Generation of three isogenic gene-edited Huntington's disease human embryonic stem cell lines with DOX-inducible NGN2 expression cassette in the AAVS1 safe locus.

Neurogenin 2 (NGN2), a neuronal transcription factor, can expedite differentiation of stem cells into mature glutamatergic neurons. We have utilized an allelic series of previously published and characterized isogenic Huntington's disease (IsoHD) human embryonic stem cell lines (Ooi et al., 2019), carrying different CAG repeat lengths in the first exon of the huntingtin gene. These IsoHDs were modified using CRISPR/Cas9 to insert NGN2 under the TET-ON doxycycline inducible promoter. The resulting IsoHD-NGN2 cell lines retained pluripotency in the absence of doxycycline (DOX), and via addition of DOX to the culturing media differentiation to neurons was achieved within 14 days.

CHCHD2 up-regulation in Huntington disease mediates a compensatory protective response against oxidative stress.

Huntington disease (HD) is a neurodegenerative disease caused by the abnormal expansion of a polyglutamine tract resulting from a mutation in the HTT gene. Oxidative stress has been identified as a significant contributing factor to the development of HD and other neurodegenerative diseases, and targeting anti-oxidative stress has emerged as a potential therapeutic approach. CHCHD2 is a mitochondria-related protein involved in regulating cell migration, anti-oxidative stress, and anti-apoptosis. Although CHCHD2 is highly expressed in HD cells, its specific role in the pathogenesis of HD remains uncertain. We postulate that the up-regulation of CHCHD2 in HD models represents a compensatory protective response against mitochondrial dysfunction and oxidative stress associated with HD. To investigate this hypothesis, we employed HD mouse striatal cells and human induced pluripotent stem cells (hiPSCs) as models to examine the effects of CHCHD2 overexpression (CHCHD2-OE) or knockdown (CHCHD2-KD) on the HD phenotype. Our findings demonstrate that CHCHD2 is crucial for maintaining cell survival in both HD mouse striatal cells and hiPSCs-derived neurons. Our study demonstrates that CHCHD2 up-regulation in HD serves as a compensatory protective response against oxidative stress, suggesting a potential anti-oxidative strategy for the treatment of HD.

Environmental Deprivation Effects on Myelin Ultrastructure in Huntington Disease and Wildtype Mice.

Environmental deprivation can have deleterious effects on adaptive myelination and oligodendroglia function. Early stage Huntington disease (HD) is characterised by white-matter myelin abnormalities in both humans and animal models. However, whether deprived environments exacerbate myelin-related pathological features of HD is not clearly understood. Here, we investigated the impact of deprivation and social isolation on ultrastructural features of myelin in the corpus callosum of the YAC128 mouse model of HD and wildtype (WT) mice using transmission electron microscopy. HD pathology on its own leads to increased representation of altered myelin features, such as thinner sheaths and compromised morphology. Interestingly, deprivation mirrors these effects in WT mice but does not greatly exacerbate the already aberrant myelin in HD mice, indicating a disease-related floor effect in the latter animals. These novel findings indicate that environmental deprivation causes abnormalities in myelin ultrastructure in the otherwise healthy corpus callosum of wild-type mice but has distinct effects on HD mice, where compromised myelin integrity is manifest from early stages of the disease.

Dysregulated COMT Expression in Fragile X Syndrome.

Transcriptional and proteomics analyses in human fragile X syndrome (FXS) neurons identified markedly reduced expression of COMT, a key enzyme involved in the metabolism of catecholamines, including dopamine, epinephrine and norepinephrine. FXS is the most common genetic cause of intellectual disability and autism spectrum disorders. COMT encodes for catechol-o-methyltransferase and its association with neuropsychiatric disorders and cognitive function has been extensively studied. We observed a significantly reduced level of COMT in in FXS human neural progenitors and neurons, as well as hippocampal neurons from Fmr1 null mice. We show that deficits in COMT were associated with an altered response in an assay of dopaminergic activity in Fmr1 null mice. These findings demonstrate that loss of FMRP downregulates COMT expression and affects dopamine signaling in FXS, and supports the notion that targeting catecholamine metabolism may be useful in regulating certain neuropsychiatric aspects of FXS.

Age-Dependent Increase in Tau Phosphorylation at Serine 396 in Huntington's Disease Prefrontal Cortex.

BACKGROUND: To date, it is still controversial whether tau phosphorylation plays a role in Huntington's disease (HD), as previous studies demonstrated either no alterations or increases in phosphorylated tau (pTau) in HD postmortem brain and mouse models. OBJECTIVE: The goal of this study was to determine whether total tau and pTau levels are altered in HD. METHODS: Immunohistochemistry, cellular fractionations, and western blots were used to measure total tau and pTau levels in a large cohort of HD and control postmortem prefrontal cortex (PFC). Furthermore, western blots were performed to assess tau, and pTau levels in HD and control isogenic embryonic stem cell (ESC)-derived cortical neurons and neuronal stem cells (NSCs). Similarly, western blots were used to assess tau and pTau levels in HttQ111 and transgenic R6/2 mice. Lastly, total tau levels were assessed in HD and healthy control plasma using Quanterix Simoa assay. RESULTS: Our results revealed that, while there was no difference in total tau or pTau levels in HD PFC compared to controls, the levels of tau phosphorylated at S396 were increased in PFC samples from HD patients 60 years or older at time of death. Additionally, tau and pTau levels were not changed in HD ESC-derived cortical neurons and NSCs. Similarly, total tau or pTau levels were not altered in HttQ111 and transgenic R6/2 mice compared to wild-type littermates. Lastly, tau levels were not changed in plasma from a small cohort of HD patients compared to controls. CONCLUSIONS: Together these findings demonstrate that pTau-S396 levels increase significantly with age in HD PFC.

Generation of a human induced pluripotent stem cell line from a patient with hypomyelinating leukodystrophy 22 (HLD22).

Hypomyelinating Leukodystrophy 22 (HLD22) is caused by a stoploss mutation in CLDN11. To study the molecular mechanisms underlying HLD22, human induced pluripotent stem cells (hiPSCs) were generated from patient fibroblasts carrying the stop-loss mutation in CLDN11.

Oligodendrocyte pathology in Huntington's disease: from mechanisms to therapeutics.

Oligodendrocytes (OLGs), highly specialized glial cells that wrap axons with myelin sheaths, are critical for brain development and function. There is new recognition of the role of OLGs in the pathogenesis of neurodegenerative diseases (NDDs), including Huntington's disease (HD), a prototypic NDD caused by a polyglutamine tract expansion in huntingtin (HTT), which results in gain- and loss-of-function effects. Clinically, HD is characterized by a constellation of motor, cognitive, and psychiatric disturbances. White matter (WM) structures, representing myelin-rich regions of the brain, are profoundly affected in HD, and recent findings reveal oligodendroglia dysfunction as an early pathological event. Here, we focus on mechanisms that underlie oligodendroglial deficits and dysmyelination in the progression of the disease, highlighting the pathogenic contributions of mutant HTT (mHTT). We also discuss potential therapeutic implications involving these molecular pathways.

Widespread dysregulation of mRNA splicing implicates RNA processing in the development and progression of Huntington's disease.

BACKGROUND: In Huntington's disease (HD), a CAG repeat expansion mutation in the Huntingtin (HTT) gene drives a gain-of-function toxicity that disrupts mRNA processing. Although dysregulation of gene splicing has been shown in human HD post-mortem brain tissue, post-mortem analyses are likely confounded by cell type composition changes in late-stage HD, limiting the ability to identify dysregulation related to early pathogenesis. METHODS: To investigate gene splicing changes in early HD, we performed alternative splicing analyses coupled with a proteogenomics approach to identify early CAG length-associated splicing changes in an established isogenic HD cell model. FINDINGS: We report widespread neuronal differentiation stage- and CAG length-dependent splicing changes, and find an enrichment of RNA processing, neuronal function, and epigenetic modification-related genes with mutant HTT-associated splicing. When integrated with a proteomics dataset, we identified several of these differential splicing events at the protein level. By comparing with human post-mortem and mouse model data, we identified common patterns of altered splicing from embryonic stem cells through to post-mortem striatal tissue. INTERPRETATION: We show that widespread splicing dysregulation in HD occurs in an early cell model of neuronal development. Importantly, we observe HD-associated splicing changes in our HD cell model that were also identified in human HD striatum and mouse model HD striatum, suggesting that splicing-associated pathogenesis possibly occurs early in neuronal development and persists to later stages of disease. Together, our results highlight splicing dysregulation in HD which may lead to disrupted neuronal function and neuropathology. FUNDING: This research is supported by the Lee Kong Chian School of Medicine, Nanyang Technological University Singapore Nanyang Assistant Professorship Start-Up Grant, the Singapore Ministry of Education under its Singapore Ministry of Education Academic Research Fund Tier 1 (RG23/22), the BC Children's Hospital Research Institute Investigator Grant Award (IGAP), and a Scholar Award from the Michael Smith Health Research BC.

Age-dependent increase in tau phosphorylation at serine 396 in Huntington's disease pre-frontal cortex.

Background: To date, it is still controversial whether tau phosphorylation plays a role in Huntington's disease (HD), as previous studies demonstrated either no alterations or increases in phosphorylated tau (pTau) in HD post-mortem brain and mouse models. Objectives: The goal of this study was to determine whether total tau and pTau levels are altered in HD. Methods: Immunohistochemistry, cellular fractionations, and western blots were used to measure tau and pTau levels in a large cohort of HD and control post-mortem prefrontal cortex (PFC). Furthermore, western blots were performed to assess tau, and pTau levels in HD and control isogenic embryonic stem cell (ESC)-derived cortical neurons and neuronal stem cells (NSCs). Similarly, western blots were used to assess tau and pTau in Htt Q111 and transgenic R6/2 mice. Lastly, total tau levels were assessed in HD and healthy control plasma using Quanterix Simoa assay. Results: Our results revealed that, while there was no difference in tau or pTau levels in HD PFC compared to controls, tau phosphorylated at S396 levels were increased in PFC samples from HD patients 60 years or older at time of death. Additionally, tau and pTau levels were not changed in HD ESC-derived cortical neurons and NSCs. Similarly, tau or pTau levels were not altered in Htt Q111 and transgenic R6/2 mice compared to wild-type littermates. Lastly, tau levels were not changed in plasma from a small cohort of HD patients compared to controls. Conclusion: Together these findings demonstrate that pTau-S396 levels increase significantly with age in HD PFC.

Claudin-11 in health and disease: implications for myelin disorders, hearing, and fertility.

Claudin-11 plays a critical role in multiple physiological processes, including myelination, auditory function, and spermatogenesis. Recently, stop-loss mutations in CLDN11 have been identified as a novel cause of hypomyelinating leukodystrophy (HLD22). Understanding the multifaceted roles of claudin-11 and the potential pathogenic mechanisms in HLD22 is crucial for devising targeted therapeutic strategies. This review outlines the biological roles of claudin-11 and the implications of claudin-11 loss in the context of the Cldn11 null mouse model. Additionally, HLD22 and proposed pathogenic mechanisms, such as endoplasmic reticulum stress, will be discussed.

Evidences for Mutant Huntingtin Inducing Musculoskeletal and Brain Growth Impairments via Disturbing Testosterone Biosynthesis in Male Huntington Disease Animals.

Body weight (BW) loss and reduced body mass index (BMI) are the most common peripheral alterations in Huntington disease (HD) and have been found in HD mutation carriers and HD animal models before the manifestation of neurological symptoms. This suggests that, at least in the early disease stage, these changes could be due to abnormal tissue growth rather than tissue atrophy. Moreover, BW and BMI are reported to be more affected in males than females in HD animal models and patients. Here, we confirmed sex-dependent growth alterations in the BACHD rat model for HD and investigated the associated contributing factors. Our results showed growth abnormalities along with decreased plasma testosterone and insulin-like growth factor 1 (IGF-1) levels only in males. Moreover, we demonstrated correlations between growth parameters, IGF-1, and testosterone. Our analyses further revealed an aberrant transcription of testosterone biosynthesis-related genes in the testes of BACHD rats with undisturbed luteinizing hormone (LH)/cAMP/PKA signaling, which plays a key role in regulating the transcription process of some of these genes. In line with the findings in BACHD rats, analyses in the R6/2 mouse model of HD showed similar results. Our findings support the view that mutant huntingtin may induce abnormal growth in males via the dysregulation of gene transcription in the testis, which in turn can affect testosterone biosynthesis.

Maturation Delay of Human GABAergic Neurogenesis in Fragile X Syndrome Pluripotent Stem Cells.

Fragile X Syndrome (FXS), the leading monogenic cause of intellectual disability and autism spectrum disorder, is caused by expansion of a CGG trinucleotide repeat in the 5'-UTR of the Fragile X Mental Retardation-1 (FMR1) gene. Epigenetic silencing of FMR1 results in loss of the Fragile X Mental Retardation Protein (FMRP). Although most studies to date have focused on excitatory neurons, recent evidence suggests that GABAergic inhibitory networks are also affected. To investigate human GABAergic neurogenesis, we established a method to reproducibly derive inhibitory neurons from multiple FXS and control human pluripotent stem cell (hPSC) lines. Electrophysiological analyses suggested that the developing FXS neurons had a delay in the GABA functional switch, a transition in fetal development that converts the GABAA channel's function from depolarization to hyperpolarization, with profound effects on the developing brain. To investigate the cause of this delay, we analyzed 14 400 single-cell transcriptomes from FXS and control cells at 2 stages of GABAergic neurogenesis. While control and FXS cells were similar at the earlier time point, the later-stage FXS cells retained expression of neuroblast proliferation-associated genes and had lower levels of genes associated with action potential regulation, synapses, and mitochondria compared with controls. Our analysis suggests that loss of FMRP prolongs the proliferative stage of progenitors, which may result in more neurons remaining immature during the later stages of neurogenesis. This could have profound implications for homeostatic excitatory-inhibitory circuit development in FXS, and suggests a novel direction for understanding disease mechanisms that may help to guide therapeutic interventions.

Ermin deficiency leads to compromised myelin, inflammatory milieu, and susceptibility to demyelinating insult.

Ermin is an actin-binding protein found almost exclusively in the central nervous system (CNS) as a component of myelin sheaths. Although Ermin has been predicted to play a role in the formation and stability of myelin sheaths, this has not been directly examined in vivo. Here, we show that Ermin is essential for myelin sheath integrity and normal saltatory conduction. Loss of Ermin in mice caused de-compacted and fragmented myelin sheaths and led to slower conduction along with progressive neurological deficits. RNA sequencing of the corpus callosum, the largest white matter structure in the CNS, pointed to inflammatory activation in aged Ermin-deficient mice, which was corroborated by increased levels of microgliosis and astrogliosis. The inflammatory milieu and myelin abnormalities were further associated with increased susceptibility to immune-mediated demyelination insult in Ermin knockout mice. Supporting a possible role of Ermin deficiency in inflammatory white matter disorders, a rare inactivating mutation in the ERMN gene was identified in multiple sclerosis patients. Our findings demonstrate a critical role for Ermin in maintaining myelin integrity. Given its near-exclusive expression in myelinating oligodendrocytes, Ermin deficiency represents a compelling "inside-out" model of inflammatory dysmyelination and may offer a new paradigm for the development of myelin stability-targeted therapies.

Early white matter pathology in the fornix of the limbic system in Huntington disease.

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.

Rescue of aberrant huntingtin palmitoylation ameliorates mutant huntingtin-induced toxicity.

Huntington disease (HD) is a neurodegenerative disorder caused by a CAG expansion in the HTT gene that codes for an elongated polyglutamine tract in the huntingtin (HTT) protein. HTT is subject to multiple post-translational modifications (PTMs) that regulate its cellular function. Mutating specific PTM sites within mutant HTT (mHTT) in HD mouse models can modulate disease phenotypes, highlighting the key role of HTT PTMs in the pathogenesis of HD. These findings have led to increased interest in developing small molecules to modulate HTT PTMs in order to decrease mHTT toxicity. However, the therapeutic efficacy of pharmacological modulation of HTT PTMs in preclinical HD models remains largely unknown. HTT is palmitoylated at cysteine 214 by the huntingtin-interacting protein 14 (HIP14 or ZDHHC17) and 14-like (HIP14L or ZDHHC13) acyltransferases. Here, we assessed if HTT palmitoylation should be regarded as a therapeutic target to treat HD by (1) investigating palmitoylation dysregulation in rodent and human HD model systems, (2) measuring the impact of mHTT-lowering therapy on brain palmitoylation, and (3) evaluating if HTT palmitoylation can be pharmacologically modulated. We show that palmitoylation of mHTT and some HIP14/HIP14L-substrates is decreased early in multiple HD mouse models, and that mHTT palmitoylation decreases further with aging. Lowering mHTT in the brain of YAC128 mice is not sufficient to rescue aberrant palmitoylation. However, we demonstrate that mHTT palmitoylation can be normalized in COS-7 cells, in YAC128 cortico-striatal primary neurons and HD patient-derived lymphoblasts using an acyl-protein thioesterase (APT) inhibitor. Moreover, we show that modulating palmitoylation reduces mHTT aggregation and mHTT-induced cytotoxicity in COS-7 cells and YAC128 neurons.

Abnormal Spinal Cord Myelination due to Oligodendrocyte Dysfunction in a Model of Huntington's Disease.

BACKGROUND: The relative contribution of grey matter (GM) and white matter (WM) degeneration to the progressive brain atrophy in Huntington's disease (HD) has been well studied. The pathology of the spinal cord in HD is comparatively less well documented. OBJECTIVE: We aim to characterize spinal cord WM abnormalities in a mouse model of HD and evaluate whether selective removal of mutant huntingtin (mHTT) from oligodendroglia rescues these deficits. METHODS: Histological assessments were used to determine the area of GM and WM in the spinal cord of 12-month-old BACHD mice, while electron microscopy was used to analyze myelin fibers in the cervical area of the spinal cord. To investigate the impact of inactivation of mHTT in oligodendroglia on these measures, we used the previously described BACHDxNG2Cre mouse line where mHTT is specifically reduced in oligodendrocyte progenitor cells. RESULTS: We show that spinal GM and WM areas are significantly atrophied in HD mice compared to wild-type controls. We further demonstrate that specific reduction of mHTT in oligodendroglial cells rescues the atrophy of spinal cord WM, but not GM, observed in HD mice. Inactivation of mHTT in oligodendroglia had no effect on the density of oligodendroglial cells but enhanced the expression of myelin-related proteins in the spinal cord. CONCLUSION: Our findings demonstrate that the myelination abnormalities observed in brain WM structures in HD extend to the spinal cord and suggest that specific expression of mHTT in oligodendrocytes contributes to such abnormalities.

Urokinase plasminogen activator mediates changes in human astrocytes modeling fragile X syndrome.

The function of astrocytes intertwines with the extracellular matrix, whose neuron and glial cell-derived components shape neuronal plasticity. Astrocyte abnormalities have been reported in the brain of the mouse model for fragile X syndrome (FXS), the most common cause of inherited intellectual disability, and a monogenic cause of autism spectrum disorder. We compared human FXS and control astrocytes generated from human induced pluripotent stem cells and we found increased expression of urokinase plasminogen activator (uPA), which modulates degradation of extracellular matrix. Several pathways associated with uPA and its receptor function were activated in FXS astrocytes. Levels of uPA were also increased in conditioned medium collected from FXS hiPSC-derived astrocyte cultures and correlated inversely with intracellular Ca2+ responses to activation of L-type voltage-gated calcium channels in human astrocytes. Increased uPA augmented neuronal phosphorylation of TrkB within the docking site for the phospholipase-Cγ1 (PLCγ1), indicating effects of uPA on neuronal plasticity. Gene expression changes during neuronal differentiation preceding astrogenesis likely contributed to properties of astrocytes with FXS-specific alterations that showed specificity by not affecting differentiation of adenosine triphosphate (ATP)-responsive astrocyte population. To conclude, our studies identified uPA as an important regulator of astrocyte function and demonstrated that increased uPA in human FXS astrocytes modulated astrocytic responses and neuronal plasticity.

Pluripotent stem cell-derived models of neurological diseases reveal early transcriptional heterogeneity.

BACKGROUND: Many neurodegenerative diseases develop only later in life, when cells in the nervous system lose their structure or function. In many forms of neurodegenerative diseases, this late-onset phenomenon remains largely unexplained. RESULTS: Analyzing single-cell RNA sequencing from Alzheimer's disease (AD) and Huntington's disease (HD) patients, we find increased transcriptional heterogeneity in disease-state neurons. We hypothesize that transcriptional heterogeneity precedes neurodegenerative disease pathologies. To test this idea experimentally, we use juvenile forms (72Q; 180Q) of HD iPSCs, differentiate them into committed neuronal progenitors, and obtain single-cell expression profiles. We show a global increase in gene expression variability in HD. Autophagy genes become more stable, while energy and actin-related genes become more variable in the mutant cells. Knocking down several differentially variable genes results in increased aggregate formation, a pathology associated with HD. We further validate the increased transcriptional heterogeneity in CHD8+/- cells, a model for autism spectrum disorder. CONCLUSIONS: Overall, our results suggest that although neurodegenerative diseases develop over time, transcriptional regulation imbalance is present already at very early developmental stages. Therefore, an intervention aimed at this early phenotype may be of high diagnostic value.

Generation of the Human Pluripotent Stem-Cell-Derived Astrocyte Model with Forebrain Identity.

Astrocytes form functionally and morphologically distinct populations of cells with brain-region-specific properties. Human pluripotent stem cells (hPSCs) offer possibilities to generate astroglia for studies investigating mechanisms governing the emergence of astrocytic diversity. We established a method to generate human astrocytes from hPSCs with forebrain patterning and final specification with ciliary neurotrophic factor (CNTF). Transcriptome profiling and gene enrichment analysis monitored the sequential expression of genes determining astrocyte differentiation and confirmed activation of forebrain differentiation pathways at Day 30 (D30) and D60 of differentiation in vitro. More than 90% of astrocytes aged D95 in vitro co-expressed the astrocytic markers glial fibrillary acidic protein (GFAP) and S100ß. Intracellular calcium responses to ATP indicated differentiation of the functional astrocyte population with constitutive monocyte chemoattractant protein-1 (MCP-1/CCL2) and tissue inhibitor of metalloproteinases-2 (TIMP-2) expression. The method was reproducible across several hPSC lines, and the data demonstrated the usefulness of forebrain astrocyte modeling in research investigating forebrain pathology.

pS421 huntingtin modulates mitochondrial phenotypes and confers neuroprotection in an HD hiPSC model.

Huntington disease (HD) is a hereditary neurodegenerative disorder caused by mutant huntingtin (mHTT). Phosphorylation at serine-421 (pS421) of mHTT has been shown to be neuroprotective in cellular and rodent models. However, the genetic context of these models differs from that of HD patients. Here we employed human pluripotent stem cells (hiPSCs), which express endogenous full-length mHTT. Using genome editing, we generated isogenic hiPSC lines in which the S421 site in mHTT has been mutated into a phospho-mimetic aspartic acid (S421D) or phospho-resistant alanine (S421A). We observed that S421D, rather than S421A, confers neuroprotection in hiPSC-derived neural cells. Although we observed no effect of S421D on mHTT clearance or axonal transport, two aspects previously reported to be impacted by phosphorylation of mHTT at S421, our analysis revealed modulation of several aspects of mitochondrial form and function. These include mitochondrial surface area, volume, and counts, as well as improved mitochondrial membrane potential and oxidative phosphorylation. Our study validates the protective role of pS421 on mHTT and highlights a facet of the relationship between mHTT and mitochondrial changes in the context of human physiology with potential relevance to the pathogenesis of HD.