Altered Proteolytic Events in Experimental Autoimmune Encephalomyelitis Discovered by iTRAQ Shotgun Proteomics Analysis of Spinal cord

摘要

The identification of changes in regulated proteolytic events during experimental autoimmune encephalomyelitis (EAE) is crucial for uncovering activated proteases that may underline the pathological features such as inflammation and demyelination. We searched for either non-tryptic or semi-tryptic peptides indicating regulated proteolytic events from a shotgun proteomics study using isobaric tags for relative and absolute quantification (iTRAQ) to compare the proteomes of normal and EAE rat lumbar spinal cords. Several proteins such as alpha_(1)-macroglobulin, a protease inhibitor, (alpha)_(1)B-glycoprotein, (beta)_(2)-microglobulin, neurofilament light polypeptide and sulfated glycoprotein 1 had nontryptic peptide iTRAQ ratios that were significantly larger than the overall protein iTRAQ ratios, suggesting that such peptides may be markers for the proteolytic products generated by the protease(s) activated during EAE. Indeed, subsequent Western blot analysis confirmed the dysregulation of specific protein cleavages in EAE tissues.