2,5-Hexanedione induced reduction in protein content and mRNA expression of neurofilament in rat cerebral cortex

Abstract

Exposure chronically to n-hexane produces central-peripheral axonopathy mediated by 2,5-hexanedione (HD). Studies have shown neurofilament (NF) subunit proteins are decreased substantially in cerebral cortices, optic axons, spinal cords, and sciatic nerves from HD-exposed rats. To deeply investigate the alterations in NF contents in HD neuropathy, the relative levels of NF-L, NF-M, and NF-H in rat cerebral cortex were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. HD was administrated to Wistar rats by intraperitoneal injection at dosage of 200 or 400 mg/kg. Rats were sacrificed after 6 weeks of treatment, and cerebral cortices were dissected, homogenized, and used for the determination of NF subunit proteins. The results, except for supernatant NF-L and NF-M that could not be assayed, showed HD intoxication resulted in significant decreases by 32–67% ( P < 0.01) in NF subunits in both of the pellet and supernatant fractions of cerebral cortex homogenate. As an initial investigation to determine how such changes in NF proteins might occur, the gene expression of NF-L, NF-M, and NF-H subunit mRNA was quantified using reverse transcription-polymerase chain reaction (RT-PCR). Statistical analysis revealed that HD exposure caused a significant reduction in the expression of NF-L and NF-H gene ( P < 0.05 or P < 0.01), while the levels of NF-M mRNA kept unaffected ( P > 0.05). These suggest that the observed reduction in NF gene expression might be related to diminished levels of subunit proteins, while the actual contribution might be uncertain. The functional significance of the reduced protein contents and the regulation of gene expression remain to be determined.