Large-sized Fetal Striatal Grafts in Huntington’s Disease Do Stop Growing: Long-term Monitoring in the Florence Experience

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Development of six large nodules of solid tissue after bilateral human fetal striatal transplantation in four Huntington’s disease patients has raised concern about the safety of this experimental therapy in our setting. We investigated by serial MRI-based volumetric analysis the growth behaviour of such grafts. After 33-73 months from transplantation the size of five grafts was stable and one graft showed a mild decrease in size. Signs neither of intracranial hypertension nor of adjuctive focal neurological deficit have ever been observed. This supports long-term safety of the grafting procedure at our Institution.

Differential Synaptic and Extrasynaptic Glutamate-Receptor Alterations in Striatal Medium-Sized Spiny Neurons of Aged YAC128 Huntington’s Disease Mice

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Huntington’s disease (HD) is a late-onset, slowly progressing neurodegenerative disorder caused by an expansion of glutamine repeats. The YAC128 mouse model has been widely used to study the progression of HD symptoms, but little is known about synaptic alterations in very old animals. The present experiments examined synaptic properties of striatal medium-sized spiny neurons (MSNs) in 16 month-old YAC128 mice. These mice were crossed with mice expressing enhanced green fluorescent protein (EGFP) under the control of either D1 or D2 dopamine receptor promoters to identify MSNs originating the direct and indirect pathways, respectively. The input-output curves of evoked excitatory postsynaptic currents mediated by activation of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) or N-methyl-D-aspartate (NMDA) receptors were reduced in MSNs in both pathways. In the presence of DL-threo-β-Benzyloxyaspartic acid (DL-TBOA), a glutamate transporter blocker used to increase activation of extrasynaptic receptors, NMDA receptor-mediated currents displayed altered amplitudes, longer decay times, and greater charge (response areas) in both direct and indirect pathway MSNs in YAC128 mice compared to wildtype controls. Amplitudes were significantly increased, primarily in direct pathway MSNs while normalized areas were significantly increased only in indirect pathway MSNs, suggesting that the two types of MSNs are affected in different ways. It may be that indirect pathway neurons are more susceptible to changes in glutamate transport. Taken together, the present findings demonstrate differential alterations in synaptic versus extrasynaptic NMDA receptors in both direct and indirect pathway MSNs in late HD, which may contribute to the dysfunction and degeneration in both pathways.

Aggregation-Prone Proteins Modulate Huntingtin Inclusion Body Formation in Yeast

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Huntington’s disease (HD) is a fatal neurodegenerative disorder caused by a polyglutamine expansion in the huntingtin (HTT) protein. The expression of mutant HTT in the baker’s yeast Saccharomyces cerevisiae recapitulates many of the cellular phenotypes observed in mammalian HD models. Mutant HTT aggregation and toxicity in yeast is influenced by the presence of the Rnq1p and Sup35p prions, as well as other glutamine/asparagine-rich aggregation-prone proteins. Here we investigated the ability of a subset of these proteins to modulate mutant HTT aggregation and to substitute for the prion form of Rnq1p. We find that overexpression of either the putative prion Ybr016wp or the Sup35p prion restores aggregation of mutant HTT in yeast cells lacking the Rnq1p prion. These results indicate that an interchangeable suite of aggregation-prone proteins regulates mutant HTT aggregation dynamics in yeast, which may have implications for mutant HTT aggregation in human cells.

Polyglutamine Disease Modeling: Epitope Based Screen for Homologous Recombination using CRISPR/Cas9 System

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We have previously reported the genetic correction of Huntington’s disease (HD) patient-derived induced pluripotent stem cells using traditional homologous recombination (HR) approaches. To extend this work, we have adopted a CRISPR-based genome editing approach to improve the efficiency of recombination in order to generate allelic isogenic HD models in human cells. Incorporation of a rapid antibody-based screening approach to measure recombination provides a powerful method to determine relative efficiency of genome editing for modeling polyglutamine diseases or understanding factors that modulate CRISPR/Cas9 HR.

Test-Retest Reliability of Diffusion Tensor Imaging in Huntington’s Disease

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Diffusion tensor imaging (DTI) has shown microstructural abnormalities in patients with Huntington’s Disease (HD) and work is underway to characterise how these abnormalities change with disease progression. Using methods that will be applied in longitudinal research, we sought to establish the reliability of DTI in early HD patients and controls. Test-retest reliability, quantified using the intraclass correlation coefficient (ICC), was assessed using region-of-interest (ROI)-based white matter atlas and voxelwise approaches on repeat scan data from 22 participants (10 early HD, 12 controls). T1 data was used to generate further ROIs for analysis in a reduced sample of 18 participants. The results suggest that fractional anisotropy (FA) and other diffusivity metrics are generally highly reliable, with ICCs indicating considerably lower within-subject compared to between-subject variability in both HD patients and controls. Where ICC was low, particularly for the diffusivity measures in the caudate and putamen, this was partly influenced by outliers. The analysis suggests that the specific DTI methods used here are appropriate for cross-sectional research in HD, and give confidence that they can also be applied longitudinally, although this requires further investigation. An important caveat for DTI studies is that test-retest reliability may not be evenly distributed throughout the brain whereby highly anisotropic white matter regions tended to show lower relative within-subject variability than other white or grey matter regions.

The PDE1/5 Inhibitor SCH-51866 Does Not Modify Disease Progression in the R6/2 Mouse Model of Huntington’s Disease

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Huntington’s disease is a neurodegenerative disorder caused by mutations in the CAG tract of huntingtin. Several studies in HD cellular and rodent systems have identified disturbances in cyclic nucleotide signaling, which might be relevant to pathogenesis and therapeutic intervention. To investigate whether selective phosphodiesterase (PDE) inhibitors can improve some aspects of disease pathogenesis in HD models, we have systematically evaluated the effects of a variety of cAMP and cGMP selective PDE inhibitors in various HD models. Here we present the lack of effect in a variety of endpoints of the PDE subtype selective inhibitor SCH-51866, a PDE1/5 inhibitor, in the R6/2 mouse model of HD, after chronic oral dosing.

Characterization of Striatal Neuronal Loss and Atrophy in the R6/2 Mouse Model of Huntington’s Disease

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Striatal neuronal degeneration and loss is an important feature of human Huntington’s disease (HD). R6/2 HD mice recapitulate many features of human HD including striatal atrophy. While striatal neuronal atrophy and loss is reported in R6/2 HD mice the degree of neuronal loss and the characteristics of cell body atrophy are unclear. We used stereological approaches to estimate whole striatal neuronal numbers and characterize changes in striatal neuronal size distribution. R6/2 HD mice had ~126000 fewer neurons per striatum (~12% decline) at 12 weeks of age than wild-type litter-mates; differences were not present at 5 weeks. Analysis of striatal neuronal numbers per cell body size category revealed declines in neuron numbers in the size ranges 550-1050 µm3 suggesting that larger striatal neurons are more susceptible to atrophy or loss in late stages of disease. R6/2 HD mice have a striatal neuronal loss phenotype. As striatal neuronal loss in human HD is dramatic, neuronal loss in R6/2 striatum provides an important late-stage outcome measure for study of disease modifying interventions

JAK/STAT Signalling in Huntington’s Disease Immune Cells

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Huntington’s disease (HD) is an inherited neurodegenerative disorder caused by a CAG repeat expansion in the huntingtin (HTT) gene. Both central and peripheral innate immune activation have been described as features of the disease. Isolated human HD monocytes have been shown to produce more cytokines upon LPS stimulation compared to control monocytes. Understanding alterations in the signalling cascades responsible and activated by this increase in pro-inflammatory cytokine production is crucial in understanding the molecular basis of this phenomenon. Here we investigated the signalling cascade most commonly activated by pro-inflammatory cytokines such as IL-6 – the JAK/STAT signalling cascade. Using flow cytometry, we show that one out of three key transcription factors activated by JAK/STAT signalling is altered in primary human HD innate immune cells, suggesting that this pathway may only play a minor, additive role in the immune cell dysfunction in HD.

Deficits in a Simple Visual Go/No-go Discrimination Task in Two Mouse Models of Huntington’s Disease

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Huntington’s disease (HD), a devastating neurodegenerative disorder caused by a CAG repeat expansion on the HTT gene located on chromosome 4, is associated with a characteristic pattern of progressive cognitive dysfunction known to involve early deficits in executive function. A modified Go/No-go successive discrimination task was designed to assess the type of online response control/executive function known to be disrupted in patients with HD. The present studies show that this simple discrimination assay revealed early and robust deficits in two mouse models of HD, the zQ175 KI mouse (deficits from 28 weeks of age) and the R6/2 mouse, carrying ~240 CAG repeats (deficits from 9 weeks of age). These deficits are not due to gross motor dysfunction in the test animals, but instead appear to measure some inability to inhibit responding in the HD mouse models, suggesting this assay may measure deficits in underlying attentional and/or behavioral inhibition processes. Accordingly, this assay may be well suited to evaluation of simple deficits in cognitive function in mouse HD models, providing a potential platform for preclinical screening.

Effects of a One Year Intensive Multidisciplinary Rehabilitation Program for Patients with Huntington’s Disease: a Prospective Intervention Study

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Objective: To assess the effects of an intensive, multidisciplinary rehabilitation program for patients with early to mid-stage Huntington’s disease.

Design: A prospective intervention study.

Setting: Two Norwegian inpatient rehabilitation centers.

Subjects: 37 patients, with early- to midstage Huntington’s disease

Interventions: A one year rehabilitation program, consisting of three admissions of three weeks each, and a five-day evaluation stay approximately 3 months after the last rehabilitation admission. Focus was on physical exercise, social activities, and group/teaching sessions. There was also emphasis to implement of coordinated health care and social services for the patients.

Main outcome measures: standard measures for motor function, including gait and balance, cognitive function, including MMSE and UHDRS cognitive assessment, anxiety and depression, activities of daily living (ADL), health related quality of life and Body Mass Index (BMI).

Results: Significant improvements were observed in gait function, balance, in physical quality of life, anxiety and depression, as well as in BMI. ADL-function remained stable with no significant decline. Only one cognitive measure (SDMT) showed significant decline, while no decline was observed for the remaining cognitive measures.

Conclusion: A multidisciplinary intensive rehabilitation program in patients with early and mid stage HD is associated with improved balance, gait function, physical quality of life and with reduced depressive and anxiety symptoms. Longer follow-up is needed to assess if these positive effects are sustained. There should be emphasis to establishment of long term and coordinated health care services for the HD patient