Hp Nguyen

Cellular and subcellular localization of Huntington aggregates in the brain of a rat transgenic for Huntington disease

Huntington disease (HD) is a progressive neurodegenerative disorder characterized by emotional, cognitive, and motor dysfunctions. Aggregation of huntingtin is a hallmark of HD and, therefore, a crucial parameter for the evaluation of HD animal models. We investigated here the regional, cellular, and subcellular distribution of N‐terminal huntingtin aggregates and associated neuropathological changes in the forebrain of a rat transgenic for HD (tgHD). The tgHD rat brain showed enormously enlarged lateral ventricles and a similar atrophy of cortical and subcortical areas as known in HD patients. Huntingtin aggregates of varying size and forms were regionally identified in neuronal nuclei, cytoplasm, dendrites, dendritic spines, axons, and synaptic terminals, closely resembling the results described earlier for human HD brains and in established HD mouse models. Huntingtin aggregates in mitochondria support mitochondrial dysfunction as contributing to the disease pathogenesis. Dark cell degeneration was reminiscent of results in HD individuals and HD mouse models. Interestingly, huntingtin aggregates were especially well accumulated in two interacting limbic forebrain systems, the ventral striatopallidum and the extended amygdala, which may contribute to the early onset of emotional changes observed in the tgHD rat. In conclusion, the tgHD rat model reflects to a remarkable extent the cellular and subcellular neuropathological key features as observed in human HD and HD mouse brains and hints of changes in limbic forebrain systems, which may elucidate the emotional dysfunction in the tgHD rat and affective disturbances in HD patients. J. Comp. Neurol. 501:716–730, 2007.

Neuropeptide Y Y1 receptor-mediated anxiolysis in the dorsocaudal lateral septum: functional antagonism of corticotropin-releasing hormone-induced anxiety.

Neuropeptide Y and corticotropin-releasing hormone are involved in the regulation of various physiological functions including the expression of anxiety and fear. The anxiogenic effects of corticotropin-releasing hormone can be modulated by neuropeptide Y, yet the brain regions involved in this interaction are only partly understood. By utilizing antibodies raised against neuropeptide Y and the Y1 receptor protein we identified a densely labeled cell group in the dorsal zone of caudal part of the rat lateral septum. Bilateral microinjections of neuropeptide Y into the dorsocaudal lateral septum but not into the intramedial septum dose-dependently decreased anxiety in the social interaction test of rats, whereas the effects of corticotropin-releasing hormone were opposite. The anxiogenic-like effect of corticotropin-releasing hormone was reversed by neuropeptide Y pretreatment. Local microinjection of the neuropeptide Y receptor selective antagonists revealed that neither Y1 receptor nor Y2 receptor selective antagonists had effects on experimental anxiety on their own suggesting that neuropeptide Y-induced anxiolysis is not tonic. The Y1 receptor antagonist blocked the anxiolytic-like effect of neuropeptide Y, while the Y2 receptor antagonist was ineffective.We conclude that neuropeptide Y in the dorsocaudal lateral septum may act as an endogenous anxiolytic and antagonize corticotropin-releasing hormone (stress)-induced anxiety. This functional antagonism probably shapes behavior under aversive conditions, as neuropeptide Y-induced anxiolysis is not tonic in nature. An imbalance between these two neuropeptide systems in the septum may lead to a maladaptive expression of anxiety after stress exposure.

Huntingtin-associated protein-1 is a modifier of the age-at-onset of Huntington's disease

A polyglutamine repeat expansion of more than 36 units in a protein called huntingtin (htt) is the only known cause of Huntingtons disease (HD). The expanded repeat length is inversely correlated with the age-at-onset (AAO), however, the onset age among HD patients with CAG repeats below 60 units varies considerably. In addition to environmental factors, genetic factors different from the expanded CAG repeat length can modify the AAO of HD. We hypothezised that htt interacting proteins might contribute to this variation in the AAO and investigated human htt-associated protein-1 (HAP1) using genetic and functional assays. We identified six polymorphisms in the HAP1 gene including one that substitutes methionine (M441) for threonine (T441) at amino acid 441. Analyzing 980 European HD patients, we found that patients homozygous for the M441 genotype show an 8-year delay in the AAO. Functional assays demonstrated that human M441-HAP1 interacts with mutant htt more tightly than does human T441-HAP1, reduces soluble htt degraded products and protects against htt-mediated toxicity. We thus provide genetic and functional evidence that the M441-HAP1 polymorphism modifies the AAO of HD.

Personalized peptide vaccine-induced immune response associated with long-term survival of a metastatic cholangiocarcinoma patient

Graphical abstract

Genetic Rodent Models of Huntington Disease

The monogenic nature of Huntington disease (HD) has led to the development of a spectrum of useful genetically modified models. In particular, rodents have pioneered as the first HD model being generated and have since been the most widely used animal model for HD in both basic research and preclinical therapeutic studies. Based on the generation strategies, these rodent models can be classified into 3 major groups, the transgenic fragment models, the transgenic full-length models and the knock-in models. These models display a range of HD-like characteristics which resemble the clinical symptoms of HD patients. Their applications in research are thus regarded as an invaluable approach to speeding up the unraveling of the underlying pathological mechanisms of HD and for finding a disease-modifying treatment for this devastating disease. In this chapter, the similarities and differences of the most commonly used rodent HD models and their relevance to human HD will be compared and discussed. This also serves to guide the selection of an appropriate rodent HD model according to the nature of investigation.

B21 Characterisation Of Mice Overexpressing Mutant Huntingtin Lacking The Spine Morphogenesis Regulator Kalirin-7

Background Kalirin-7, a postsynaptic density protein which interacts with the huntingtin-associated-protein-1, has several common interactors with huntingtin as PSD95 and NMDAR2B. Kalirin-7 is important for the generation and maintenance of synaptic dendrites and glutamatergic synapses. Glutamatergic N-methyl-D-aspartate receptor (NMDAR)-mediated excitotoxicity has been implicated in Huntington’s disease (HD) pathophysiology for a long time. But the mechanisms mediating NMDAR toxicity in HD are still not clear. Aim In this study,we investigated the effect of kalirin-7 deficiency in a HD mouse model Methods Kalirin-7 knockout (Kal7KO) mice were crossbred with a transgenic huntington mouse model (YAC128) carrying a YAC construct with the fulllength human huntingtin with 128CAG repeats. Male mice underwent a behavioural characterisation including activity, motor learning, cognition and anxiety testing. Protein biochemistry and histology were used to assess effects of kalirin-7-deficiency on huntingtin aggregation, glutamate receptor subunit levels and synapse morphology in cortex and striatum. Results Kalirin-7 deficiency does not affect the motor performance of YAC128 mice on the rotarod nor does it influence activity in an automated home cage system. Interestingly, Kalirin-7 deficiency reduces anxiety of YAC128 on the elevated plus maze. Spatial learning, memory or relearning in YAC128 mice in a simple swimming test are not modulated by the lack of Kalirin-7. On protein level, Kalirin-7 deficiency leads to the upregulation of larger Kalirin isoforms in YAC128 mice whereas levels of NMDAR subunits are not affected. The aggregate load of YAC128 mice is not changed by lack of kalirin-7. Moreover, the absence of Kalirin-7 does not change nor cause a difference in synapse morphology of YAC128 mice. Conclusions Kalirin-7 deficiency specifically influences the anxiety phenotype of YAC128 mice but does not affect aggregate load or NMDAR toxicity linked parameters.

C06 Motor, Emotional And Cognitive Phenotyping Of Bachd Rats

Background Huntington disease (HD) is an inherited neurodegenerative disorder caused by an expanded CAG repeat in the huntingtin (HTT) gene. Patients affected by HD are impaired in motor and cognitive abilities and present neuropsychiatric symptoms. BACHD rats are a model of HD expressing the full length mutant human HTT under the control of the human HTT promoter.AimThe purpose of this study was to phenotype BACHD rats for anxiety, cognition and motor functionality. Methods BACHD rats and WT controls, between 1 and 6 months of age, were subjected to open field (OF), elevated plus maze (EPM), passive avoidance (PA), and rotarod tests. Body weight measurements were taken once a month. Results BACHD rats, compared to controls, showed: decreased time spent in the inner area as well as lower number of zone transitions in OF at 1 and 3 months; decreased distance and speed in OF at 3 months; alterations in EPM behaviour at 4 months; unchanged emotional and associative learning at 6 months and progressively impaired motor coordination starting from month 2. Body weight did not change throughout the experiment. Conclusions The impairment in motor function as well as the behavioural alterations indicates that BACHD rats may be a model representative of the human disease.

B05 New Light On The Role Of Calpain-mediated Proteolysis Of Mutant Huntingtin

Background Calpains (CAPNs), a class of intracellular calcium-dependent cysteine proteases, belong to the group of proteolytic enzymes which have been associated with the cleavage of mutant huntingtin (Htt), the source for toxic fragments of the disease protein. As excitation-induced calcium influx occurring in neurons can activate CAPNs and excitotoxicity represents an important factor in the pathomechanism of Huntington’s disease (HD), these proteases might explain the neuronal determination of the disease. Aims To improve our knowledge on the involvement of CAPNs in the molecular mechanism of HD pathogenesis by examining the impact of different members of the CAPN system and by dissecting linked activation pathways. Methods The CAPN system and proteolytic fragmentation of Htt were analysed in different in vitroand in vivomodels of HD via protein biochemical and calcium imaging techniques. Results We observed an over-activation of the CAPN system in cell and mouse models of HD. Activation of CAPNs in cell-free and cell-based assays promoted the fragmentation of wild type and mutant Htt generating specific N- and C-terminal fragments. Conversely, suppressing CAPN activity using specific inhibitors improved the viability of STHdh Q111 cells. These results stress the importance of CAPNs on mutant Htt cleavage and disease development. Conclusions Our enhanced understanding of calpain-mediated proteolysis as a key event in the molecular pathogenesis of HD, will promote the quest for new therapeutical targets as a treatment for this fatal disease.

C10 Generation and characterisation of a transgenic rat model of huntington's disease-like 4 (HDL4), also called spinocerebellar ataxia type 17 (SCA17)

Huntingtons disease-like 4 (HDL4), also called spinocerebellar ataxia type 17, is an autosomal-dominant, late-onset neurodegenerative disorder caused by CAG repeat expansions in the TATA-box-binding protein (TBP), an ubiquitously expressed transcription factor. The clinical features of HDL4 closely resembled those of Huntingtons disease, including uncontrolled movements, emotional problems, and loss of thinking ability. To further investigate this devastating disease and additionally find a good model for treatment studies, we generated the first transgenic rat model of HDL4, which carries a full human cDNA fragment of the TBP gene with 64 CAG repeats under the control of the murine prion protein promoter (PrP-TBP64Q). In total we obtained ten positive founder animals, whereof only five transmitted the transgene. On the basis of the expression level of mutant TBP as well as the distribution of mutant TBP in the different brain regions we chose one of these five lines (line 8.4) for further characterisation. This line shows a strong expression of the mutant TBP protein in the cerebellum and a moderate expression in the olfactory bulb, brainstem and cortex. HDL4 rats showed an ataxia-like phenotype and impaired motor coordination and balance capabilities in the beam walking test. Additionally, HDL4 rats had a significantly lower body weight than their wildtype littermates and showed decreased activity. At the age of 10 months neuropathological changes, such as misshaped Purkinje cells, degenerated dendrites as well as missing basket and stellate neurons were observed in the cerebellum of heterozygous transgenic HDL4 rats compared to wildtype littermates. By electron microscopy we demonstrated dark cell degeneration as well as moderate fibre degeneration in the striatum. Moreover, we observed decreased levels of dopamine receptor (D2R) by performing PET imaging. Altogether the present data shows that this animal model exhibits symptoms which mimic well the human HDL4 phenotype indicating that this rat model will be of great value for further studies of pathomechanisms in Huntingtons disease-like 4 and for therapeutic trials.

E04 Autophagy-related gene Atg7 is a genetic modifier of AAO specific in the Italian population

Background The only known cause of Huntingtons disease (HD) is a polyglutamine repeat expansion of more than 36 units in the huntingtin protein, which is inversely correlated with the age-at-onset (AAO) of the disease. However, additional genetic factors apart from the expanded CAG repeat length can modify the course and the AAO in HD. Since autophagy, the major way for the degradation of mutant huntingtin, is thought to exert influence on the pathogenesis of HD, we hypothesised that autophagy-related (Atg) genes might contribute to the variation in the AAO. Aim/method In this regard, we analysed the association of the V471A polymorphism in the autophagy-related gene Atg7 gene in two large cohorts (1st study group: 952 patients; 2nd study group (EHDN REGISTRY): 1464 patients), composed of HD patients descending from different European countries. Results Although a polymorphism in the Atg7 gene that substitutes alanine for valine (V471A) showed a significant effect on the AAO in the first patient cohort mainly consisting of Italian and German HD patients, the significant effect of the Atg7 V471A polymorphism could not be confirmed in the second study group that was composed of European patients other than Italian and German patients. A more detailed analysis revealed a significant effect of the Atg7 V471A polymorphism on the AAO especially in the Italian population (327 patients) and was associated with a 4 years earlier disease onset. Conclusion Therefore, we identified a genetic modifier for HD in relationship to the autophagic pathway, but with a specific effect in a single population. This result affirms the influence of genetic modifiers on the course of HD, but also suggests population-specific modifying mechanisms in the HD pathogenesis.