Neuron-specific ablation of the Krabbe disease gene galactosylceramidase in mice results in neurodegeneration.
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ABSTRACT:
SUBMITTER: Kreher C
PROVIDER: S-EPMC9255775 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Krabbe disease is an inherited neurodegenerative disease caused by mutations in the galactosylceramidase gene. In the infantile form, patients die before 3 years of age. Systemic adeno-associated virus serotype 9 (AAV9) gene therapy was recently shown to reverse the disease course in human patients in another lethal infantile neurodegenerative disease. To explore AAV9 therapy for Krabbe disease, we engineered a codon-optimized AAV9 galactosylceramidase vector. We further incorporated features to allow AAV9-derived galactosylceramidase to more efficiently cross the blood-brain barrier and be secreted from transduced cells. We tested the optimized vector by a single systemic injection in the twitcher mouse, an authentic Krabbe disease model. Untreated twitcher mice showed characteristic neuropathology and motion defects. They died prematurely with a median life span of 41 days. Intravenous injection in 2-day-old twitcher mice reduced central and peripheral neuropathology and significantly improved the gait pattern and body weight. Noticeably, the median life span was extended to 150 days. Intraperitoneal injection in 6- to 12-day-old twitcher mice also significantly improved the motor function, body weight, and median life span (to 104 days). Our results far exceed the ≤70 days median life span seen in all reported stand-alone systemic AAV therapies. Our study highlights the importance of vector engineering for Krabbe disease gene therapy. The engineered vector warrants further development.
| S-EPMC6761594 | biostudies-literature
Project description:Krabbe disease (KD) is caused by a deficiency of galactosylceramidase (GALC), which induces demyelination and neurodegeneration due to accumulation of cytotoxic psychosine. Hematopoietic stem cell transplantation (HSCT) improves clinical outcomes in KD patients only if delivered pre-symptomatically. Here, we hypothesize that the restricted temporal efficacy of HSCT reflects a requirement for GALC in early brain development. Using a novel Galc floxed allele, we induce ubiquitous GALC ablation (Galc-iKO) at various postnatal timepoints and identify a critical period of vulnerability to GALC ablation between P4-6 in mice. Early Galc-iKO induction causes a worse KD phenotype, higher psychosine levels in the rodent brainstem and spinal cord, and a significantly shorter life-span of the mice. Intriguingly, GALC expression peaks during this critical developmental period in mice. Further analysis of this mouse model reveals a cell autonomous role for GALC in the development and maturation of immature T-box-brain-1 positive brainstem neurons. These data identify a perinatal developmental period, in which neuronal GALC expression influences brainstem development that is critical for KD pathogenesis.
| S-EPMC7584660 | biostudies-literature
Project description:Human Ag R (HuR) is an RNA binding protein in the ELAVL protein family. To study the neuron-specific function of HuR, we generated inducible, neuron-specific HuR-deficient mice of both sexes. After tamoxifen-induced deletion of HuR, these mice developed a phenotype consisting of poor balance, decreased movement, and decreased strength. They performed significantly worse on the rotarod test compared with littermate control mice, indicating coordination deficiency. Using the grip-strength test, it was also determined that the forelimbs of neuron-specific HuR-deficient mice were much weaker than littermate control mice. Immunostaining of the brain and cervical spinal cord showed that HuR-deficient neurons had increased levels of cleaved caspase-3, a hallmark of cell apoptosis. Caspase-3 cleavage was especially strong in pyramidal neurons and ? motor neurons of HuR-deficient mice. Genome-wide microarray and real-time PCR analysis further indicated that HuR deficiency in neurons resulted in altered expression of genes in the brain involved in cell growth, including trichoplein keratin filament-binding protein, Cdkn2c, G-protein signaling modulator 2, immediate early response 2, superoxide dismutase 1, and Bcl2. The additional enriched Gene Ontology terms in the brain tissues of neuron-specific HuR-deficient mice were largely related to inflammation, including IFN-induced genes and complement components. Importantly, some of these HuR-regulated genes were also significantly altered in the brain and spinal cord of patients with amyotrophic lateral sclerosis. Additionally, neuronal HuR deficiency resulted in the redistribution of TDP43 to cytosolic granules, which has been linked to motor neuron disease. Taken together, we propose that this neuron-specific HuR-deficient mouse strain can potentially be used as a motor neuron disease model.
| S-EPMC6008238 | biostudies-literature
Project description:Globoid cell leukodystrophy (GLD), or Krabbe disease, is a neurodegenerative sphingolipidosis caused by genetic deficiency of lysosomal β-galactosylceramidase (GALC), characterized by neuroinflammation and demyelination of the central (CNS) and peripheral nervous system. The acute phase protein long pentraxin-3 (PTX3) is a soluble pattern recognition receptor and a regulator of innate immunity. Growing evidence points to the involvement of PTX3 in neurodegeneration. However, the expression and role of PTX3 in the neurodegenerative/neuroinflammatory processes that characterize GLD remain unexplored. Here, immunohistochemical analysis of brain samples from Krabbe patients showed that macrophages and globoid cells are intensely immunoreactive for PTX3. Accordingly, Ptx3 expression increases throughout the course of the disease in the cerebrum, cerebellum, and spinal cord of GALC-deficient twitcher (Galctwi/twi) mice, an authentic animal model of GLD. This was paralleled by the upregulation of proinflammatory genes and M1-polarized macrophage/microglia markers and of the levels of PTX3 protein in CNS and plasma of twitcher animals. Crossing of Galctwi/twi mice with transgenic PTX3 overexpressing animals (hPTX3 mice) demonstrated that constitutive PTX3 overexpression reduced the severity of clinical signs and the upregulation of proinflammatory genes in the spinal cord of P35 hPTX3/Galctwi/twi mice when compared to Galctwi/twi littermates, leading to a limited increase of their life span. However, this occurred in the absence of a significant impact on the histopathological findings and on the accumulation of the neurotoxic metabolite psychosine when evaluated at this late time point of the disease. In conclusion, our results provide the first evidence that PTX3 is produced in the CNS of GALC-deficient Krabbe patients and twitcher mice. PTX3 may exert a protective role by reducing the neuroinflammatory response that occurs in the spinal cord of GALC-deficient animals.
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Project description:Multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE), are chronic inflammatory demyelinating and neurodegenerative diseases of the CNS. Although neurodegeneration is the major contributor to chronic disability in MS, mechanisms governing the viability of axons and neurons in MS and EAE remain elusive. Data indicate that activation of pancreatic endoplasmic reticulum kinase (PERK) influences, positively or negatively, neuron and axon viability in various neurodegenerative diseases through induction of ATF4. In this study, we demonstrate that the PERK pathway was activated in neurons during EAE. We found that neuron-specific PERK inactivation impaired EAE resolution and exacerbated EAE-induced axon degeneration, neuron loss, and demyelination. Surprisingly, neuron-specific ATF4 inactivation did not alter EAE disease course or EAE-induced axon degeneration, neuron loss, and demyelination. These results suggest that PERK activation in neurons protects axons and neurons against inflammation in MS and EAE through ATF4-independent mechanisms.
| S-EPMC6413796 | biostudies-literature
Project description:The progressive loss of neurons causes neurodegenerative diseases. Because the accumulation of DNA breaks results in neuronal apoptosis, the lack of a variety of DNA damage repair-related proteins contributes to neurodegeneration. The ubiquitin ligase RNF8 plays an important role in DNA double-strand break repair via histone ubiquitination. However, the function of RNF8 in terminally differentiated neurons remains unknown. This study aimed to determine whether RNF8 is involved in the DNA damage response in neurons and contributes to neurodegeneration. Here, we present evidence suggesting that RNF8 deficiency results in DNA damage accumulation and neuronal apoptosis. RNF8(-/-) mice exhibit neuronal degeneration and reactive astrocytosis. Neurons from RNF8(-/-) mice appear to be more susceptible to X-ray-induced DNA damage. These changes were consistent with the behavioral performances of the RNF8-deficient mice, which included impaired performances in the open-field test and step-down avoidance task. Overall, these findings show that RNF8 is required for DNA damage repair in neurons. RNF8 deficiency is sufficient to cause neuronal pathology and cognitive decline, and the loss of RNF8 results in neuron degeneration.
| S-EPMC4851709 | biostudies-literature
Project description:We generated a mouse model (cKO) with a conditional deletion of TGF-beta signaling in the retinal neurons by crossing TGF-beta receptor I (TGF-beta RI) floxed mice with nestin-Cre mice. Almost all of the newborn cKO mice had retinal detachment at the retinal pigment epithelium (RPE)/photoreceptor layer junction of the neurosensory retina (NSR). The immunostaining for chondroitin-6-sulfate showed a very weak reaction in cKO mice in contrast to intense staining in the photoreceptor layer in wild-type mice. Macroscopic cataracts, in one or both eyes, were observed in 50% of the mice by 6 months of age, starting as early as the first month after birth. The cKO mouse model demonstrates that the TGF-beta signaling deficiency in retinal cells leads to decreased levels of chondroitin sulfate proteoglycan in the retinal interphotoreceptor matrix. This in turn causes retinal detachment due to the loss of adhesion of the NSR to RPE.
| S-EPMC2062516 | biostudies-literature
Project description:Alzheimer's disease (AD) is a neurodegenerative disease caused by accumulation of amyloid beta (A?) plaque and neurofibrillary tangle formation. We have shown in vitro, that knock-down and blockade of the 37 kDa/67 kDa Laminin Receptor (LRP/LR) resulted in reduced A? induced cytotoxicity and A? accumulation. In order to test the effect of blocking LRP/LR on A? formation and AD associated symptoms, AD transgenic mice received the anti-LRP/LR specific antibody, IgG1-iS18 through intranasal administration. We show that this treatment resulted in an improvement in memory, and decreased A? plaque formation. Moreover, a significant decrease in A?42 protein expression with a concomitant increase in amyloid precursor protein (APP) and telomerase reverse transcriptase (mTERT) levels was observed. These data recommend IgG1-iS18 as a potentially powerful therapeutic antibody for AD treatment.
| S-EPMC6007457 | biostudies-literature
Project description:Krabbe disease is a lysosomal storage disease caused by mutations in the gene that encodes galactosylceramidase, in which galactosylsphingosine (psychosine) accumulation drives demyelination in the central and peripheral nervous systems, ultimately progressing to death in early childhood. Gene therapy, alone or in combination with transplant, has been developed for almost two decades in mouse models, with increasing therapeutic benefit paralleling the improvement of next-generation adeno-associated virus (AAV) vectors. This effort has recently shown remarkable efficacy in the canine model of the disease by two different groups that used either systemic or cerebrospinal fluid (CSF) administration of AAVrh10 or AAV9. Building on our experience developing CSF-delivered, AAV-based drug products for a variety of neurodegenerative disorders, we conducted efficacy, pharmacology, and safety studies of AAVhu68 delivered to the CSF in two relevant natural Krabbe animal models, and in nonhuman primates. In newborn Twitcher mice, the highest dose (1 × 1011 genome copies [GC]) of AAVhu68.hGALC injected into the lateral ventricle led to a median survival of 130 days compared to 40.5 days in vehicle-treated mice. When this dose was administered intravenously, the median survival was 49 days. A single intracisterna magna injection of AAVhu68.cGALC at 3 × 1013 GC into presymptomatic Krabbe dogs increased survival for up to 85 weeks compared to 12 weeks in controls. It prevented psychosine accumulation in the CSF, preserved peripheral nerve myelination, ambulation, and decreased brain neuroinflammation and demyelination, although some regions remained abnormal. In a Good Laboratory Practice-compliant toxicology study, we administered the clinical candidate into the cisterna magna of 18 juvenile rhesus macaques at 3 doses that displayed efficacy in mice. We observed no dose-limiting toxicity and sporadic minimal degeneration of dorsal root ganglia (DRG) neurons. Our studies demonstrate the efficacy, scalability, and safety of a single cisterna magna AAVhu68 administration to treat Krabbe disease. ClinicalTrials.Gov ID: NCT04771416.
| S-EPMC9142772 | biostudies-literature
Project description:BackgroundPantothenate kinase-associated neurodegeneration, PKAN, is an inherited disorder characterized by progressive impairment in motor coordination and caused by mutations in PANK2, a human gene that encodes one of four pantothenate kinase (PanK) isoforms. PanK initiates the synthesis of coenzyme A (CoA), an essential cofactor that plays a key role in energy metabolism and lipid synthesis. Most of the mutations in PANK2 reduce or abolish the activity of the enzyme. This evidence has led to the hypothesis that lower CoA might be the underlying cause of the neurodegeneration in PKAN patients; however, no mouse model of the disease is currently available to investigate the connection between neuronal CoA levels and neurodegeneration. Indeed, genetic and/or dietary manipulations aimed at reducing whole-body CoA synthesis have not produced a desirable PKAN model, and this has greatly hindered the discovery of a treatment for the disease.Objective, methods, results and conclusionsCellular CoA levels are tightly regulated by a balance between synthesis and degradation. CoA degradation is catalyzed by two peroxisomal nudix hydrolases, Nudt7 and Nudt19. In this study we sought to reduce neuronal CoA in mice through the alternative approach of increasing Nudt7-mediated CoA degradation. This was achieved by combining the use of an adeno-associated virus-based expression system with the synapsin (Syn) promoter. We show that mice with neuronal overexpression of a cytosolic version of Nudt7 (scAAV9-Syn-Nudt7cyt) exhibit a significant decrease in brain CoA levels in conjunction with a reduction in motor coordination. These results strongly support the existence of a link between CoA levels and neuronal function and show that scAAV9-Syn-Nudt7cyt mice can be used to model PKAN.
| S-EPMC4460045 | biostudies-literature