| 1. |
- Maxwell, Christopher A., et al.
(författare)
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Interplay between BRCA1 and RHAMM Regulates Epithelial Apicobasal Polarization and May Influence Risk of Breast Cancer
- 2011
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Ingår i: PLoS Biology. - : Public Library of Science (PLoS). - 1545-7885 .- 1544-9173. ; 9:11
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Tidskriftsartikel (refereegranskat)abstract
- Differentiated mammary epithelium shows apicobasal polarity, and loss of tissue organization is an early hallmark of breast carcinogenesis. In BRCA1 mutation carriers, accumulation of stem and progenitor cells in normal breast tissue and increased risk of developing tumors of basal-like type suggest that BRCA1 regulates stem/progenitor cell proliferation and differentiation. However, the function of BRCA1 in this process and its link to carcinogenesis remain unknown. Here we depict a molecular mechanism involving BRCA1 and RHAMM that regulates apicobasal polarity and, when perturbed, may increase risk of breast cancer. Starting from complementary genetic analyses across families and populations, we identified common genetic variation at the low-penetrance susceptibility HMMR locus (encoding for RHAMM) that modifies breast cancer risk among BRCA1, but probably not BRCA2, mutation carriers: n = 7,584, weighted hazard ratio ((w)HR) = 1.09 (95% CI 1.02-1.16), p(trend) = 0.017; and n = 3,965, (w)HR = 1.04 (95% CI 0.94-1.16), p(trend) = 0.43; respectively. Subsequently, studies of MCF10A apicobasal polarization revealed a central role for BRCA1 and RHAMM, together with AURKA and TPX2, in essential reorganization of microtubules. Mechanistically, reorganization is facilitated by BRCA1 and impaired by AURKA, which is regulated by negative feedback involving RHAMM and TPX2. Taken together, our data provide fundamental insight into apicobasal polarization through BRCA1 function, which may explain the expanded cell subsets and characteristic tumor type accompanying BRCA1 mutation, while also linking this process to sporadic breast cancer through perturbation of HMMR/RHAMM.
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| 2. |
- Ast, Anne, et al.
(författare)
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mHTT Seeding Activity : A Marker of Disease Progression and Neurotoxicity in Models of Huntington's Disease
- 2018
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Ingår i: Molecular Cell. - : Elsevier BV. - 1097-2765. ; 71:5, s. 6-688
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Tidskriftsartikel (refereegranskat)abstract
- Self-propagating, amyloidogenic mutant huntingtin (mHTT) aggregates may drive progression of Huntington's disease (HD). Here, we report the development of a FRET-based mHTT aggregate seeding (FRASE) assay that enables the quantification of mHTT seeding activity (HSA) in complex biosamples from HD patients and disease models. Application of the FRASE assay revealed HSA in brain homogenates of presymptomatic HD transgenic and knockin mice and its progressive increase with phenotypic changes, suggesting that HSA quantitatively tracks disease progression. Biochemical investigations of mouse brain homogenates demonstrated that small, rather than large, mHTT structures are responsible for the HSA measured in FRASE assays. Finally, we assessed the neurotoxicity of mHTT seeds in an inducible Drosophila model transgenic for HTTex1. We found a strong correlation between the HSA measured in adult neurons and the increased mortality of transgenic HD flies, indicating that FRASE assays detect disease-relevant, neurotoxic, mHTT structures with severe phenotypic consequences in vivo. Ast et al. present the development of a FRET-based aggregate seeding (FRASE) assay that facilitates the detection and quantification of mHTT seeding activity (HSA) in complex biosamples. They show that HSA is detectable in brains of presymptomatic Huntington's disease (HD) mice and correlates with disease progression and neurotoxicity in HD transgenic flies.
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| 3. |
- Baldo, Barbara, et al.
(författare)
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Effects of Deletion of Mutant Huntingtin in Steroidogenic Factor 1 Neurons on the Psychiatric and Metabolic Phenotype in the BACHD Mouse Model of Huntington Disease.
- 2014
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 9:10
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Tidskriftsartikel (refereegranskat)abstract
- Psychiatric and metabolic features appear several years before motor disturbances in the neurodegenerative Huntington's disease (HD), caused by an expanded CAG repeat in the huntingtin (HTT) gene. Although the mechanisms leading to these aspects are unknown, dysfunction in the hypothalamus, a brain region controlling emotion and metabolism, has been suggested. A direct link between the expression of the disease causing protein, huntingtin (HTT), in the hypothalamus and the development of metabolic and psychiatric-like features have been shown in the BACHD mouse model of HD. However, precisely which circuitry in the hypothalamus is critical for these features is not known. We hypothesized that expression of mutant HTT in the ventromedial hypothalamus, an area involved in the regulation of metabolism and emotion would be important for the development of these non-motor aspects. Therefore, we inactivated mutant HTT in a specific neuronal population of the ventromedial hypothalamus expressing the transcription factor steroidogenic factor 1 (SF1) in the BACHD mouse using cross-breeding based on a Cre-loxP system. Effects on anxiety-like behavior were assessed using the elevated plus maze and novelty-induced suppressed feeding test. Depressive-like behavior was assessed using the Porsolt forced swim test. Effects on the metabolic phenotype were analyzed using measurements of body weight and body fat, as well as serum insulin and leptin levels. Interestingly, the inactivation of mutant HTT in SF1-expressing neurons exerted a partial positive effect on the depressive-like behavior in female BACHD mice at 4 months of age. In this cohort of mice, no anxiety-like behavior was detected. The deletion of mutant HTT in SF1 neurons did not have any effect on the development of metabolic features in BACHD mice. Taken together, our results indicate that mutant HTT regulates metabolic networks by affecting hypothalamic circuitries that do not involve the SF1 neurons of the ventromedial hypothalamus.
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| 4. |
- Baldo, Barbara, et al.
(författare)
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Maintenance of Basal Levels of Autophagy in Huntington's Disease Mouse Models Displaying Metabolic Dysfunction.
- 2013
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:12
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Tidskriftsartikel (refereegranskat)abstract
- Huntington's disease (HD) is a fatal neurodegenerative disorder caused by an expanded polyglutamine repeat in the huntingtin protein. Neuropathology in the basal ganglia and in the cerebral cortex has been linked to the motor and cognitive symptoms whereas recent work has suggested that the hypothalamus might be involved in the metabolic dysfunction. Several mouse models of HD that display metabolic dysfunction have hypothalamic pathology, and expression of mutant huntingtin in the hypothalamus has been causally linked to the development of metabolic dysfunction in mice. Although the pathogenic mechanisms by which mutant huntingtin exerts its toxic functions in the HD brain are not fully known, several studies have implicated a role for the lysososomal degradation pathway of autophagy. Interestingly, changes in autophagy in the hypothalamus have been associated with the development of metabolic dysfunction in wild-type mice. We hypothesized that expression of mutant huntingtin might lead to changes in the autophagy pathway in the hypothalamus in mice with metabolic dysfunction. We therefore investigated whether there were changes in basal levels of autophagy in a mouse model expressing a fragment of 853 amino acids of mutant huntingtin selectively in the hypothalamus using a recombinant adeno-associate viral vector approach as well as in the transgenic BACHD mice. We performed qRT-PCR and Western blot to investigate the mRNA and protein expression levels of selected autophagy markers. Our results show that basal levels of autophagy are maintained in the hypothalamus despite the presence of metabolic dysfunction in both mouse models. Furthermore, although there were no major changes in autophagy in the striatum and cortex of BACHD mice, we detected modest, but significant differences in levels of some markers in mice at 12 months of age. Taken together, our results indicate that overexpression of mutant huntingtin in mice do not significantly perturb basal levels of autophagy.
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| 5. |
- Baldo, Barbara, et al.
(författare)
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Quantification of Total and Mutant Huntingtin Protein Levels in Biospecimens Using a Novel alphaLISA Assay
- 2018
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Ingår i: eNeuro. - 2373-2822. ; 5:4
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Tidskriftsartikel (refereegranskat)abstract
- The neurodegenerative Huntington's disease (HD) is caused by a polyglutamine (polyQ) amplification in the huntingtin protein (HTT). Currently there is no effective therapy available for HD; however, several efforts are directed to develop and optimize HTT-lowering methods to improve HD phenotypes. To validate these approaches, there is an immediate need for reliable, sensitive, and easily accessible methods to quantify HTT expression. Using the AlphaLISA platform, we developed two novel sensitive and robust assays for quantification of HTT in biological samples using commercially available antibodies. The first, a polyQ-independent assay, measures the total pool of HTT, while the second, a polyQ-dependent assay, preferentially detects the mutant form of HTT. Using purified HTT protein standards and brain homogenates from an HD mouse model, we determine a lower limit of quantification of 1 and 3 pm and optimal reproducibility with CV values lower than 7% for intra- and 20% for interassay. In addition, we used the assays to quantify HTT in neural stem cells generated from patient-derived induced pluripotent stem cells in vitro and in human brain tissue lysates. Finally, we could detect changes in HTT levels in a mouse model where mutant HTT was conditionally deleted in neural tissue, verifying the potential to monitor the outcome of HTT-lowering strategies. This analytical platform is ideal for high-throughput screens and thus has an added value for the HD community as a tool to optimize novel therapeutic approaches aimed at modulating HTT protein levels.
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| 6. |
- Cheong, Rachel Y., et al.
(författare)
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Effects of mutant huntingtin inactivation on Huntington disease-related behaviours in the BACHD mouse model
- 2021
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Ingår i: Neuropathology and Applied Neurobiology. - : Wiley. - 0305-1846 .- 1365-2990. ; 47:4, s. 564-578
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Tidskriftsartikel (refereegranskat)abstract
- Aims: Huntington disease (HD) is a fatal neurodegenerative disorder with no disease-modifying treatments approved so far. Ongoing clinical trials are attempting to reduce huntingtin (HTT) expression in the central nervous system (CNS) using different strategies. Yet, the distribution and timing of HTT-lowering therapies required for a beneficial clinical effect is less clear. Here, we investigated whether HD-related behaviours could be prevented by inactivating mutant HTT at different disease stages and to varying degrees in an experimental model. Methods: We generated mutant BACHD mice with either a widespread or circuit-specific inactivation of mutant HTT by using Cre recombinase (Cre) under the nestin promoter or the adenosine A2A receptor promoter respectively. We also simulated a clinical gene therapy scenario with allele-specific HTT targeting by injections of recombinant adeno-associated viral (rAAV) vectors expressing Cre into the striatum of adult BACHD mice. All mice were assessed using behavioural tests to investigate motor, metabolic and psychiatric outcome measures at 4–6 months of age. Results: While motor deficits, body weight changes, anxiety and depressive-like behaviours are present in BACHD mice, early widespread CNS inactivation during development significantly improves rotarod performance, body weight changes and depressive-like behaviour. However, conditional circuit-wide mutant HTT deletion from the indirect striatal pathway during development and focal striatal-specific deletion in adulthood failed to rescue any of the HD-related behaviours. Conclusions: Our results indicate that widespread targeting and the timing of interventions aimed at reducing mutant HTT are important factors to consider when developing disease-modifying therapies for HD.
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| 7. |
- Gabery, Sanaz, et al.
(författare)
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Early white matter pathology in the fornix of the limbic system in Huntington disease
- 2021
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Ingår i: Acta Neuropathologica. - : Springer Science and Business Media LLC. - 0001-6322 .- 1432-0533. ; 142:5, s. 791-806
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Tidskriftsartikel (refereegranskat)abstract
- Huntington disease (HD) is a fatal neurodegenerative disorder caused by an expanded CAG repeat in the huntingtin (HTT) gene. The typical motor symptoms have been associated with basal ganglia pathology. However, psychiatric and cognitive symptoms often precede the motor component and may be due to changes in the limbic system. Recent work has indicated pathology in the hypothalamus in HD but other parts of the limbic system have not been extensively studied. Emerging evidence suggests that changes in HD also include white matter pathology. Here we investigated if the main white matter tract of the limbic system, the fornix, is affected in HD. We demonstrate that the fornix is 34% smaller already in prodromal HD and 41% smaller in manifest HD compared to controls using volumetric analyses of MRI of the IMAGE-HD study. In post-mortem fornix tissue from HD cases, we confirm the smaller fornix volume in HD which is accompanied by signs of myelin breakdown and reduced levels of the transcription factor myelin regulating factor but detect no loss of oligodendrocytes. Further analyses using RNA-sequencing demonstrate downregulation of oligodendrocyte identity markers in the fornix of HD cases. Analysis of differentially expressed genes based on transcription-factor/target-gene interactions also revealed enrichment for binding sites of SUZ12 and EZH2, components of the Polycomb Repressive Complex 2, as well as RE1 Regulation Transcription Factor. Taken together, our data show that there is early white matter pathology of the fornix in the limbic system in HD likely due to a combination of reduction in oligodendrocyte genes and myelin break down.
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| 8. |
- Iacomussi, Sofia, et al.
(författare)
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Governance of Access in Biobanking: The Case of Telethon Network of Genetic Biobanks
- 2021
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Ingår i: Biopreservation and Biobanking. - : Mary Ann Liebert. - 1947-5535 .- 1947-5543. ; 19:6, s. 483-492
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Tidskriftsartikel (refereegranskat)abstract
- The discussion concerning the measure of the quality of a biobank should focus not only on the number of stored samples and their quality but also on the assessment of their access arrangements and governance. This article aims at contributing to the ongoing debate on samples and data access governance in biobanking by presenting the case of the Telethon Network of Genetic Biobanks (TNGB). We attempt to contribute to the need for clear and available access criteria and harmonization in access arrangements to maximize the influence of biobanks in the progress of biomedical research. We reviewed all the sample requests submitted to the TNGB from 2008 to 2020, focusing on those rejected by the Access Committee and the reasons behind the rejections. The analysis of the reasons behind the rejected requests allowed us to analyze how those relate to the issues of scientific misconduct, prioritization, and noncompliance with the biobank's mission. We discuss those issues in light of the actions and motivations used by TNGB in the access decision-making process. Based on this analysis, we suggest that a cross-implementation of a checklist for access assessment would improve the whole access process, ensuring a more transparent and smoother governance. Finally, we conclude that the TNGB's Charter and approach toward access governance could contribute as an important reference point to deal with the issues that have emerged in the international discussion on the topic.
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| 9. |
- Martrat, Griselda, et al.
(författare)
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Exploring the link between MORF4L1 and risk of breast cancer
- 2011
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Ingår i: Breast Cancer Research. - : Springer Science and Business Media LLC. - 1465-5411 .- 1465-542X. ; 13:2
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Tidskriftsartikel (refereegranskat)abstract
- Introduction: Proteins encoded by Fanconi anemia (FA) and/or breast cancer (BrCa) susceptibility genes cooperate in a common DNA damage repair signaling pathway. To gain deeper insight into this pathway and its influence on cancer risk, we searched for novel components through protein physical interaction screens. Methods: Protein physical interactions were screened using the yeast two-hybrid system. Co-affinity purifications and endogenous co-immunoprecipitation assays were performed to corroborate interactions. Biochemical and functional assays in human, mouse and Caenorhabditis elegans models were carried out to characterize pathway components. Thirteen FANCD2-monoubiquitinylation-positive FA cell lines excluded for genetic defects in the downstream pathway components and 300 familial BrCa patients negative for BRCA1/2 mutations were analyzed for genetic mutations. Common genetic variants were genotyped in 9,573 BRCA1/2 mutation carriers for associations with BrCa risk. Results: A previously identified co-purifying protein with PALB2 was identified, MRG15 (MORF4L1 gene). Results in human, mouse and C. elegans models delineate molecular and functional relationships with BRCA2, PALB2, RAD51 and RPA1 that suggest a role for MRG15 in the repair of DNA double-strand breaks. Mrg15-deficient murine embryonic fibroblasts showed moderate sensitivity to g-irradiation relative to controls and reduced formation of Rad51 nuclear foci. Examination of mutants of MRG15 and BRCA2 C. elegans orthologs revealed phenocopy by accumulation of RPA-1 (human RPA1) nuclear foci and aberrant chromosomal compactions in meiotic cells. However, no alterations or mutations were identified for MRG15/MORF4L1 in unclassified FA patients and BrCa familial cases. Finally, no significant associations between common MORF4L1 variants and BrCa risk for BRCA1 or BRCA2 mutation carriers were identified: rs7164529, P-trend = 0.45 and 0.05, P-2df = 0.51 and 0.14, respectively; and rs10519219, P-trend = 0.92 and 0.72, P-2df = 0.76 and 0.07, respectively. Conclusions: While the present study expands on the role of MRG15 in the control of genomic stability, weak associations cannot be ruled out for potential low-penetrance variants at MORF4L1 and BrCa risk among BRCA2 mutation carriers.
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| 10. |
- Soylu-Kucharz, Rana, et al.
(författare)
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Metabolic and behavioral effects of mutant huntingtin deletion in Sim1 neurons in the BACHD mouse model of Huntington's disease
- 2016
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 6
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Tidskriftsartikel (refereegranskat)abstract
- Hypothalamic pathology, metabolic dysfunction and psychiatric symptoms are part of Huntington disease (HD), which is caused by an expanded CAG repeat in the huntingtin (HTT) gene. Inactivation of mutant HTT selectively in the hypothalamus prevents the development of metabolic dysfunction and depressive-like behavior in the BACHD mouse model. The hypothalamic paraventricular nucleus (PVN) is implicated in metabolic and emotional control, therefore we here tested whether inactivation of mutant HTT in the PVN affects metabolic and psychiatric manifestations of HD in BACHD mice. BACHD mice were crossed with mice expressing Cre-recombinase under the Sim1 promoter (Sim1-Cre) to inactivate mutant HTT in Sim1 expressing cells, i.e. the PVN of the hypothalamus. We found that inactivation of mutant HTT in Sim1 cells had a sex-specific effect on both the metabolic and the psychiatric phenotype, as these phenotypes were no longer different in male BACHD/Sim1-Cre mice compared to wild-type littermates. We also found a reduced number of GnRH neurons specifically in the anterior hypothalamus and an increased testes weight in male BACHD mice compared to wild-type littermates. Taken together, expression of mutant HTT in Sim1 cells may play a role for the development of metabolic dysfunction and depressive-like behavior in male BACHD mice.
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