Cytoskeletal protein carbonylation and degradation in experimental autoimmune encephalomyelitis.
Protein carbonylation, the non-enzymatic addition of aldehydes or ketones to specific amino acid residues, has been implicated in the pathophysiology of multiple sclerosis. In this study, we investigated whether protein carbonyls also accumulate in the spinal cord of Lewis rats with acute experimental autoimmune encephalomyelitis (EAE). Western blots analysis after derivatization with dinitrophenyl hydrazine (oxyblot) showed elevated protein carbonylation at the time of maximal clinical disability. During the same period glutathione levels were substantially reduced, suggesting a causal relationship between these two markers. In contrast, lipid peroxidation products accumulated in EAE spinal cord well before the appearance of neurological symptoms. Carbonyl staining was not restricted to inflammatory lesions but present throughout the spinal cord particularly in neuronal cell bodies and axons. By 2-dimensional-oxyblot, we identified several cytoskeletal proteins, including beta-actin, glial acidic fibrillary protein, and the neurofilament proteins as the major targets of carbonylation. These findings were confirmed by pull-down experiments, which also showed an increase in the number of carbonylated beta-actin molecules and a decrease in that of oxidized neurofilament proteins in EAE. These data suggest the possibility that oxidation targets neurofilament proteins for degradation, which may contribute to axonal pathology observed in multiple sclerosis and EAE.
Smerjac
Suzanne M
SM
Department of Cell Biology and Physiology, University of New Mexico - Health Sciences Center, Albuquerque, New Mexico 87131, USA.
Bizzozero
Oscar A
OA
D000199:Actins / Q000378:metabolism; D000818:Animals; D001369:Axons / Q000378:metabolism / Q000473:pathology; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D004195:Disease Models, Animal; D004681:Encephalomyelitis, Autoimmune, Experimental / Q000378:metabolism* / Q000503:physiopathology; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D015227:Lipid Peroxidation / Q000502:physiology; D008297:Male; D009103:Multiple Sclerosis / Q000378:metabolism / Q000503:physiopathology; D009187:Myelitis / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology; D016900:Neurofilament Proteins / Q000378:metabolism; D009474:Neurons / Q000378:metabolism / Q000473:pathology; D018384:Oxidative Stress; D050050:Protein Carbonylation; D051381:Rats; D011917:Rats, Inbred Lew; D013116:Spinal Cord / Q000378:metabolism* / Q000503:physiopathology; D014855:Wallerian Degeneration / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology
D000199:Actins; D003598:Cytoskeletal Proteins; D005904:Glial Fibrillary Acidic Protein; D016900:Neurofilament Proteins; D005978:Glutathione