Tunica et al, 2009;Zimman et al, 2010) and microarrays (Sukhanov et al, 2005).…”