Huntington Disease (HD) is a dominantly inherited neurological disease attributed to a CAG expansion within the HD gene. The HD mutation gives rise to a polyglutamine expansion in exon 1 of the protein huntingtin (Htt). Since the discovery of the HD mutation in 1993, various HD gene mouse models have been developed to contain either fragments or full-length copies of the mutant HD gene. The existence of these HD mouse models enables focused therapeutic testing to develop potential treatments for HD. RNA interference (RNAi) therapy is a targeted gene silencing approach whereby synthetic RNA constructs are shuttled into the cell by viral vectors and used by the cell’s endogenous RNAi machinery to silence a gene of interest. RNAi therapy holds promise for mutant huntingtin (muHtt) allele-specific silencing as a treatment for HD. The purpose of this thesis was to develop the tools for pre-clinical testing of RNAi-mediated gene silencing of human muHtt in the YAC128 mouse model of HD. First, AAV serotypes were compared for delivery to striatal neurons, the neurons most affected in HD. From this work AAV serotype 1 was selected as the most effective serotype for construct delivery. Second, synthetic RNAi constructs including short-hairpin RNA (shRNA) and microRNA-based constructs (miR-shRNAs) were compared for silencing of human muHtt expression in vivo. Here, miR-shRNAs were found to have increased gene silencing and improved tolerance in avoiding immune activation compared to shRNAs. Alternatively, the shRNAs induced dramatic immune activation and morbidity in some cases. Ultimately these findings will contribute to a pre-clinical trial in YAC128 mice investigating Htt RNAi-mediated gene silencing in the treatment of HD, which is also discussed in this thesis. This future work provides proof-of-principle for muHtt allele-specific silencing as a treatment of HD.

The purpose of this dissertation is to examine the use, role and function of medical pedigrees as part of extended networks of genetic practices. Integral to my argument is a description of geneticisation (i.e., the redefinition of family problems as genetic in origin), grounded in a set of detailed case studies of the development and use of visual tools in genetic practices. / In recent years, medical sociologists have tended to link geneticisation to medicalisation (i.e., the social control by doctors over patients accompanied by the translation of social problems into medical issues). I argue that the twin notions of geneticisation and medicalisation are problematic, insofar as they embody a simplistic and negative understanding of medical activities and they prevent a sociological inquiry into the technical content of genetic practices. / Medical pedigrees are visual tools used to translate family problems into visual inscriptions, in order to show the genetic nature of a given disease. The use of medical pedigrees in genetic counselling and research rests on a chain of genetic practices including the inscription of family trees, the standardisation of medical pedigrees, the combination of specialised forms of medical pedigrees with other diagnostic inscriptions, and the circulation of published pedigrees. The analysis is based on a genealogical approach, as built on a combination of historical and ethnographic methods. The genealogical approach was applied to the analysis of a long network of genetic practices centred on Huntington's disease. The analysis spans over 120 years and compares two different international settings (North America and Japan). / The thesis examines how lay support group members and family members collect family narratives, family inscriptions and family trees, which were first translated by genetic counsellors into various forms of medical pedigrees, and then circulated as educational material among lay and medical practitioners. On the basis of these case studies, the conclusion is reached that the notion of geneticisation should be understood as a specific process resulting from an emerging cooperative practice between medical practitioners and lay support group members, rather than as a process of medicalisation.

Huntington Disease (HD) is a dominantly inherited neurological disease attributed to a CAG expansion within the HD gene. The HD mutation gives rise to a polyglutamine expansion in exon 1 of the protein huntingtin (Htt). Since the discovery of the HD mutation in 1993, various HD gene mouse models have been developed to contain either fragments or full-length copies of the mutant HD gene. The existence of these HD mouse models enables focused therapeutic testing to develop potential treatments for HD. RNA interference (RNAi) therapy is a targeted gene silencing approach whereby synthetic RNA constructs are shuttled into the cell by viral vectors and used by the cell’s endogenous RNAi machinery to silence a gene of interest. RNAi therapy holds promise for mutant huntingtin (muHtt) allele-specific silencing as a treatment for HD. The purpose of this thesis was to develop the tools for pre-clinical testing of RNAi-mediated gene silencing of human muHtt in the YAC128 mouse model of HD. First, AAV serotypes were compared for delivery to striatal neurons, the neurons most affected in HD. From this work AAV serotype 1 was selected as the most effective serotype for construct delivery. Second, synthetic RNAi constructs including short-hairpin RNA (shRNA) and microRNA-based constructs (miR-shRNAs) were compared for silencing of human muHtt expression in vivo. Here, miR-shRNAs were found to have increased gene silencing and improved tolerance in avoiding immune activation compared to shRNAs. Alternatively, the shRNAs induced dramatic immune activation and morbidity in some cases. Ultimately these findings will contribute to a pre-clinical trial in YAC128 mice investigating Htt RNAi-mediated gene silencing in the treatment of HD, which is also discussed in this thesis. This future work provides proof-of-principle for muHtt allele-specific silencing as a treatment of HD.

Huntington Disease (HD) is a dominantly inherited neurological disease attributed to a CAG expansion within the HD gene. The HD mutation gives rise to a polyglutamine expansion in exon 1 of the protein huntingtin (Htt). Since the discovery of the HD mutation in 1993, various HD gene mouse models have been developed to contain either fragments or full-length copies of the mutant HD gene. The existence of these HD mouse models enables focused therapeutic testing to develop potential treatments for HD. RNA interference (RNAi) therapy is a targeted gene silencing approach whereby synthetic RNA constructs are shuttled into the cell by viral vectors and used by the cell’s endogenous RNAi machinery to silence a gene of interest. RNAi therapy holds promise for mutant huntingtin (muHtt) allele-specific silencing as a treatment for HD. The purpose of this thesis was to develop the tools for pre-clinical testing of RNAi-mediated gene silencing of human muHtt in the YAC128 mouse model of HD. First, AAV serotypes were compared for delivery to striatal neurons, the neurons most affected in HD. From this work AAV serotype 1 was selected as the most effective serotype for construct delivery. Second, synthetic RNAi constructs including short-hairpin RNA (shRNA) and microRNA-based constructs (miR-shRNAs) were compared for silencing of human muHtt expression in vivo. Here, miR-shRNAs were found to have increased gene silencing and improved tolerance in avoiding immune activation compared to shRNAs. Alternatively, the shRNAs induced dramatic immune activation and morbidity in some cases. Ultimately these findings will contribute to a pre-clinical trial in YAC128 mice investigating Htt RNAi-mediated gene silencing in the treatment of HD, which is also discussed in this thesis. This future work provides proof-of-principle for muHtt allele-specific silencing as a treatment of HD. / Medicine, Faculty of / Medical Genetics, Department of / Graduate

Amino acids and other ninhydrin-positive compounds were measured in post-mortem thalamus from 25 Huntington's choreics, 10 schizophrenics, 5 schizophrenic-like psychotics, and 23 controls dying without neurological disease. Gamma-aminobutyric acid (GABA) was significantly reduced in choreic thalami, in accord with deficiencies found in other brain regions choreics (Perry et al., 1973a,b). GABA was also significantly reduced in schizophrenic thalami, suggesting a biochemical link between these two diseases, and supporting the hypothesis of a defect in the GABA system in schizophrenia (Roberts, 1972). Homocarnosine, a GABA-containing dipeptide, was also low in choreic and 9 out of 10 schizophrenic thalami. One schizophrenic had extremely high homocarnosine. Glycerophosphoethanolamine was significantly elevated in Huntington's choreics, but not in schizophrenics. A number of other variables were considered for their potential influence on amino acid concentrations in thalamus. The majority of amino acids were found to rise in a significantly linear fashion in the interval 3 to 49 hours post-mortem, although other models might have described the change better. GABA, ornithine, histidine and tyrosine were found to decrease significantly with increasing age between 21 and 80 years, in controls. The effects of pre-mortem hypoxia, regional variation within the thalamus, and neuroleptic drug treatment could not be rigorously tested with these data. Neuroleptics were unlikely to have been the cause of group differences in GABA concentration, since they failed to deplete GABA in brain of chronically treated rats. On the other hand, bronchopneumonia and other causes of pre-mortem hypoxia could not be ruled out as potential contributers to reduced GABA in thalamus. / Medicine, Faculty of / Medical Genetics, Department of / Graduate

Huntington disease (HD) is an inherited neurodegenerative disease caused by a trinucleotide repeat expansion in the huntingtin (HTT) gene. Metabolic dysfunction is a feature of HD that is recapitulated in HD mouse models. Our lab has shown that circadian feeding rhythms are disrupted in humanized HD mice and restored by suppression of brain HTT. Furthermore, when circadian feeding rhythm is artificially restored, in addition to normalization of metabolic function, liver and striatal HTT is temporarily reduced, demonstrating that HTT is involved in gut-brain feedback. The gut microbiome, which can regulate gut-brain feedback, has been implicated in the pathogenesis of other central nervous system disorders and we hypothesize it also plays a role in HD. The objective of this study is to investigate alterations in relative abundance of HD gut microbiota using existing plasma metabolomics data to identify candidate bacteria. If distinct microbiota profiles are demonstrated, this would provide the basis for future unbiased studies to investigate the complete HD microbiome.

No description available.

O objetivo deste trabalho foi caracterizar as alterações de linguagem na doença de Huntington e correlacioná-las com os transtornos motores, cognitivos, psiquiátricos e, também, com o tempo de doença. Foram estudados 26 indivíduos, divididos em grupo leve (11 doentes) e moderado (15 doentes), comparados com dois grupos controle. Foram encontradas alterações em provas de compreensão e expressão da linguagem oral e gráfica para os dois grupos de doentes. Evidências sugerem que não há prejuízo nas representações semânticas, e que as dificuldades de linguagem estão relacionadas com o declínio cognitivo global e, especialmente, com o prejuízo das funções executivas. As alterações de linguagem se correlacionaram com o desempenho em tarefas cognitivas, mas não com as alterações motoras ou psiquiátricas da doença. Também não foi encontrada correlação entre o desempenho de linguagem e o tempo de doença / The objective of this study was to characterize the language alterations in Huntington's disease and how they relate to severity of motor, cognitive, psychiatric disturbances and also with the disease duration. Twenty-six (26) individuals were divided into groups characterized as lightly (11) and moderately ill (15) and compared with two control groups. Alterations in exams of language comprehension and expression were noticed for both groups of sick individuals. The result indicates no evident loss in semantic representation. The language difficulties are related to a global cognitive decline and, principally, to loss of executive functions. The language alterations were significantly correlated to performance in cognitive tasks, but not to the motor or psychiatric alterations of the disease. There was also no correlation observed between the language performance and duration of illness

Knowledge of genetic predisposition to future illness and disability creates uncertainties that shape and influence life decisions about reproduction, career, health behavior, and the need for care. Current research has not yet identified the meaning of the experience of feeling uncertain among intimate partners of persons who have received genetic information pertaining to future health status. The purpose of this phenomenological study was to understand the meaning of uncertainty as a lived experienced among intimate partners of persons who have tested positive for a mutation in the gene causing Huntington disease (HD) but have not yet been clinically diagnosed with HD. The specific aims were to create a rich, vivid description of uncertainty as experienced by this population and to present these findings within an existential phenomenological perspective. Using van Manen's hermeneutic-phenomenological methodology, experiential descriptions from 10 intimate partners of persons in the prodromal phase of HD were obtained. Thematic aspects of the lived experience of uncertainty were uncovered and isolated; essential themes were determined; and linguistic transformations were composed. The analysis revealed four essential themes, indicating that the meaning of the lived experience of uncertainty was 1) an intangible reality characterized by 2) anticipating with ebbing and flowing disquietude while feeling 3) a weighty pull to dwell upon, towards inner turmoil and 4) a subdued presence with freeing possibilities. The implications of these findings are that nurses need to ensure adequate opportunity is created in which the meaning of the lived experience of uncertainty can be ascertained and explored among persons who are on the cusp of the inevitable but not yet graspable. Continued research is needed to further address the implications of being situated in this potentially fracturing phase of the disease trajectory and to determine appropriate interventions.

Polyglutamine-expanded fragments, derived from the human huntingtin protein, are aggregation-prone and toxic in yeast cells, bearing endogenous QN-rich proteins in the aggregated (prion) form. Attachment of the proline-rich region targets polyglutamine aggregates to the large perinuclear deposit (aggresome). Aggresome targeting ameliorates polyglutamine cytotoxicity in the presence of the prion form of Rnq1 protein, however, aggresome-forming construct remains toxic in the presence of the prion form of translation termination (release) factor Sup35 (eRF3). Disomy by chromosome II partly ameliorates polyglutamine toxicity in the strains containing Sup35 prion. The chromosome II gene, coding for another release factor, and interaction partner of Sup35, named Sup45 (eRF1), is responsible for amelioration of toxicity. Plasmid-mediated overproduction of Sup45, or expression of the Sup35 derivative that lacks the QN-rich domain and is unable to be incorporated into prion aggregates, also ameliorate polyglutamine toxicity. Protein analysis indicates that polyglutamines alter aggregation patterns of the Sup35 prion and promote aggregation of Sup45, while excess Sup45 counteracts these effects. In the absence of Sup35 prion, disomy by chromosome II is still able to decrease polyglutamine toxicity. However, SUP45 is no longer the gene responsible for such an effect. Taken together with the finding that the presence of both the Rnq1 prion and the Sup35 prion has an additive effect on polyQ toxicity, one gene or few genes on chromosome II are able to ameliorate polyQ toxicity through a SUP45-independent pathway. The identification of such a gene is currently ongoing. Monosomy by chromosome VIII in diploid heterozygous by AQT (Anti-polyQ Toxicity mutants that are disomic by chromosome II) counteracted the effect of AQT. Similarly, deletion of the arg4 gene in chromosome VIII in AQT haploid was able to eliminate the AQT effect. Moreover, analysis of genes involved in the arginine and polyamine synthesis indicated that loss of genes in later stages of arginine biosynthesis causes increase of polyglutamine toxicity. Deletion of genes arg1, arg4, arg8 (arginine pathway) and spe1 (polyamine pathway) all suppressed the Sup35 prion phenotype expression in the nonsense suppression system. Further analysis regarding the mechanisms behind those effects is needed. Our data uncover the mechanisms by which genetic and epigenetic factors may influence polyglutamine toxicity, and demonstrate that one and the same type of polyglutamine deposits could be cytoprotective or cytotoxic, depending on the prion composition of a eukaryotic cell.