Huu Phuc Nguyen

Overexpression of human wildtype torsinA and human ΔGAG torsinA in a transgenic mouse model causes phenotypic abnormalities

Primary torsion dystonia is an autosomal-dominant inherited movement disorder. Most cases are caused by an in-frame deletion (GAG) of the DYT1 gene encoding torsinA. Reduced penetrance and phenotypic variability suggest that alteration of torsinA amino acid sequence is necessary but not sufficient for development of clinical symptoms and that additional factors must contribute to the factual manifestation of the disease. We generated 4 independent transgenic mouse lines, two overexpressing human mutant torsinA and two overexpressing human wildtype torsinA using a strong murine prion protein promoter. Our data provide for the first time in vivo evidence that not only mutant torsinA is detrimental to neuronal cells but that also wildtype torsinA can lead to neuronal dysfunction when overexpressed at high levels. This hypothesis is supported by (i) neuropathological findings, (ii) neurochemistry, (iii) behavioral abnormalities and (iv) DTI-MRI analysis.

Cholesterol Defect Is Marked across Multiple Rodent Models of Huntington's Disease and Is Manifest in Astrocytes

Brain cholesterol, which is synthesized locally, is a major component of myelin and cell membranes and participates in neuronal functions, such as membrane trafficking, signal transduction, neurotransmitter release, and synaptogenesis. Here we show that brain cholesterol biosynthesis is reduced in multiple transgenic and knock-in Huntingtons disease (HD) rodent models, arguably dependent on deficits in mutant astrocytes. Mice carrying a progressively increased number of CAG repeats show a more evident reduction in cholesterol biosynthesis. In postnatal life, the cholesterol-dependent activities of neurons mainly rely on the transport of cholesterol from astrocytes on ApoE-containing particles. Our data show that mRNA levels of cholesterol biosynthesis and efflux genes are severely reduced in primary HD astrocytes, along with impaired cellular production and secretion of ApoE. Consistently, in CSF of HD mice, ApoE is mostly associated with smaller lipoproteins, indicating reduced cholesterol transport on ApoE-containing lipoproteins circulating in the HD brain. These findings indicate that cholesterol defect is robustly marked in HD animals, implying that strategies aimed at selectively modulating brain cholesterol metabolism might be of therapeutic significance.

Sex differences in a transgenic rat model of Huntington's disease: decreased 17β-estradiol levels correlate with reduced numbers of DARPP32+ neurons in males

Recent clinical studies have highlighted that female sex hormones represent potential neuroprotective mediators against damage caused by acute and chronic brain diseases. This evidence has been confirmed by experimental studies documenting the protective role of female sex hormones both in vitro and in vivo, although these studies did not specifically focus on Huntingtons disease (HD). We therefore investigated the onset and course of HD in female and male transgenic (tg) HD (CAG(n51)) and control rats across age and focused on three aspects: (i) behavioral and physiological alterations (energy expenditure, home-cage activity, emotional disturbance and motor dysfunction), (ii) morphological markers (numbers and characteristics of striatal DARPP32(+) medium-sized spiny neurons (MSNs) and dopamine receptor autoradiography) and (iii) peripheral sex hormone levels as well as striatal estrogen receptor expression. Independent of their sex, tgHD rats exhibited increased levels of food intake, elevated home-cage activity scores and anxiolytic-like behavior, whereas only males showed an impairment of motor function. In line with the latter finding, loss and atrophy of DARPP32(+) MSNs were apparent only in male tgHD rats. This result was associated with a decreased striatal dopamine D1 receptor density and lower plasma levels of 17beta-estradiol at the age of 14 months. As DARPP32(+) MSNs expressed both alpha- and beta-estrogen receptors and showed a correlation between cell numbers and 17beta-estradiol levels, our findings suggest sex-related differences in the HD phenotype pointing to a substantial neuroprotective effect of sex hormones and opening new perspectives on the therapy of HD.

Localization of sequence variations in PGC-1α influence their modifying effect in Huntington disease

BackgroundHuntington disease (HD) is caused by a polyglutamine expansion of more than 35 units in the huntingtin protein. This expanded repeat length inversely correlates with the age-at-onset (AAO), however, additional genetic factors apart from the expanded CAG repeat size are thought to influence the course and the AAO in HD. Until now, among others, the gene encoding PCG-1α (PPARGC1A) was shown to modify the AAO in two independent, however small, populations. PGC-1α is involved in the induction of various mechanisms regulating mitochondrial biogenesis and oxidative stress defence. Furthermore, several studies have linked impairment of its function and/or its expression to HD pathogenesis. As the identification of distinct modifiers in association studies is largely dependent on the size of the observed population, we investigated nine different single nucleotide polymorphisms (SNPs) in PPARGC1A in order to replicate the disease modifying effect in more than 800 European HD patients and to identify an association with AAO in HD.ResultsTwo SNPs, one in the promoter and one in the transcribed region of the gene, showed a significant effect on the AAO. While the minor allele of SNP rs7665116 (g.38570C), located in the transcribed gene region, was associated with a delay in disease onset, especially in HD patients with Italian ancestry, the minor allele of SNP rs2970870 (g.-1437C) in the promoter region leads to an earlier onset of HD in its homozygous state. Additionally, global testing of haplotype block 2, which covers the main part of the transcribed region of the gene, revealed an association between block 2 haplotypes and the disease onset.ConclusionTherefore, our results indicate opposing modifying influences of two SNPs within one gene on AAO and support the idea that PGC-1α dysfunction is involved in HD pathology.

Selective striatal neuron loss and alterations in behavior correlate with impaired striatal function in Huntington's disease transgenic rats

Huntingtons disease (HD) is an inherited neurodegenerative disorder characterized by selective striatal neuron loss and motor, cognitive and affective disturbances. The present study aimed to test the hypothesis of adult-onset neuron loss in striatum and frontal cortical layer V as well as alterations in behavior pointing to impaired striatal function in a recently developed transgenic rat model of HD (tgHD rats) exhibiting enlarged ventricles, striatal atrophy and pycnotic pyramidal cells in frontal cortical layer V. High-precision design-based stereological analysis revealed a reduced mean total number of neurons in the striatum but not in frontal cortical layer V of 12-month-old tgHD rats compared with age-matched wild-type controls. No alterations in mean total numbers of striatal neurons were found in 6-month-old animals. Testing 14-month-old animals in a choice reaction time task indicated impaired striatal function of tgHD rats compared with controls.

Stem Cell Quiescence in the Hippocampal Neurogenic Niche Is Associated With Elevated Transforming Growth Factor-β Signaling in an Animal Model of Huntington Disease

Cellular proliferation, differentiation, integration, and survival within the adult neural stem cell niche are altered under pathological conditions, but the molecular cues regulating the biology of this niche are mostly unknown. We examined the hippocampal neural stem cell niche in a transgenic rat model of Huntington disease. In this model, progressive cognitive deficits develop at the age of 9months, suggesting possible hippocampal dysfunction. We found a disease-associated progressive decline in hippocampal progenitor cell proliferation accompanied by an expansion of the pool of 5-bromo-2-deoxyuridine label-retaining Sox-2-positive quiescent stem cells in the transgenic animals. Increments in quiescent stem cells occurred at the expense of cAMP-responsive element-binding protein-mediated neuronal differentiation and survival. Because elevated levels of transforming growth factor-&bgr;1 (TGF-&bgr;1) impair neural progenitor proliferation, we investigated hippocampal TGF-&bgr; signaling and determined that TGF-&bgr;1 induces the neural progenitors to exit the cell cycle. Although phospho-Smad2, an effector of TGF-&bgr; signaling, is normally absent in subgranular stem cells, it accumulated progressively in Sox2/glial fibrillary acidic protein-expressing cells of the subgranular zone in the transgenic rats. These results indicate that alterations in neurogenesis in transgenic Huntington disease rats occur in successive phases that are associated with increasing TGF-&bgr; signaling. Thus, TGF-&bgr;1 signaling seems to be a crucial modulator of neurogenesis in Huntington disease and may represent a target for future therapy.

A Novel BACHD Transgenic Rat Exhibits Characteristic Neuropathological Features of Huntington Disease

Huntington disease (HD) is an inherited progressive neurodegenerative disorder, characterized by motor, cognitive, and psychiatric deficits as well as neurodegeneration and brain atrophy beginning in the striatum and the cortex and extending to other subcortical brain regions. The genetic cause is an expansion of the CAG repeat stretch in the HTT gene encoding huntingtin protein (htt). Here, we generated an HD transgenic rat model using a human bacterial artificial chromosome (BAC), which contains the full-length HTT genomic sequence with 97 CAG/CAA repeats and all regulatory elements. BACHD transgenic rats display a robust, early onset and progressive HD-like phenotype including motor deficits and anxiety-related symptoms. In contrast to BAC and yeast artificial chromosome HD mouse models that express full-length mutant huntingtin, BACHD rats do not exhibit an increased body weight. Neuropathologically, the distribution of neuropil aggregates and nuclear accumulation of N-terminal mutant huntingtin in BACHD rats is similar to the observations in human HD brains. Aggregates occur more frequently in the cortex than in the striatum and neuropil aggregates appear earlier than mutant htt accumulation in the nucleus. Furthermore, we found an imbalance in the striatal striosome and matrix compartments in early stages of the disease. In addition, reduced dopamine receptor binding was detectable by in vivo imaging. Our data demonstrate that this transgenic BACHD rat line may be a valuable model for further understanding the disease mechanisms and for preclinical pharmacological studies.

Age at onset in Huntington’s disease is modified by the autophagy pathway: implication of the V471A polymorphism in Atg7

Huntington’s disease (HD) is caused by an expansion of a polyglutamine repeat of more than 35xa0units in the huntingtin protein. The expanded repeat length is inversely correlated with the age at onset (AAO); however, additional genetic factors apart from the expanded CAG repeat length can modify the course and the AAO in HD. Aberrations in macroautophagy have been observed in Huntington, Alzheimer, Parkinson, motor neuron and prion diseases. Therefore, we hypothesized that polymorphisms in autophagy-related (Atg) genes might contribute to the variation in the AAO. We initially tested eight single nucleotide polymorphisms in five Atg genes (Atg3, Atg5, Atg7, Atg16L1 and Beclin-1) for their frequency of ≥1%. Subsequently, we investigated the polymorphisms Atg7 V471A and Atg16L1 T281A for a disease-modifying effect in more than 900 European HD patients (including 2 populations consisting of 346 German patients and 327 patients of Italian descent). One polymorphism in the Atg7 gene that substitutes alanine for valine (V471A) showed a significant effect on the AAO (Pxa0=xa00.0050) and was associated with an earlier disease onset of 4xa0years. Our results further support the important pathophysiological role of autophagy in HD.

Progressive deterioration of reaction time performance and choreiform symptoms in a new Huntington's disease transgenic ratmodel

We tested the hypothesis that a recently developed transgenic rat model of Huntingtons disease (tgHD rats) showed an age-and genotype-dependent change in psychomotor performance and in the frequency of choreiform movements similar to HD patients. Wild type and tgHD (homozygotic and heterozygotic) rats were behaviorally tested at an age of 15 and 20 months. Our results show that tgHD rats exhibit an age-, and genotype-dependent deterioration of the psychomotor performance and choreiform symptoms, closely mimicking the clinical time course changes of psychomotor symptoms of HD patients. These data provide further experimental evidence that the tgHD rat can be considered as a relevant animal model of HD.

Behavioral and In Vivo Electrophysiological Evidence for Presymptomatic Alteration of Prefrontostriatal Processing in the Transgenic Rat Model for Huntington Disease

Cognitive decline precedes motor symptoms in Huntington disease (HD). A transgenic rat model for HD carrying only 51 CAG repeats recapitulates the late-onset HD phenotype. Here, we assessed prefrontostriatal function in this model through both behavioral and electrophysiological assays. Behavioral examination consisted in a temporal bisection task within a supra-second range (2 vs.8 s), which is thought to involve prefrontostriatal networks. In two independent experiments, the behavioral analysis revealed poorer temporal sensitivity as early as 4 months of age, well before detection of overt motor deficits. At a later symptomatic age, animals were impaired in their temporal discriminative behavior. In vivo recording of field potentials in the dorsomedial striatum evoked by stimulation of the prelimbic cortex were studied in 4- to 5-month-old rats. Input/output curves, paired-pulse function, and plasticity induced by theta-burst stimulation (TBS) were assessed. Results showed an altered plasticity, with higher paired-pulse facilitation, enhanced short-term depression, as well as stronger long-term potentiation after TBS in homozygous transgenic rats. Results from the heterozygous animals mostly fell between wild-type and homozygous transgenic rats. Our results suggest that normal plasticity in prefrontostriatal circuits may be necessary for reliable and precise timing behavior. Furthermore, the present study provides the first behavioral and electrophysiological evidence of a presymptomatic alteration of prefrontostriatal processing in an animal model for Huntington disease and suggests that supra-second timing may be the earliest cognitive dysfunction in HD.