Oxidative damage to proteins in the spinal cord in amyotrophic lateral sclerosis (ALS).
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease, which has been linked to the generation of free radicals and oxidative stress. Oxidative damage to spinal cord proteins is suggested to be a contributory factor to neuronal death in ALS. Since proteins are the major targets for free radicals and the so-called "reactive species", therefore the objective of our study was to identify oxidatively damaged spinal cord proteins. The material consisted of spinal cords of 8 sporadic ALS cases and 5 controls. We estimated the level of protein carbonyl moieties, which react quantitatively with 2,4-dinitrophenylhydrazine (DNPH). Afterwards proteins were separated by SDS-polyacrylamide gel electrophoresis and the protein bound DNPH moieties were detected immunochemically. We also morphologically examined spinal cords after immune staining against DNPH. The protein carbonyl content of the ALS spinal cords significantly increased in all examined cases. In most ALS patients, proteins with 125 kDa, 70 kDa and 36kDa were highly oxidized. The 70-kDa protein was identified immunochemically to be neurofilament 68. The morphological examination of ALS spinal cords indicated a pronounced anti-DNPH immune reaction in neurones of the anterior horns; the reaction in the posterior horns was less intense. Microglia in the white matter was immunoreactive; astroglia was DNPH-negative. Although the exact mechanism by which reactive oxygen species induce motor neurones to die is not known yet, the presented data indicate that they affect spinal cord cellular proteins, including neurofilament 68. In this study, we successfully examined the neurochemical features accompanying motor neuron injury in ALS, and the results may help to develop a rationale anti-oxidative neuroprotective strategy.
Niebrój-Dobosz
Irena
I
Medical Research Centre, Polish Academy of Sciences, Warszawa, Poland. neuromyol@cmdik.pan.pl
Dziewulska
Dorota
D
Kwieciński
Hubert
H
D000368:Aged; D000369:Aged, 80 and over; D000690:Amyotrophic Lateral Sclerosis / Q000378:metabolism*; D001253:Astrocytes / Q000378:metabolism; D015153:Blotting, Western; D016923:Cell Death / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D017628:Microglia / Q000378:metabolism; D008875:Middle Aged; D009419:Nerve Tissue Proteins / Q000032:analysis* / Q000378:metabolism; D016900:Neurofilament Proteins / Q000378:metabolism; D009474:Neurons / Q000378:metabolism; D010084:Oxidation-Reduction; D010659:Phenylhydrazines; D017382:Reactive Oxygen Species; D013116:Spinal Cord / Q000737:chemistry* / Q000378:metabolism*
D009419:Nerve Tissue Proteins; D016900:Neurofilament Proteins; D010659:Phenylhydrazines; D017382:Reactive Oxygen Species; C066858:neurofilament protein NF 68; C004787:2,4-dinitrophenylhydrazine