4-Hydroxy-2-nonenal induces mitochondrial dysfunction and aberrant axonal outgrowth in adult sensory neurons that mimics features of diabetic neuropathy

Modification of proteins by 4-hydroxy-2-nonenal (4-HNE) has been proposed to cause neurotoxicity in a number of neurodegenerative diseases, including distal axonopathy in diabetic sensory neuropathy. We tested the hypothesis that exposure of cultured adult rat sensory neurons to 4-HNE would result in the formation of amino acid adducts on mitochondrial proteins and that this process would be associated with impaired mitochondrial physiology and axonal function. In addition, we compared 4-HNE-induced axon pathology with that exhibited by neurons isolated from diabetic rats. Cultured adult rat dorsal root ganglion (DRG) sensory neurons were incubated with varying concentrations of 4-HNE. Cell survival, axonal morphology and level of axon outgrowth were assessed. In addition, video microscopy of live cells, Western blot and immunofluorescent staining were utilized to detect protein adduct formation by 4-HNE and to localize actively respiring mitochondria. 4-HNE induced formation of protein adducts on cytoskeletal and mitochondrial proteins, and impaired axon regeneration by approximately 50% at 3 μM whilst having no effect on neuronal survival. 4-HNE initiated formation of aberrant axonal structures and caused the accumulation of mitochondria in these dystrophic structures. Neurons treated with 4-HNE exhibited a distal loss of active mitochondria and treatment of mitochondrial preparations from cortex induced a significant and acute reduction in the rate of electron transport. Finally, the distal axonopathy and the associated aberrant axonal structure generated by 4-HNE treatment mimicked axon pathology observed in DRG sensory neurons isolated from diabetic rats and replicated aspects of neurodegeneration observed in human diabetic sensory neuropathy.