Expression analysis of novel striatal-enriched genes in Huntington disease

Selective degeneration of striatal neurons is a pathologic hallmark of Huntington disease (HD). The exact mechanism(s) behind this specific neurodegeneration is still unknown. Expression studies of diseased human post-mortem brain, as well as different mouse models exhibiting striatal degeneration, have demonstrated changes in the expression of many important genes with a large proportion of changes being observed in the striatal-enriched genes. These investigations have raised questions about how enrichment of particular transcripts in the striatum can lead to its selective vulnerability to neurodegeneration. Monitoring the expression changes of striatal-enriched genes during the course of the disease may be informative about their potential involvement in selective degeneration. In this study, we analyzed a Serial Analysis of Gene Expression (SAGE) database (www.mouseatlas.org) and compared the mouse striatum to 18 other brain regions to generate a novel list of striatal-enriched transcripts. These novel striatal-enriched transcripts were subsequently evaluated for expression changes in the YAC128 mouse model of HD, and differentially expressed transcripts were further examined in human post-mortem caudate samples. We identified transcripts with altered expression in YAC128 mice, which also showed consistent expression changes in human post-mortem tissue. The identification of novel striatal-enriched genes with altered expression in HD offers new avenues of study, leading towards a better understanding of specific pathways involved in the selective degeneration of striatal neurons in H

Transcriptional changes in Huntington disease identified using genome-wide expression profiling and cross-platform analysis

Evaluation of transcriptional changes in the striatum may be an effective approach to understanding the natural history of changes in expression contributing to the pathogenesis of Huntington disease (HD). We have performed genome-wide expression profiling of the YAC128 transgenic mouse model of HD at 12 and 24 months of age using two platforms in parallel: Affymetrix and Illumina. The data from these two powerful platforms were integrated to create a combined rank list, thereby revealing the identity of additional genes that proved to be differentially expressed between YAC128 and control mice. Using this approach, we identified 13 genes to be differentially expressed between YAC128 and controls which were validated by quantitative real-time PCR in independent cohorts of animals. In addition, we analyzed additional time points relevant to disease pathology: 3, 6 and 9 months of age. Here we present data showing the evolution of changes in the expression of selected genes: Wt1, Pcdh20 and Actn2 RNA levels change as early as 3 months of age, whereas Gsg1l, Sfmbt2, Acy3, Polr2a and Ppp1r9a RNA expression levels are affected later, at 12 and 24 months of age. We also analyzed the expression of these 13 genes in human HD and control brain, thereby revealing changes in SLC45A3, PCDH20, ACTN2, DDAH1 and PPP1R9A RNA expression. Further study of these genes may unravel novel pathways contributing to HD pathogenesis. DDBJ/EMBL/GenBank accession no: GSE1967

In vivo cell-autonomous transcriptional abnormalities revealed in mice expressing mutant huntingtin in striatal but not cortical neurons

Huntington's disease (HD), caused by a CAG repeat expansion in the huntingtin (HTT) gene, is characterized by abnormal protein aggregates and motor and cognitive dysfunction. Htt protein is ubiquitously expressed, but the striatal medium spiny neuron (MSN) is most susceptible to dysfunction and death. Abnormal gene expression represents a core pathogenic feature of HD, but the relative roles of cell-autonomous and non-cell-autonomous effects on transcription remain unclear. To determine the extent of cell-autonomous dysregulation in the striatum in vivo, we examined genome-wide RNA expression in symptomatic D9-N171-98Q (a.k.a. DE5) transgenic mice in which the forebrain expression of the first 171 amino acids of human Htt with a 98Q repeat expansion is limited to MSNs. Microarray data generated from these mice were compared with those generated on the identical array platform from a pan-neuronal HD mouse model, R6/2, carrying two different CAG repeat lengths, and a relatively high degree of overlap of changes in gene expression was revealed. We further focused on known canonical pathways associated with excitotoxicity, oxidative stress, mitochondrial dysfunction, dopamine signaling and trophic support. While genes related to excitotoxicity, dopamine signaling and trophic support were altered in both DE5 and R6/2 mice, which may be either cell autonomous or non-cell autonomous, genes related to mitochondrial dysfunction, oxidative stress and the peroxisome proliferator-activated receptor are primarily affected in DE5 transgenic mice, indicating cell-autonomous mechanisms. Overall, HD-induced dysregulation of the striatal transcriptome can be largely attributed to intrinsic effects of mutant Htt, in the absence of expression in cortical neuron

Cell loss in the motor and cingulate cortex correlates with symptomatology in Huntington's disease

Huntington's disease is an autosomal dominant inherited neurodegenerative disease with motor symptoms that are variably co-expressed with mood and cognitive symptoms, and in which variable neuronal degeneration is also observed in the basal ganglia and the cerebral cortex. We have recently shown that the variable symptomatology in Huntington's disease correlates with the variable compartmental pattern of GABAA receptor and cell loss in the striatum. To determine whether the phenotypic variability in Huntington's disease is also related to variable neuronal degeneration in the cerebral cortex, we undertook a double-blind study using unbiased stereological cell counting methods to determine the pattern of cell loss in the primary motor and anterior cingulate cortices in the brains of 12 cases of Huntington's disease and 15 controls, and collected detailed data on the clinical symptomatology of the patients with Huntington's disease from family members and clinical records. The results showed a significant association between: (i) pronounced motor dysfunction and cell loss in the primary motor cortex; and (ii) major mood symptomatology and cell loss in the anterior cingulate cortex. This association held for both total neuronal loss (neuronal N staining) and pyramidal cell loss (SMI32 staining), and also correlated with marked dystrophic changes in the remaining cortical neurons. There was also an association between cortical cell loss and striatal neuropathological grade, but no significant association with CAG repeat length in the Huntington's disease gene. These findings suggest that the heterogeneity in clinical symptomatology that characterizes Huntington's disease is associated with variation in the extent of cell loss in the corresponding functional regions of the cerebral cortex whereby motor dysfunction correlates with primary motor cortex cell loss and mood symptomatology is associated with cell loss in the cingulate corte

Decreased Striatal RGS2 Expression Is Neuroprotective in Huntington's Disease (HD) and Exemplifies a Compensatory Aspect of HD-Induced Gene Regulation

The molecular phenotype of Huntington's disease (HD) is known to comprise highly reproducible changes in gene expression involving striatal signaling genes. Here we test whether individual changes in striatal gene expression are capable of mitigating HD-related neurotoxicity.We used protein-encoding and shRNA-expressing lentiviral vectors to evaluate the effects of RGS2, RASD2, STEP and NNAT downregulation in HD. Of these four genes, only RGS2 and RASD2 modified mutant htt fragment toxicity in cultured rat primary striatal neurons. In both cases, disease modulation was in the opposite of the predicted direction: whereas decreased expression of RGS2 and RASD2 was associated with the HD condition, restoring expression enhanced degeneration of striatal cells. Conversely, silencing of RGS2 or RASD2 enhanced disease-related changes in gene expression and resulted in significant neuroprotection. These results indicate that RGS2 and RASD2 downregulation comprises a compensatory response that allows neurons to better tolerate huntingtin toxicity. Assessment of the possible mechanism of RGS2-mediated neuroprotection showed that RGS2 downregulation enhanced ERK activation. These results establish a novel link between the inhibition of RGS2 and neuroprotective modulation of ERK activity.Our findings both identify RGS2 downregulation as a novel compensatory response in HD neurons and suggest that RGS2 inhibition might be considered as an innovative target for neuroprotective drug development

Modulation of nucleosome dynamics in Huntington's disease

Transcriptional dysregulation and aberrant chromatin remodeling are central features in the pathology of Huntington's disease (HD). In order to more fully characterize these pathogenic events, an assessment of histone profiles and associated gene changes were performed in transgenic N171-82Q (82Q) and R6/2 HD mice. Analyses revealed significant chromatin modification, resulting in reduced histone acetylation with concomitant increased histone methylation, consistent with findings observed in HD patients. While there are no known interventions that ameliorate or arrest HD progression, DNA/RNA-binding anthracyclines may provide significant therapeutic potential by correcting pathological nucleosome changes and realigning transcription. Two such anthracyclines, chromomycin and mithramycin, improved altered nucleosome homeostasis in HD mice, normalizing the chromatin pattern. There was a significant shift in the balance between methylation and acetylation in treated HD mice to that found in wild-type mice, resulting in greater acetylation of histone H3 at lysine 9 and promoting gene transcription. Gene expression profiling in anthracycline-treated HD mice showed molecular changes that correlate with disease correction, such that a subset of downregulated genes were upregulated with anthracycline treatment. Improved nucleosomal dynamics were concurrent with a significant improvement in the behavioral and neuropathological phenotype observed in HD mice. These data show the ability of anthracycline compounds to rebalance epigenetic histone modification and, as such, may provide the rationale for the design of human clinical trials in HD patient

HTT-lowering reverses Huntington's disease immune dysfunction caused by NFκB pathway dysregulation

The peripheral immune response is altered in Huntington's disease, but the underlying mechanisms are unclear. Using RNA interference to lower huntingtin levels in leucocytes from patients, Träger et al. reverse disease-associated phenotypes including cytokine elevation and transcriptional dysregulation, and argue for a direct effect of mutant huntingtin on NFκΒ signallin

Cell loss in the motor and cingulate cortex correlates with symptomatology in Huntington's disease

Huntington's disease is an autosomal dominant inherited neurodegenerative disease with motor symptoms that are variably co-expressed with mood and cognitive symptoms, and in which variable neuronal degeneration is also observed in the basal ganglia and the cerebral cortex. We have recently shown that the variable symptomatology in Huntington's disease correlates with the variable compartmental pattern of GABA(A) receptor and cell loss in the striatum. To determine whether the phenotypic variability in Huntington's disease is also related to variable neuronal degeneration in the cerebral cortex, we undertook a double-blind study using unbiased stereological cell counting methods to determine the pattern of cell loss in the primary motor and anterior cingulate cortices in the brains of 12 cases of Huntington's disease and 15 controls, and collected detailed data on the clinical symptomatology of the patients with Huntington's disease from family members and clinical records. The results showed a significant association between: (i) pronounced motor dysfunction and cell loss in the primary motor cortex; and (ii) major mood symptomatology and cell loss in the anterior cingulate cortex. This association held for both total neuronal loss (neuronal N staining) and pyramidal cell loss (SMI32 staining), and also correlated with marked dystrophic changes in the remaining cortical neurons. There was also an association between cortical cell loss and striatal neuropathological grade, but no significant association with CAG repeat length in the Huntington's disease gene. These findings suggest that the heterogeneity in clinical symptomatology that characterizes Huntington's disease is associated with variation in the extent of cell loss in the corresponding functional regions of the cerebral cortex whereby motor dysfunction correlates with primary motor cortex cell loss and mood symptomatology is associated with cell loss in the cingulate cortex

A potent and selective Sirtuin 1 inhibitor alleviates pathology in multiple animal and cell models of Huntington's disease

Protein acetylation, which is central to transcriptional control as well as other cellular processes, is disrupted in Huntington's disease (HD). Treatments that restore global acetylation levels, such as inhibiting histone deacetylases (HDACs), are effective in suppressing HD pathology in model organisms. However, agents that selectively target the disease-relevant HDACs have not been available. SirT1 (Sir2 in Drosophila melanogaster) deacetylates histones and other proteins including transcription factors. Genetically reducing, but not eliminating, Sir2 has been shown to suppress HD pathology in model organisms. To date, small molecule inhibitors of sirtuins have exhibited low potency and unattractive pharmacological and biopharmaceutical properties. Here, we show that highly selective pharmacological inhibition of Drosophila Sir2 and mammalian SirT1 using the novel inhibitor selisistat (selisistat; 6-chloro-2,3,4,9-tetrahydro-1H-carbazole-1-carboxamide) can suppress HD pathology caused by mutant huntingtin exon 1 fragments in Drosophila, mammalian cells and mice. We have validated Sir2 as the in vivo target of selisistat by showing that genetic elimination of Sir2 eradicates the effect of this inhibitor in Drosophila. The specificity of selisistat is shown by its effect on recombinant sirtuins in mammalian cells. Reduction of HD pathology by selisistat in Drosophila, mammalian cells and mouse models of HD suggests that this inhibitor has potential as an effective therapeutic treatment for human disease and may also serve as a tool to better understand the downstream pathways of SirT1/Sir2 that may be critical for H

Regional and cellular gene expression changes in human Huntington's disease brain

Huntington's disease (HD) pathology is well understood at a histological level but a comprehensive molecular analysis of the effect of the disease in the human brain has not previously been available. To elucidate the molecular phenotype of HD on a genome-wide scale, we compared mRNA profiles from 44 human HD brains with those from 36 unaffected controls using microarray analysis. Four brain regions were analyzed: caudate nucleus, cerebellum, prefrontal association cortex [Brodmann's area 9 (BA9)] and motor cortex [Brodmann's area 4 (BA4)]. The greatest number and magnitude of differentially expressed mRNAs were detected in the caudate nucleus, followed by motor cortex, then cerebellum. Thus, the molecular phenotype of HD generally parallels established neuropathology. Surprisingly, no mRNA changes were detected in prefrontal association cortex, thereby revealing subtleties of pathology not previously disclosed by histological methods. To establish that the observed changes were not simply the result of cell loss, we examined mRNA levels in laser-capture microdissected neurons from Grade 1 HD caudate compared to control. These analyses confirmed changes in expression seen in tissue homogenates; we thus conclude that mRNA changes are not attributable to cell loss alone. These data from bona fide HD brains comprise an important reference for hypotheses related to HD and other neurodegenerative disease