A proteomics study of MHC-associated peptide profiles displayed by macrophages after stimulation in vitro with human low density lipoproteins (78.12)

Abstract

Abstract Recognition of modified self proteins by macrophages usually induces the generation of peptide fragments derived cytoplasmic pool of cells that can be presented by the major histocompatibility complex (MHC). Despite the potential for presentation of a wide range of peptides only a limited number of peptides are displayed by the complexes. This generates peptide patterns that imprint a signature associated with particular cellular or disease processes. The aim of this study was to investigate whether stimulation of macrophages with human low density lipoprotein (LDL) generates peptide patterns on macrophage MHC class I and II in vitro. The study also assessed the effectiveness of peptide elution methods for peptide extraction. To accomplish this we stimulated J774.2 cells with native or oxidised LDL particles and extracted the MHC-associated peptides using citrate-phosphate buffer at pH 3, 4 & 5. Samples were then analysed by LC/MS/MS. MASCOT analysis was carried out to identify the protein source of peptides and IEDB and NetMHCII 1.0 were used to predict the peptide-binding affinity to MHC class I and II complexes. We identified 54 peptides derived from macrophages stimulated with ox-LDL, 33 peptides derived from cells treated with n-LDL and 10 peptides identified from untreated cells. Peptide derived from proteins involved in macrophage DNA and RNA replication, signal transduction, cytoskeleton and lipid metabolism were identified. We found a differential pattern of MHC-associated peptide expression in treated macrophages. These findings contribute to future identification of disease-associated peptides for their potential use in the development of therapeutic and/or diagnostic tools.