No abstract

Background In Huntington's disease (HD), alterations in energy metabolism and mitochondrial function may result in oxidative stress. Increased levels of oxidative damage products have been reported in HD. The transcriptional co-activator PGC-1 α stimulates mitochondrial biogenesis and induces expression of many reactive oxygen species (ROS) eliminating enzymes including SOD1, SOD2, catalase and glutathione peroxidase (Gpx1). In addition, it was shown by two independent research groups that PGC-1 α transcription is changed in HD. Aims Here we analysed the PGC-1 α dependent ROS stress response in two different mouse models for HD. Methods/techniques The expression levels of ROS detoxifying enzymes in brain and skeletal muscle of 12-week-old R6/2 and WT mice and in 1-year-old HdhQ20/ HdhQ111 knock-in mice were determined by qPCR. The response to oxidative stress stimuli (H2O2, glutamate) were studied using primary neurons of HdhQ20 and HdhQ111. To detect a failure of ROS stress response induction, primary neurons were treated with oxidative stressors and cells were harvested after different time points of recovery for mRNA isolation and qPCR expression analysis of ROS detoxifying enzymes. Results/outcome We found that PGC-1 α, PGC-1β, catalase, SOD1 and SOD2 were downregulated in R6/2 brain. Furthermore, PGC-1 α was also downregulated in the HdhQ111 knock-in mouse brain, but no changes in the oxidative stress response system was found. In contrast, in muscle tissue of R6/2, Pgc-1 α expression levels were unchanged while catalase was upregulated and SOD1, SOD2 and GPx1 were downregulated. In skeletal muscle of HdhQ111, PGC-1 α and PGC-1 β were upregulated and all other tested genes were unchanged. HdhQ20 and HdhQ111 primary neurons which were exposed to glutamate did not show any significant induction of the ROS detoxifying system. It is possible that the culturing conditions with regular air oxygen concentrations might have resulted in an ROS stress response and a subsequent failure of a further stress response induction being measurable. Experiments are repeated with 3% oxygen culturing conditions. Conclusions In summary, we found more pronounced alterations of the ROS defence system in the R6/2 mouse compared with the HdhQ111 knock-in mouse. Alterations of the ROS detox system differed between skeletal muscle and brain tissue.

Background Several observations indicate that mitochondrial dysfunction plays an important role in the pathogenesis of Huntington's disease (HD). HD patients lose significant body weight despite normal or increased food intake and impairment of ATP synthesis occurs even in pre-motor manifest HD expansion mutation carriers. Alterations in mitochondrial function and structure include impairment of respiratory chain activity, decrease of Ca2+ buffering capacity and an increase in size of mitochondria. Until now the molecular mechanisms linking mutant huntingtin to mitochondria dysfunction are not known. Aim Purify mitochondria from brain and skeletal muscle of two HD mouse models and analyse the proteome using a two dimensional differential in-gel electrophoresis (2D-DIGE) approach. Methods/techniques To get a more detailed picture of mitochondrial changes in HD we isolated mitochondria of brain and skeletal muscle of the exon-1 R6/2 mouse model and of the knock-in mouse models HdhQ20/HdhQ111. First, protocols for the isolation of mitochondria using subcellular fractionation/differential centrifugation were optimised and the purity of the mitochondrial fraction was confirmed by western blot. Mitochondrial lysates were then used for proteome analysis using a 2D-DIGE approach. Results In mitochondria of R6/2 mouse brains, mitochondrial proteins of the citric acid cycle, the amino acid degradation and mitochondria fusion were increased. Few selected mitochondrial proteins were downregulated. These results will be further corroborated by western blot and compared with changes in skeletal muscle and to proteome analysis of mitochondria from brain and skeletal muscle in the HdhQ knock-in mice. Conclusions These first results of a mitochondrial proteome analysis on mitochondria from R6/2 mouse brains demonstrating an increase of certain mitochondrial proteins involved in energy metabolism suggests that the observed alterations reflect at least in part compensatory changes.

No abstract

No abstract