Abstract

Huntington's disease (HD) is an autosomal dominant neurodegenerative disease caused by elongation of a CAG-repeat within the first exon of the huntingtin gene. This mutation leads to a toxic gain-of-function of the huntingtin protein (htt). The exact mechanism of HD pathogenesis remains elusive, but it is thought that proteolytic cleavage of the mutant htt protein is an important step in HD pathogenesis. However, studies involving htt cleavage fragments in human brain tissue could be complicated by non-disease specific degradation of the htt protein during post-mortem delay. To elucidate the effects of post-mortem delay, we have conducted a study using human HD caudate nucleus tissue and human temporal lobe tissue as control with low initial post-mortem delays (3 and 1 h resp). To mimic post-mortem delay, specimens were brought to room-temperature and every 2 h samples were taken for a minimum of 8 h. Analysis of these samples was performed by Western-blotting using an antibody that recognises the first 17 amino acids. For both brain regions, the majority of fragments did not change between time points, apart from fragments at 52 kD and 70 kD which increased over time. Only in the HD caudate nucleus specimen, we observed several htt fragments between 80 to 100 kD that decreased over time. We conclude that post-mortem delay only has moderate effects. Next, we analysed interpersonal differences between the sensory/motor cortex and caudate nucleus region from nine different HD and control subjects. First results on Western blotting indicate that there are no striking differences between HD and controls for the sensory/motor cortex. For the HD caudate nucleus however, we observed an increase in protein fragments compared to the control samples. These initial results suggest that there is a regional variation in htt protein fragmentation in the human brain which may be related to pathogenesis.