BACKGROUND: Huntington's disease (HD) is a devastating neurodegenerative disorder caused by CAG triplet expansions in the huntingtin gene. Oxidative stress is linked to HD pathology, although it is not clear whether this is an effect or a mediator of disease. The transgenic (TgHD) minipig expresses the N-terminal part of human-mutated huntingtin and represents a unique model to investigate therapeutic strategies towards HD. A more detailed characterization of this model is needed to fully utilize its potential. METHODS: In this study, we focused on the molecular and cellular features of fibroblasts isolated from TgHD minipigs and the wild-type (WT) siblings at different ages, pre-symptomatic at the age of 24-36 months and with the onset of behavioural symptoms at the age of 48 months. We measured oxidative stress, the expression of oxidative stress-related genes, proliferation capacity along with the expression of cyclin B1 and D1 proteins, cellular permeability, and the integrity of the nuclear DNA (nDNA) and mitochondrial DNA in these cells. RESULTS: TgHD fibroblasts isolated from 48-month-old animals showed increased oxidative stress, which correlated with the overexpression of SOD2 encoding mitochondrial superoxide dismutase 2, and the NEIL3 gene encoding DNA glycosylase involved in replication-associated repair of oxidized DNA. TgHD cells displayed an abnormal proliferation capacity and permeability. We further demonstrated increased nDNA damage in pre-symptomatic TgHD fibroblasts (isolated from animals aged 24-36 months). CONCLUSIONS: Our results unravel phenotypic alterations in primary fibroblasts isolated from the TgHD minipig model at the age of 48 months. Importantly, nDNA damage appears to precede these phenotypic alterations. Our results highlight the impact of fibroblasts from TgHD minipigs in studying the molecular mechanisms of HD pathophysiology that gradually occur with age.

• Klíčová slova
• Permeability, DNA damage, Huntington’s disease, Large-animal model, Minipig model, Mutated huntingtin, Oxidative stress, Primary fibroblasts, Proliferation,
• MeSH
• buněčné dělení MeSH
• fenotyp MeSH
• fibroblasty metabolismus   MeSH
• geneticky modifikovaná zvířata MeSH
• lidé MeSH
• miniaturní prasata MeSH
• mitochondriální DNA genetika   MeSH
• N-glykosylhydrolasy biosyntéza   genetika   MeSH
• oxidační stres MeSH
• peroxidace lipidů MeSH
• poškození DNA MeSH
• prasata MeSH
• primární buněčná kultura MeSH
• protein huntingtin genetika   metabolismus   MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• regulace genové exprese MeSH
• stárnutí metabolismus   MeSH
• superoxiddismutasa biosyntéza   genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• HTT protein, human MeSH Prohlížeč
• mitochondriální DNA MeSH
• N-glykosylhydrolasy MeSH
• protein huntingtin MeSH
• reaktivní formy kyslíku MeSH
• superoxiddismutasa MeSH
• superoxide dismutase 2 MeSH Prohlížeč

BACKGROUND: Huntington's disease is induced by CAG expansion in a single gene coding the huntingtin protein. The mutated huntingtin (mtHtt) primarily causes degeneration of neurons in the brain, but it also affects peripheral tissues, including testes. OBJECTIVE: We studied sperm and testes of transgenic boars expressing the N-terminal region of human mtHtt. METHODS: In this study, measures of reproductive parameters and electron microscopy (EM) images of spermatozoa and testes of transgenic (TgHD) and wild-type (WT) boars of F1 (24-48 months old) and F2 (12-36 months old) generations were compared. In addition, immunofluorescence, immunohistochemistry, Western blot, hormonal analysis and whole-genome sequencing were done in order to elucidate the effects of mtHtt. RESULTS: Evidence for fertility failure of both TgHD generations was observed at the age of 13 months. Reproductive parameters declined and progressively worsened with age. EM revealed numerous pathological features in sperm tails and in testicular epithelium from 24- and 36-month-old TgHD boars. Moreover, immunohistochemistry confirmed significantly lower proliferation activity of spermatogonia in transgenic testes. mtHtt was highly expressed in spermatozoa and testes of TgHD boars and localized in all cells of seminiferous tubules. Levels of fertility-related hormones did not differ in TgHD and WT siblings. Genome analysis confirmed that insertion of the lentiviral construct did not interrupt any coding sequence in the pig genome. CONCLUSIONS: The sperm and testicular degeneration of TgHD boars is caused by gain-of-function of the highly expressed mtHtt.

• MeSH
• genetické vektory MeSH
• geneticky modifikovaná zvířata MeSH
• Huntingtonova nemoc metabolismus   patologie   MeSH
• Lentivirus genetika   MeSH
• lidé MeSH
• miniaturní prasata MeSH
• modely nemocí na zvířatech MeSH
• mutace * MeSH
• počet spermií MeSH
• prasata MeSH
• proliferace buněk fyziologie   MeSH
• protein huntingtin genetika   metabolismus   MeSH
• spermie metabolismus   patologie   MeSH
• stárnutí metabolismus   patologie   MeSH
• testis metabolismus   patologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• HTT protein, human MeSH Prohlížeč
• protein huntingtin MeSH

Huntington's disease (HD) is a fatal neurodegenerative disorder caused by a CAG trinucleotide repeat expansion in the huntingtin gene. Previously, we showed strong huntingtin reduction and prevention of neuronal dysfunction in HD rodents using an engineered microRNA targeting human huntingtin, delivered via adeno-associated virus (AAV) serotype 5 vector with a transgene encoding an engineered miRNA against HTT mRNA (AAV5-miHTT). One of the challenges of rodents as a model of neurodegenerative diseases is their relatively small brain, making successful translation to the HD patient difficult. This is particularly relevant for gene therapy approaches, where distribution achieved upon local administration into the parenchyma is likely dependent on brain size and structure. Here, we aimed to demonstrate the translation of huntingtin-lowering gene therapy to a large-animal brain. We investigated the feasibility, efficacy, and tolerability of one-time intracranial administration of AAV5-miHTT in the transgenic HD (tgHD) minipig model. We detected widespread dose-dependent distribution of AAV5-miHTT throughout the tgHD minipig brain that correlated with the engineered microRNA expression. Both human mutant huntingtin mRNA and protein were significantly reduced in all brain regions transduced by AAV5-miHTT. The combination of widespread vector distribution and extensive huntingtin lowering observed with AAV5-miHTT supports the translation of a huntingtin-lowering gene therapy for HD from preclinical studies into the clinic.

• Klíčová slova
• AAV, Huntington disease, gene silencing, microRNA, transgenic minipig,
• MeSH
• Dependovirus genetika   MeSH
• expanze trinukleotidových repetic genetika   MeSH
• genetická terapie metody   MeSH
• genetické vektory genetika   MeSH
• geneticky modifikovaná zvířata MeSH
• Huntingtonova nemoc genetika   metabolismus   terapie   MeSH
• lidé MeSH
• mikro RNA genetika   metabolismus   MeSH
• miniaturní prasata MeSH
• modely nemocí na zvířatech MeSH
• prasata MeSH
• protein huntingtin genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• mikro RNA MeSH
• protein huntingtin MeSH

The pathogenic effect of mutant HTT (mHTT) which causes Huntington disease (HD) are not restricted to nervous system. Such phenotypes include aberrant immune responses observed in the HD models. However, it is still unclear how this immune dysregulation influences the innate immune response against pathogenic infection. In the present study, we used transgenic Drosophila melanogaster expressing mutant HTT protein (mHTT) with hemocyte-specific drivers and examined the immune responses and hemocyte function. We found that mHTT expression in the hemocytes did not affect fly viability, but the numbers of circulating hemocytes were significantly decreased. Consequently, we observed that the expression of mHTT in the hemocytes compromised the immune responses including clot formation and encapsulation which lead to the increased susceptibility to entomopathogenic nematode and parasitoid wasp infections. In addition, mHTT expression in Drosophila macrophage-like S2 cells in vitro reduced ATP levels, phagocytic activity and the induction of antimicrobial peptides. Further effects observed in mHTT-expressing cells included the altered production of cytokines and activation of JAK/STAT signaling. The present study shows that the expression of mHTT in Drosophila hemocytes causes deficient cellular and humoral immune responses against invading pathogens. Our findings provide the insight into the pathogenic effects of mHTT in the immune cells.

• Klíčová slova
• Drosophila melanogaster, Huntington's disease, antimicrobial peptide (AMPs), cytokines, immunity, infection, phagocytosis,
• MeSH
• buněčné linie MeSH
• Drosophila melanogaster MeSH
• exprese genu * MeSH
• geneticky modifikovaná zvířata MeSH
• hemocyty imunologie   MeSH
• humorální imunita * MeSH
• lidé MeSH
• protein huntingtin genetika   imunologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• HTT protein, human MeSH Prohlížeč
• protein huntingtin MeSH

Huntington's disease (HD) is a fatal neurodegenerative disease caused by a genetic expansion of the CAG repeat region in the huntingtin (HTT) gene. Studies in HD mouse models have shown that artificial miRNAs can reduce mutant HTT, but evidence for their effectiveness and safety in larger animals is lacking. HD transgenic sheep express the full-length human HTT with 73 CAG repeats. AAV9 was used to deliver unilaterally to HD sheep striatum an artificial miRNA targeting exon 48 of the human HTT mRNA under control of two alternative promoters: U6 or CβA. The treatment reduced human mutant (m) HTT mRNA and protein 50-80% in the striatum at 1 and 6 months post injection. Silencing was detectable in both the caudate and putamen. Levels of endogenous sheep HTT protein were not affected. There was no significant loss of neurons labeled by DARPP32 or NeuN at 6 months after treatment, and Iba1-positive microglia were detected at control levels. It is concluded that safe and effective silencing of human mHTT protein can be achieved and sustained in a large-animal brain by direct delivery of an AAV carrying an artificial miRNA.

• Klíčová slova
• AAV, Huntington's disease, RNAi, large animal models,
• MeSH
• Dependovirus genetika   MeSH
• elektrolyty metabolismus   MeSH
• genetické vektory metabolismus   MeSH
• geneticky modifikovaná zvířata MeSH
• genom virový MeSH
• Huntingtonova nemoc genetika   patologie   MeSH
• imunoanalýza MeSH
• injekce MeSH
• játra patofyziologie   MeSH
• ledviny patofyziologie   MeSH
• lidé MeSH
• messenger RNA genetika   metabolismus   MeSH
• mikro RNA genetika   metabolismus   MeSH
• mikroglie metabolismus   MeSH
• modely nemocí na zvířatech MeSH
• mutantní proteiny metabolismus   MeSH
• neostriatum metabolismus   MeSH
• neurony metabolismus   MeSH
• ovce MeSH
• protein huntingtin metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• elektrolyty MeSH
• HTT protein, human MeSH Prohlížeč
• messenger RNA MeSH
• mikro RNA MeSH
• mutantní proteiny MeSH
• protein huntingtin MeSH

(1) Background: Huntington's disease (HD) is rare incurable hereditary neurodegenerative disorder caused by CAG repeat expansion in the gene coding for the protein huntingtin (HTT). Mutated huntingtin (mHTT) undergoes fragmentation and accumulation, affecting cellular functions and leading to neuronal cell death. Porcine models of HD are used in preclinical testing of currently emerging disease modifying therapies. Such therapies are aimed at reducing mHTT expression, postpone the disease onset, slow down the progression, and point out the need of biomarkers to monitor disease development and therapy efficacy. Recently, extracellular vesicles (EVs), particularly exosomes, gained attention as possible carriers of disease biomarkers. We aimed to characterize HTT and mHTT forms/fragments in blood plasma derived EVs in transgenic (TgHD) and knock-in (KI-HD) porcine models, as well as in HD patients' plasma. (2) Methods: Small EVs were isolated by ultracentrifugation and HTT forms were visualized by western blotting. (3) Results: The full length 360 kDa HTT co-isolated with EVs from both the pig model and HD patient plasma. In addition, a ~70 kDa mutant HTT fragment was specific for TgHD pigs. Elevated total huntingtin levels in EVs from plasma of HD groups compared to controls were observed in both pig models and HD patients, however only in TgHD were they significant (p = 0.02). (4) Conclusions: Our study represents a valuable initial step towards the characterization of EV content in the search for HD biomarkers.

• Klíčová slova
• Huntington´s disease, KI-HD, TgHD, biomarker, exosome, extracellular vesicle, fragment, huntingtin, neurodegenerative disease, pig model,
• MeSH
• biologické markery MeSH
• extracelulární vezikuly * metabolismus   MeSH
• Huntingtonova nemoc * metabolismus   MeSH
• krevní plazma metabolismus   MeSH
• lidé MeSH
• prasata MeSH
• proteiny nervové tkáně genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• biologické markery MeSH
• proteiny nervové tkáně MeSH

In Germany at least 8000 and probably up to ca. 14,000 people currently suffer from clinically manifest Huntington's disease (HD). In addition, an estimated 24,000 Germans carry the HD mutation in the huntingtin (HTT) gene and will develop HD during their lifetime. Although HD is a rare neurodegenerative disease, it is currently in the focus of general medical interest: clinical trials have begun that provide a rational basis for hope to slow down the so far relentless progression of the disease, ultimately resulting in patients becoming entirely dependent on nursing care. If treatment is started early enough it may be possible to mitigate the clinical manifestation of HD. These innovative therapeutic approaches aim at inhibiting the de novo production of mutant HTT gene products. A first clinical drug trial to demonstrate the efficacy (phase III) of intrathecal antisense oligonucleotides (ASO, active substance RG6042) was started in 2019. Additional clinical studies on alternative treatment approaches with allele-selective ASOs as well as gene therapeutic approaches using RNA molecules and zinc finger repressor complexes are imminent. This article gives an overview of the current gene-selective therapeutic approaches in HD under discussion.

• Klíčová slova
• Adeno-associated viruses, Antisense oligonucleotide, Gene therapy, Huntingtin, Zinc finger repressor complexes,
• MeSH
• antisense oligonukleotidy terapeutické užití   MeSH
• genetická terapie * trendy   MeSH
• Huntingtonova nemoc * terapie   MeSH
• klinické zkoušky jako téma MeSH
• lidé MeSH
• protein huntingtin genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Geografické názvy
• Německo MeSH
• Názvy látek
• antisense oligonukleotidy MeSH
• protein huntingtin MeSH

Huntington's disease (HD) is a progressive neurodegenerative disorder characterized by cognitive, motor, and psychiatric symptoms. Despite significant advances in understanding the underlying molecular mechanisms of HD, there is currently no cure or disease-modifying treatment available. Emerging pharmacological approaches offer promising strategies to alleviate symptoms and slow down disease progression. This comprehensive review aims to provide a critical appraisal of the latest developments in pharmacological interventions for HD. The review begins by discussing the pathogenesis of HD, focusing on the role of mutant huntingtin protein, mitochondrial dysfunction, excitotoxicity, and neuro-inflammation. It then explores emerging therapeutic targets, including the modulation of protein homeostasis, mitochondrial function, neuro-inflammation, and neurotransmitter systems. Pharmacological agents targeting these pathways are discussed, including small molecules, gene-based therapies, and neuroprotective agents. In recent years, several clinical trials have been conducted to evaluate the safety and efficiency of novel compounds for HD. This review presents an update on the outcomes of these trials, highlighting promising results and challenges encountered. Additionally, it discusses the potential of repurposing existing drugs approved for other indications as a cost-effective approach for HD treatment. The review concludes by summarizing the current state of pharmacological approaches for HD and outlining future directions in drug development. The integration of multiple therapeutic strategies, personalized medicine approaches, and combination therapies are highlighted as potential avenues to maximize treatment effectiveness.

• Klíčová slova
• Huntington's disease, Mitochondrial dysfunction, Mutant huntingtin protein, Neurodegenerative disorders, Pharmacological approaches,
• MeSH
• genetická terapie metody   MeSH
• Huntingtonova nemoc * farmakoterapie   MeSH
• lidé MeSH
• mitochondrie účinky léků   metabolismus   MeSH
• neuroprotektivní látky * terapeutické užití   farmakologie   MeSH
• protein huntingtin genetika   antagonisté a inhibitory   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• neuroprotektivní látky * MeSH
• protein huntingtin MeSH

Huntington's disease (HD) is a rare hereditary autosomal dominant neurodegenerative disorder, which is caused by expression of mutant huntingtin protein (mHTT) with an abnormal number of glutamine repeats in its N terminus, and characterized by intracellular mHTT aggregates (inclusions) in the brain. Exosomes are small extracellular vesicles that are secreted generally by all cell types and can be isolated from almost all body fluids such as blood, urine, saliva, and cerebrospinal fluid. Exosomes may participate in the spreading of toxic misfolded proteins across the central nervous system in neurodegenerative diseases. In HD, such propagation of mHTT was observed both in vitro and in vivo. On the other hand, exosomes might carry molecules with neuroprotective effects. In addition, due to their capability to cross blood-brain barrier, exosomes hold great potential as sources of biomarkers available from periphery or carriers of therapeutics into the central nervous system. In this review, we discuss the emerging roles of exosomes in HD pathogenesis, diagnosis, and therapy.

• Klíčová slova
• Huntington’s disease, biomarker, exosome, extracellular vesicle, huntingtin, neurodegeneration, polyQ, therapy,
• MeSH
• biologické modely MeSH
• exozómy metabolismus   MeSH
• Huntingtonova nemoc metabolismus   MeSH
• lidé MeSH
• protein huntingtin metabolismus   MeSH
• sbalování proteinů MeSH
• systémy cílené aplikace léků MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• protein huntingtin MeSH

BACKGROUND: Although the highest expression of mutant huntingtin (mtHtt) was observed in the brain, its negative effects were also apparent in other tissues. Specifically, mtHtt impairs metabolic homeostasis and causes transcriptional dysregulation in adipose tissue. Adipogenic differentiation can be induced by the activation of two transcription factors: CCAAT/enhancer-binding protein alpha (CEBPα) and peroxisome proliferator-activated receptor gamma (PPARγ). These same transcription factors were found to be compromised in some tissues of Huntington's disease (HD) mouse models and in lymphocytes of HD patients. OBJECTIVE: This study investigated the adipogenic potential of mesenchymal stem cells (MSCs) derived from transgenic Huntington's disease (TgHD) minipigs expressing human mtHtt (1-548aa) containing 124 glutamines. Two differentiation conditions were used, employing PPARγ agonist rosiglitazone or indomethacin. METHODS: Bone marrow MSCs were isolated from TgHD and WT minipig siblings and compared by their cluster of differentiation using flow cytometry. Their adipogenic potential in vitro was analyzed using quantitative immunofluorescence and western blot analysis of transcription factors and adipogenic markers. RESULTS: Flow cytometry analysis did not reveal any significant difference between WT and TgHD MSCs. Nevertheless, following differentiation into adipocytes, the expression of CEBPα nuclear, PPARγ and adipogenic marker FABP4/AP2 were significantly lower in TgHD cells compared to WT cells. In addition, we proved both rosiglitazone and indomethacin to be efficient for adipogenic differentiation of porcine MSCs, with rosiglitazone showing a better adipogenic profile. CONCLUSIONS: We demonstrated a negative influence of mtHtt on adipogenic differentiation of porcine MSCs in vitro associated with compromised expression of adipogenic transcription factors.

• Klíčová slova
• Huntington’s disease, adipogenesis, mesenchymal stem cells, minipigs,
• MeSH
• adipogeneze * MeSH
• buňky kostní dřeně cytologie   MeSH
• geneticky modifikovaná zvířata genetika   MeSH
• Huntingtonova nemoc genetika   patologie   MeSH
• kultivované buňky MeSH
• lidé MeSH
• mezenchymální kmenové buňky cytologie   MeSH
• prasata MeSH
• transkripční faktory genetika   MeSH
• tukové buňky cytologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• transkripční faktory MeSH