Evidence for multiple mechanisms responsible for 2,5-hexanedione-induced neuropathy.
The present studies were carried out to investigate the comparative roles of protein cross-linking and alteration in protein phosphorylation in the accumulation of neurofilaments due to aliphatic hexacarbons. In these studies, rats were given 2,5-hexanedione (0, 0.1, 0.25 and 1.0%) for 70 days in their drinking water. In a separate study of in vitro protein phosphorylation rats were given 1% 2,5-hexanedione for 14 days in their drinking water. Spinal cord neurofilaments were isolated and analyzed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblotting using anti-neurofilament antibodies, radioimmunoassays (RIAs) of phosphorylated epitopes on neurofilament proteins and protein phosphorylation. Protein cross-linking of neurofilaments was found in all animals treated with 2,5-hexanedione including the lowest dose (0.1%) which did not produce clinical signs of intoxication. Protein phosphorylation of neurofilament proteins, as well as MAP-2 was significantly decreased upon treatment. Protein staining revealed a decreased amount of neurofilament protein and immunoblotting demonstrated neurofilament protein cross-linking in these animals. Protein staining of glial fibrillary acidic protein (GFAP) was unaltered by this treatment. RIAs of phosphorylated and non-phosphorylated epitopes of neurofilament proteins indicated that in vivo phosphorylation of these proteins was also decreased. Two-dimensional gel electrophoresis indicated a shift of the neurofilament proteins to a basic pI, indicating a dephosphorylation of neurofilament proteins. Cross-linked neurofilament proteins also exhibited a pI which was more basic than any of the individual neurofilament proteins. This report demonstrates differential effects of 2,5-hexanedione on neurofilament proteins and indicates that several mechanisms may be responsible for their accumulation.
Lapadula
D M
DM
Department of Pharmacology, Duke University Medical Center, NC 27710.
Suwita
E
E
Abou-Donia
M B
MB
D000818:Animals; D004305:Dose-Response Relationship, Drug; D006588:Hexanones / Q000009:adverse effects*; D007381:Intermediate Filament Proteins / Q000302:isolation & purification / Q000378:metabolism*; D007382:Intermediate Filaments / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D007659:Ketones / Q000009:adverse effects*; D008297:Male; D008970:Molecular Weight; D016900:Neurofilament Proteins; D009498:Neurotoxins; D010766:Phosphorylation; D051381:Rats; D011919:Rats, Inbred Strains; D012016:Reference Values; D013116:Spinal Cord / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*
D006588:Hexanones; D007381:Intermediate Filament Proteins; D007659:Ketones; D016900:Neurofilament Proteins; D009498:Neurotoxins; C011269:2,5-hexanedione