Mutations in the glucocerebrosidase (gba) gene cause Gaucher disease (GD), the most common lysosomal storage disorder, and increase susceptibility to Parkinson’s disease (PD). While the clinical and ...pathological features of idiopathic PD and PD related to gba (PD-GBA) mutations are very similar, cellular mechanisms underlying neurodegeneration in each are unclear. Using a mouse model of neuronopathic GD, we show that autophagic machinery and proteasomal machinery are defective in neurons and astrocytes lacking gba. Markers of neurodegeneration—p62/SQSTM1, ubiquitinated proteins, and insoluble α-synuclein—accumulate. Mitochondria were dysfunctional and fragmented, with impaired respiration, reduced respiratory chain complex activities, and a decreased potential maintained by reversal of the ATP synthase. Thus a primary lysosomal defect causes accumulation of dysfunctional mitochondria as a result of impaired autophagy and dysfunctional proteasomal pathways. These data provide conclusive evidence for mitochondrial dysfunction in GD and provide insight into the pathogenesis of PD and PD-GBA.
Display omitted
•Autophagic and proteasomal pathways are suppressed in gba knockout mice•α-Synuclein accumulates and forms deposits in gba knockout mouse brainstem•Neurons and astrocytes from gba knockout mice harbor dysfunctional mitochondria•Mitochondria do not recruit Parkin and accumulate in gba knockout neurons
Full text
Available for:
GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Over the last decade, pioneering liver-directed gene therapy trials for haemophilia B have achieved sustained clinical improvement after a single systemic injection of adeno-associated virus (AAV) ...derived vectors encoding the human factor IX cDNA. These trials demonstrate the potential of AAV technology to provide long-lasting clinical benefit in the treatment of monogenic liver disorders. Indeed, with more than ten ongoing or planned clinical trials for haemophilia A and B and dozens of trials planned for other inherited genetic/metabolic liver diseases, clinical translation is expanding rapidly. Gene therapy is likely to become an option for routine care of a subset of severe inherited genetic/metabolic liver diseases in the relatively near term. In this review, we aim to summarise the milestones in the development of gene therapy, present the different vector tools and their clinical applications for liver-directed gene therapy. AAV-derived vectors are emerging as the leading candidates for clinical translation of gene delivery to the liver. Therefore, we focus on clinical applications of AAV vectors in providing the most recent update on clinical outcomes of completed and ongoing gene therapy trials and comment on the current challenges that the field is facing for large-scale clinical translation. There is clearly an urgent need for more efficient therapies in many severe monogenic liver disorders, which will require careful risk-benefit analysis for each indication, especially in paediatrics.
Full text
Available for:
FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OBVAL, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
The potential of induced pluripotent stem cells (iPSCs) in disease modeling and regenerative medicine is vast, but current methodologies remain inefficient. Understanding the cellular mechanisms ...underlying iPSC reprogramming, such as the metabolic shift from oxidative to glycolytic energy production, is key to improving its efficiency. We have developed a lentiviral reporter system to assay longitudinal changes in cell signaling and transcription factor activity in living cells throughout iPSC reprogramming of human dermal fibroblasts. We reveal early NF-κB, AP-1, and NRF2 transcription factor activation prior to a temporal peak in hypoxia inducible factor α (HIFα) activity. Mechanistically, we show that an early burst in oxidative phosphorylation and elevated reactive oxygen species generation mediates increased NRF2 activity, which in turn initiates the HIFα-mediated glycolytic shift and may modulate glucose redistribution to the pentose phosphate pathway. Critically, inhibition of NRF2 by KEAP1 overexpression compromises metabolic reprogramming and results in reduced efficiency of iPSC colony formation.
Display omitted
•Cells increase proliferation, OXPHOS, and ROS production early in reprogramming•The antioxidant response is therefore active at this stage, prior to HIFα activation•NRF2 promotes HIFα activation, the metabolic switch, and colony formation•NRF2 activation is concomitant with glucose redistribution to the PPP
Hawkins et al. examine the metabolic shift during iPSC reprogramming. They propose that increased proliferation of cells driven by transgene expression can lead to increased oxidative phosphorylation resulting in ROS production. Elevated ROS activates NRF2, promoting HIFα activation and the switch to glycolysis.
Full text
Available for:
GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
•The role of bioluminescent imaging in models of infection is discussed.•Light-producing reporter animals are used to study host response to infection.•Methodology for developing neonatal somatic ...light producing reporter mice described.
Infectious diseases are one of the leading causes of death worldwide. Modelling and understanding human infection is imperative to developing treatments to reduce the global burden of infectious disease. Bioluminescence imaging is a highly sensitive, non-invasive technique based on the detection of light, produced by luciferase-catalysed reactions. In the study of infectious disease, bioluminescence imaging is a well-established technique; it can be used to detect, localize and quantify specific immune cells, pathogens or immunological processes. This enables longitudinal studies in which the spectrum of the disease process and its response to therapies can be monitored. Light producing transgenic rodents are emerging as key tools in the study of host response to infection. Here, we review the strategies for identifying biological processes in vivo, including the technology of bioluminescence imaging and illustrate how this technique is shedding light on the host-pathogen relationship.
Full text
Available for:
GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
Preterm birth is a serious global health problem and the leading cause of infant death before 5 years of age. At least 40% of cases are associated with infection. The most common way for pathogens to ...access the uterine cavity is by ascending from the vagina. Bioluminescent pathogens have revolutionized the understanding of infectious diseases. We hypothesized that bioluminescent Escherichia coli can be used to track and monitor ascending vaginal infections. Two bioluminescent strains were studied: E. coli K12 MG1655-lux, a nonpathogenic laboratory strain, and E. coli K1 A192PP-lux2, a pathogenic strain capable of causing neonatal meningitis and sepsis in neonatal rats. On embryonic day 16, mice received intravaginal E. coli K12, E. coli K1, or phosphate-buffered saline followed by whole-body bioluminescent imaging. In both cases, intravaginal delivery of E. coli K12 or E. coli K1 led to bacterial ascension into the uterine cavity, but only E. coli K1 induced preterm parturition. Intravaginal administration of E. coli K1 significantly reduced the proportion of pups born alive compared with E. coli K12 and phosphate-buffered saline controls. However, in both groups of viable pups born after bacterial inoculation, there was evidence of comparable brain inflammation by postnatal day 6. This study ascribes specific mechanisms by which exposure to intrauterine bacteria leads to premature delivery and neurologic inflammation in neonates.
Full text
Available for:
GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
ABSTRACT
Activation of uterine inflammatory pathways leads to preterm labor (PTL), associated with high rates of neonatal mortality and morbidity. The transcription factors nuclear factor κB (NFκB) ...and activator protein 1 (AP‐1) regulate key proinflammatory and procontractile genes involved in normal labor and PTL. Here we show that (NFκB) activation normally occurs in the mouse myometrium at gestation day E18, prior to labor, whereas AP‐1 and JNK activation occurs at labor onset. Where labor was induced using the progesterone receptor antagonist RU486, NFkB and AP‐1/JNK activation both occurred at the time of labor (20 h compared to 60 h in DMSO‐treated controls). Using an LPS (Escherichia coli: serotype O111)‐induced PTL model that selectively activates AP‐1 but not NFkB, we show that myometrial AP‐1 activation drives production of cytokines (Il‐6, Il‐8, and Il‐1β), metalloproteinases (Mmp3 and Mmp10), and procontractile proteins (Cox‐2 and Cx43) resulting in PTL after 7 h. Protein levels of CX43 and IL‐1β, and IL‐1β cleavage, were increased following LPS‐induced activation of AP‐1. Inhibition of JNK by SP600125 (30 mg/kg) delayed PTL by 6 h (7.5 vs. 13.5 h P<0.05). Our data reveal that (NFκB) activation is not a functional requirement for infection/inflammation‐induced preterm labor and that AP‐1 activation is sufficient to drive inflammatory pathways that cause PTL.—MacIntyre, D. A., Lee, Y. S., Migale, R., Herbert, B. R., Waddington, S. N., Peebles, D., Hagberg, H., Johnson, M. R., Bennett, P. R. Activator protein 1 is a key terminal mediator of inflammation‐induced preterm labor in mice. FASEB J. 28, 2358–2368 (2014). www.fasebj.org
Full text
Available for:
BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Parkinson's disease (PD) is a progressive late-onset neurodegenerative disease leading to physical and cognitive decline. Mutations of leucine-rich repeat kinase 2 (LRRK2) are the most common genetic ...cause of PD. LRRK2 is a complex scaffolding protein with known regulatory roles in multiple molecular pathways. Two prominent examples of LRRK2-modulated pathways are Wingless/Int (Wnt) and nuclear factor of activated T-cells (NFAT) signaling. Both are well described key regulators of immune and nervous system development as well as maturation. The aim of this study was to establish the physiological and pathogenic role of LRRK2 in Wnt and NFAT signaling in the brain, as well as the potential contribution of the non-canonical Wnt/Calcium pathway. In vivo cerebral Wnt and NFATc1 signaling activity was quantified in LRRK2 G2019S mutant knock-in (KI) and LRRK2 knockout (KO) male and female mice with repeated measures over 28 weeks, employing lentiviral luciferase biosensors, and analyzed using a mixed-effect model. To establish spatial resolution, we investigated tissues, and primary neuronal cell cultures from different brain regions combining luciferase signaling activity, immunohistochemistry, qPCR and western blot assays. Results were analyzed by unpaired t-test with Welch's correction or 2-way ANOVA with post hoc corrections. In vivo Wnt signaling activity in LRRK2 KO and LRRK2 G2019S KI mice was increased significantly ~ threefold, with a more pronounced effect in males (~ fourfold) than females (~ twofold). NFATc1 signaling was reduced ~ 0.5-fold in LRRK2 G2019S KI mice. Brain tissue analysis showed region-specific expression changes in Wnt and NFAT signaling components. These effects were predominantly observed at the protein level in the striatum and cerebral cortex of LRRK2 KI mice. Primary neuronal cell culture analysis showed significant genotype-dependent alterations in Wnt and NFATc1 signaling under basal and stimulated conditions. Wnt and NFATc1 signaling was primarily dysregulated in cortical and hippocampal neurons respectively. Our study further built on knowledge of LRRK2 as a Wnt and NFAT signaling protein. We identified complex changes in neuronal models of LRRK2 PD, suggesting a role for mutant LRRK2 in the dysregulation of NFAT, and canonical and non-canonical Wnt signaling.
Full text
Available for:
IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
9.
Gene therapy for neurotransmitter‐related disorders Chu, Wing Sum; Ng, Joanne; Waddington, Simon N. ...
Journal of inherited metabolic disease,
January 2024, 2024-Jan, 2024-01-00, 20240101, Volume:
47, Issue:
1
Journal Article
Peer reviewed
Open access
Inborn errors of neurotransmitter (NT) metabolism are a group of rare, heterogenous diseases with predominant neurological features, such as movement disorders, autonomic dysfunction, and ...developmental delay. Clinical overlap with other disorders has led to delayed diagnosis and treatment, and some conditions are refractory to oral pharmacotherapies. Gene therapies have been developed and translated to clinics for paediatric inborn errors of metabolism, with 38 interventional clinical trials ongoing to date. Furthermore, efforts in restoring dopamine synthesis and neurotransmission through viral gene therapy have been developed for Parkinson's disease. Along with the recent European Medicines Agency (EMA) and Medicines and Healthcare Products Regulatory Agency (MHRA) approval of an AAV2 gene supplementation therapy for AADC deficiency, promising efficacy and safety profiles can be achieved in this group of diseases. In this review, we present preclinical and clinical advances to address NT‐related diseases, and summarise potential challenges that require careful considerations for NT gene therapy studies.
Full text
Available for:
FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
For inherited genetic diseases, fetal gene therapy offers the potential of prophylaxis against early, irreversible and lethal pathological change. To explore this, we studied neuronopathic Gaucher ...disease (nGD), caused by mutations in GBA. In adult patients, the milder form presents with hepatomegaly, splenomegaly and occasional lung and bone disease; this is managed, symptomatically, by enzyme replacement therapy. The acute childhood lethal form of nGD is untreatable since enzyme cannot cross the blood-brain barrier. Patients with nGD exhibit signs consistent with hindbrain neurodegeneration, including neck hyperextension, strabismus and, often, fatal apnea
. We selected a mouse model of nGD carrying a loxP-flanked neomycin disruption of Gba plus Cre recombinase regulated by the keratinocyte-specific K14 promoter. Exclusive skin expression of Gba prevents fatal neonatal dehydration. Instead, mice develop fatal neurodegeneration within 15 days
. Using this model, fetal intracranial injection of adeno-associated virus (AAV) vector reconstituted neuronal glucocerebrosidase expression. Mice lived for up to at least 18 weeks, were fertile and fully mobile. Neurodegeneration was abolished and neuroinflammation ameliorated. Neonatal intervention also rescued mice but less effectively. As the next step to clinical translation, we also demonstrated the feasibility of ultrasound-guided global AAV gene transfer to fetal macaque brains.
PDF
