Cholesterol redistribution triggered by CYP46A1 gene therapy improves major hallmarks of Niemann-Pick type C disease but is not sufficient to halt neurodegeneration.

Cholesterol 24-hydroxylase (CYP46A1) is an exclusively neuronal cytochrome P450 enzyme responsible for converting cholesterol into 24S-hydroxycholesterol, which serves as the primary pathway for eliminating cholesterol in the brain. We and others have shown that increased activity of CYP46A1 leads to reduced levels of cholesterol and has a positive effect on cognition. Therefore, we hypothesized that CYP46A1 could be a potential therapeutic target in Niemann-Pick type C (NPC) disease, a rare and fatal neurodegenerative disorder, characterized by cholesterol accumulation in endolysosomal compartments. Herein, we show that CYP46A1 ectopic expression, in cellular models of NPC and in Npc1tm(I1061T) mice by adeno-associated virus-mediated gene therapy improved NPC disease phenotype. Amelioration in functional, biochemical, molecular and neuropathological hallmarks of NPC disease were characterized. In vivo, CYP46A1 expression partially prevented weight loss and hepatomegaly, corrected the expression levels of genes involved in cholesterol homeostasis, and promoted a redistribution of brain cholesterol accumulated in late endosomes/lysosomes. Moreover, concomitant with the amelioration of cholesterol metabolism dysregulation, CYP46A1 attenuated microgliosis and lysosomal dysfunction in mouse cerebellum, favoring a pro-resolving phenotype. In vivo CYP46A1 ectopic expression improves important features of NPC disease and may represent a valid therapeutic approach to be used concomitantly with other drugs. However, promoting cholesterol redistribution does not appear to be enough to prevent Purkinje neuronal death in the cerebellum. This indicates that cholesterol buildup in neurons might not be the main cause of neurodegeneration in this human lipidosis.

Positron Emission Tomography Quantitative Assessment of Off-Target Whole-Body Biodistribution of I-124-Labeled Adeno-Associated Virus Capsids Administered to Cerebral Spinal Fluid.

Based on studies in experimental animals demonstrating that administration of adeno-associated virus (AAV) vectors to the cerebrospinal fluid (CSF) is an effective route to transfer genes to the nervous system, there are increasing number of clinical trials using the CSF route to treat nervous system disorders. With the knowledge that the CSF turns over four to five times daily, and evidence in experimental animals that at least some of CSF administered AAV vectors are distributed to systemic organs, we asked: with AAV administration to the CSF, what fraction of the total dose remains in the nervous system and what fraction goes off target and is delivered systemically? To quantify the biodistribution of AAV capsids immediately after administration, we covalently labeled AAV capsids with iodine 124 (I-124), a cyclotron generated positron emitter, enabling quantitative positron emission tomography scanning of capsid distribution for up to 96 h after AAV vector administration. We assessed the biodistribution to nonhuman primates of I-124-labeled capsids from different AAV clades, including 9 (clade F), rh.10 (E), PHP.eB (F), hu68 (F), and rh91(A). The analysis demonstrated that 60-90% of AAV vectors administered to the CSF through either the intracisternal or intrathecal (lumbar) routes distributed systemically to major organs. These observations have potentially significant clinical implications regarding accuracy of AAV vector dosing to the nervous system, evoking systemic immunity at levels similar to that with systemic administration, and potential toxicity of genes designed to treat nervous system disorders being expressed in non-nervous system organs. Based on these data, individuals in clinical trials using AAV vectors administered to the CSF should be monitored for systemic as well as nervous system adverse events and CNS dosing considerations should account for a significant AAV systemic distribution.

Bioelectronic cell-based device provides a strategy for the treatment of the experimental model of multiple sclerosis.

Wireless powered optogenetic cell-based implant provides a strategy to deliver subcutaneously therapeutic proteins. Immortalize Human Mesenchymal Stem Cells (hMSC-TERT) expressing the bacteriophytochrome diguanylate cyclase (DGCL) were validated for optogenetic controlled interferon-ß delivery (Optoferon cells) in a bioelectronic cell-based implant. Optoferon cells transcriptomic profiling was used to elaborate an in-silico model of the recombinant interferon-ß production. Wireless optoelectronic device integration was developed using additive manufacturing and injection molding. Implant cell-based optoelectronic interface manufacturing was established to integrate industrial flexible compact low-resistance screen-printed Near Field Communication (NFC) coil antenna. Optogenetic cell-based implant biocompatibility, and device performances were evaluated in the Experimental Autoimmune Encephalomyelitis (EAE) mouse model of multiple sclerosis.

Genetically modified macrophages accelerate myelin repair.

Preventing neurodegeneration-associated disability progression in patients with multiple sclerosis (MS) remains an unmet therapeutic need. As remyelination prevents axonal degeneration, promoting this process in patients might enhance neuroprotection. In demyelinating mouse lesions, local overexpression of semaphorin 3F (Sema3F), an oligodendrocyte progenitor cell (OPC) attractant, increases remyelination. However, molecular targeting to MS lesions is a challenge. A clinically relevant paradigm for delivering Sema3F to demyelinating lesions could be to use blood-derived macrophages as vehicles. Thus, we chose transplantation of genetically modified hematopoietic stem cells (HSCs) as means of obtaining chimeric mice with circulating Sema3F-overexpressing monocytes. We demonstrated that Sema3F-transduced HSCs stimulate OPC migration in a neuropilin 2 (Nrp2, Sema3F receptor)-dependent fashion, which was conserved in middle-aged OPCs. While demyelinating lesions induced in mice with Sema3F-expressing blood cells showed no changes in inflammation and OPC survival, OPC recruitment was enhanced which accelerated the onset of remyelination. Our results provide a proof of concept that blood cells, particularly monocytes/macrophages, can be used to deliver pro-remyelinating agents "at the right time and place," suggesting novel means for remyelination-promoting strategies in MS.

Hematopoietic stem cell transplantation chemotherapy causes microglia senescence and peripheral macrophage engraftment in the brain.

Hematopoietic stem cell transplantation (HSCT) is a therapy used for multiple malignant and nonmalignant diseases, with chemotherapy used for pretransplantation myeloablation. The post-HSCT brain contains peripheral engrafted parenchymal macrophages, despite their absence in the normal brain, with the engraftment mechanism still undefined. Here we show that HSCT chemotherapy broadly disrupts mouse brain regenerative populations, including a permanent loss of adult neurogenesis. Microglial density was halved, causing microglial process expansion, coinciding with indicators of broad senescence. Although microglia expressed cell proliferation markers, they underwent cell cycle arrest in S phase with a majority expressing the senescence and antiapoptotic marker p21. In vivo single-cell tracking of microglia after recovery from chemical depletion showed loss of their regenerative capacity, subsequently replaced with donor macrophages. We propose that HSCT chemotherapy causes microglial senescence with a gradual decrease to a critical microglial density, providing a permissive niche for peripheral macrophage engraftment of the brain.

Cholesterol Hydroxylating Cytochrome P450 46A1: From Mechanisms of Action to Clinical Applications.

Cholesterol, an essential component of the brain, and its local metabolism are involved in many neurodegenerative diseases. The blood-brain barrier is impermeable to cholesterol; hence, cholesterol homeostasis in the central nervous system represents a balance between in situ biosynthesis and elimination. Cytochrome P450 46A1 (CYP46A1), a central nervous system-specific enzyme, converts cholesterol to 24-hydroxycholesterol, which can freely cross the blood-brain barrier and be degraded in the liver. By the dual action of initiating cholesterol efflux and activating the cholesterol synthesis pathway, CYP46A1 is the key enzyme that ensures brain cholesterol turnover. In humans and mouse models, CYP46A1 activity is altered in Alzheimer's and Huntington's diseases, spinocerebellar ataxias, glioblastoma, and autism spectrum disorders. In mouse models, modulations of CYP46A1 activity mitigate the manifestations of Alzheimer's, Huntington's, Nieman-Pick type C, and Machao-Joseph (spinocerebellar ataxia type 3) diseases as well as amyotrophic lateral sclerosis, epilepsy, glioblastoma, and prion infection. Animal studies revealed that the CYP46A1 activity effects are not limited to cholesterol maintenance but also involve critical cellular pathways, like gene transcription, endocytosis, misfolded protein clearance, vesicular transport, and synaptic transmission. How CYP46A1 can exert central control of such essential brain functions is a pressing question under investigation. The potential therapeutic role of CYP46A1, demonstrated in numerous models of brain disorders, is currently being evaluated in early clinical trials. This review summarizes the past 70 years of research that has led to the identification of CYP46A1 and brain cholesterol homeostasis as powerful therapeutic targets for severe pathologies of the CNS.

Optimized Protocol for Subcutaneous Implantation of Encapsulated Cells Device and Evaluation of Biocompatibility.

Improving a drug delivery system is critical to treat central nervous system disorders. Here we studied an innovative approach based on implantation of a wireless-powered cell-based device in mice. This device, coupling biologic material and electronics, is the first of its kind. The advantage of this technology is its ability to control the secretion of a therapeutic molecule and to switch the classical permanent delivery to activation on demand. In diseases with relapsing-remitting phases such as multiple sclerosis, such activation could be selectively achieved in relapsing phases. However, the safety (tolerance to biomaterials and surgical procedure) of such a clinical device needs to be verified. Therefore, the development of tools to assess the biocompatibility of the system in animal models is an essential step. We present the development of this new therapeutic approach, the challenges we encountered during the different steps of its development (such as cell loading in the chamber, surgery protocol for subcutaneous implantation of the device) and the tools we used to evaluate cell viability and biocompatibility of the device.

Complete Correction of Brain and Spinal Cord Pathology in Metachromatic Leukodystrophy Mice.

Metachromatic leukodystrophy (MLD) is a lysosomal storage disorder characterized by accumulation of sulfatides in both glial cells and neurons. MLD results from an inherited deficiency of arylsulfatase A (ARSA) and myelin degeneration in the central and peripheral nervous systems. Currently, no effective treatment is available for the most frequent late infantile (LI) form of MLD after symptom onset. The LI form results in rapid neurological degradation and early death. ARSA enzyme must be rapidly and efficiently delivered to brain and spinal cord oligodendrocytes of patients with LI MLD in order to potentially stop the progression of the disease. We previously showed that brain gene therapy with adeno-associated virus serotype rh10 (AAVrh10) driving the expression of human ARSA cDNA alleviated most long-term disease manifestations in MLD mice but was not sufficient in MLD patient to improve disease progression. Herein, we evaluated the short-term effects of intravenous AAVPHP.eB delivery driving the expression of human ARSA cDNA under the control of the cytomegalovirus/b-actin hybrid (CAG) promoter in 6-month-old MLD mice that already show marked sulfatide accumulation and brain pathology. Within 3 months, a single intravenous injection of AAVPHP.eB-hARSA-HA resulted in correction of brain and spinal cord sulfatide storage, and improvement of astrogliosis and microgliosis in brain and spinal cord of treated animals. These results strongly support to consider the use of AAVPHP.eB-hARSA vector for intravenous gene therapy in symptomatic rapidly progressing forms of MLD.

The Challenge of Gene Therapy for Neurological Diseases: Strategies and Tools to Achieve Efficient Delivery to the Central Nervous System.

For more than 10 years, gene therapy for neurological diseases has experienced intensive research growth and more recently therapeutic interventions for multiple indications. Beneficial results in several phase 1/2 clinical studies, together with improved vector technology have advanced gene therapy for the central nervous system (CNS) in a new era of development. Although most initial strategies have focused on orphan genetic diseases, such as lysosomal storage diseases, more complex and widespread conditions like Alzheimer's disease, Parkinson's disease, epilepsy, or chronic pain are increasingly targeted for gene therapy. Increasing numbers of applications and patients to be treated will require improvement and simplification of gene therapy protocols to make them accessible to the largest number of affected people. Although vectors and manufacturing are a major field of academic research and industrial development, there is a growing need to improve, standardize, and simplify delivery methods. Delivery is the major issue for CNS therapies in general, and particularly for gene therapy. The blood-brain barrier restricts the passage of vectors; strategies to bypass this obstacle are a central focus of research. In this study, we present the different ways that can be used to deliver gene therapy products to the CNS. We focus on results obtained in large animals that have allowed the transfer of protocols to human patients and have resulted in the generation of clinical data. We discuss the different routes of administration, their advantages, and their limitations. We describe techniques, equipment, and protocols and how they should be selected for safe delivery and improved efficiency for the next generation of gene therapy trials for CNS diseases.

Intra-CSF AAV9 and AAVrh10 Administration in Nonhuman Primates: Promising Routes and Vectors for Which Neurological Diseases?

The identification of the most efficient method for whole central nervous system targeting that is translatable to humans and the safest route of adeno-associated virus (AAV) administration is a major concern for future applications in clinics. Additionally, as many AAV serotypes were identified for gene introduction into the brain and the spinal cord, another key to human gene-therapy success is to determine the most efficient serotype. In this study, we compared lumbar intrathecal administration through catheter implantation and intracerebroventricular administration in the cynomolgus macaque. We also evaluated and compared two AAV serotypes that are currently used in clinical trials: AAV9 and AAVrh10. We demonstrated that AAV9 lumbar intrathecal delivery using a catheter achieved consistent transgene expression in the motor neurons of the spinal cord and in the neurons/glial cells of several brain regions, whereas AAV9 intracerebroventricular delivery led to a consistent transgene expression in the brain. In contrast, AAVrh10 lumbar intrathecal delivery led to rare motor neuron targeting. Finally, we found that AAV9 efficiently targets respiratory and skeletal muscles after injection into the cerebrospinal fluid (CSF), which represents an outstanding new property that can be useful for the treatment of diseases affecting both the central nervous system and muscle.

The cholesterol 24-hydroxylase activates autophagy and decreases mutant huntingtin build-up in a neuroblastoma culture model of Huntington's disease.

OBJECTIVE: Compromised brain cholesterol turnover and altered regulation of brain cholesterol metabolism have been allied with some neurodegenerative diseases, including Huntington's disease (HD). Following our previous studies in HD, in this study we aim to investigate in vitro in a neuroblastoma cellular model of HD, the effect of CYP46A1 overexpression, an essential enzyme in cholesterol metabolism, on huntingtin aggregation and levels. RESULTS: We found that CYP46A1 reduces the quantity and size of mutant huntingtin aggregates in cells, as well as the levels of mutant huntingtin protein. Additionally, our results suggest that the observed beneficial effects of CYP46A1 in HD cells are linked to the activation of autophagy. Taken together, our results further demonstrate that CYP46A1 is a pertinent target to counteract HD progression.

Real-Time Monitoring of Exosome Enveloped-AAV Spreading by Endomicroscopy Approach: A New Tool for Gene Delivery in the Brain.

Exosomes represent a strategy for optimizing the adeno-associated virus (AAV) toward the development of novel therapeutic options for neurodegenerative disorders. However, in vivo spreading of exosomes and AAVs after intracerebral administration is poorly understood. This study provides an assessment and comparison of the spreading into the brain of exosome-enveloped AAVs (exo-AAVs) or unassociated AAVs (std-AAVs) through in vivo optical imaging techniques like probe-based confocal laser endomicroscopy (pCLE) and ex vivo fluorescence microscopy. The std-AAV serotypes (AAV6 and AAV9) encoding the GFP were enveloped in exosomes and injected into the ipsilateral hippocampus. At 3 months post-injection, pCLE detected enhanced GFP expression of both exo-AAV serotypes in contralateral hemispheres compared to std-AAVs. Although sparse GFP-positive astrocytes were observed using exo-AAVs, our results show that the enhancement of the transgene expression resulting from exo-AAVs was largely restricted to neurons and oligodendrocytes. Our results suggest (1) the possibility of combining gene therapy with an endoscopic approach to enable tracking of exo-AAV spread, and (2) exo-AAVs allow for widespread, long-term gene expression in the CNS, supporting the use of exo-AAVs as an efficient gene delivery tool.

CYP46A1 gene therapy deciphers the role of brain cholesterol metabolism in Huntington's disease.

Dysfunctions in brain cholesterol homeostasis have been extensively related to brain disorders. The main pathway for brain cholesterol elimination is its hydroxylation into 24S-hydroxycholesterol by the cholesterol 24-hydrolase, CYP46A1. Increasing evidence suggests that CYP46A1 has a role in the pathogenesis and progression of neurodegenerative disorders, and that increasing its levels in the brain is neuroprotective. However, the mechanisms underlying this neuroprotection remain to be fully understood. Huntington's disease is a fatal autosomal dominant neurodegenerative disease caused by an abnormal CAG expansion in huntingtin's gene. Among the multiple cellular and molecular dysfunctions caused by this mutation, altered brain cholesterol homeostasis has been described in patients and animal models as a critical event in Huntington's disease. Here, we demonstrate that a gene therapy approach based on the delivery of CYP46A1, the rate-limiting enzyme for cholesterol degradation in the brain, has a long-lasting neuroprotective effect in Huntington's disease and counteracts multiple detrimental effects of the mutated huntingtin. In zQ175 Huntington's disease knock-in mice, CYP46A1 prevented neuronal dysfunctions and restored cholesterol homeostasis. These events were associated to a specific striatal transcriptomic signature that compensates for multiple mHTT-induced dysfunctions. We thus explored the mechanisms for these compensations and showed an improvement of synaptic activity and connectivity along with the stimulation of the proteasome and autophagy machineries, which participate to the clearance of mutant huntingtin (mHTT) aggregates. Furthermore, BDNF vesicle axonal transport and TrkB endosome trafficking were restored in a cellular model of Huntington's disease. These results highlight the large-scale beneficial effect of restoring cholesterol homeostasis in neurodegenerative diseases and give new opportunities for developing innovative disease-modifying strategies in Huntington's disease.

Restoring brain cholesterol turnover improves autophagy and has therapeutic potential in mouse models of spinocerebellar ataxia.

Spinocerebellar ataxias (SCAs) are devastating neurodegenerative disorders for which no curative or preventive therapies are available. Deregulation of brain cholesterol metabolism and impaired brain cholesterol turnover have been associated with several neurodegenerative diseases. SCA3 or Machado-Joseph disease (MJD) is the most prevalent ataxia worldwide. We show that cholesterol 24-hydroxylase (CYP46A1), the key enzyme allowing efflux of brain cholesterol and activating brain cholesterol turnover, is decreased in cerebellar extracts from SCA3 patients and SCA3 mice. We investigated whether reinstating CYP46A1 expression would improve the disease phenotype of SCA3 mouse models. We show that administration of adeno-associated viral vectors encoding CYP46A1 to a lentiviral-based SCA3 mouse model reduces mutant ataxin-3 accumulation, which is a hallmark of SCA3, and preserves neuronal markers. In a transgenic SCA3 model with a severe motor phenotype we confirm that cerebellar delivery of AAVrh10-CYP46A1 is strongly neuroprotective in adult mice with established pathology. CYP46A1 significantly decreases ataxin-3 protein aggregation, alleviates motor impairments and improves SCA3-associated neuropathology. In particular, improvement in Purkinje cell number and reduction of cerebellar atrophy are observed in AAVrh10-CYP46A1-treated mice. Conversely, we show that knocking-down CYP46A1 in normal mouse brain impairs cholesterol metabolism, induces motor deficits and produces strong neurodegeneration with impairment of the endosomal-lysosomal pathway, a phenotype closely resembling that of SCA3. Remarkably, we demonstrate for the first time both in vitro, in a SCA3 cellular model, and in vivo, in mouse brain, that CYP46A1 activates autophagy, which is impaired in SCA3, leading to decreased mutant ataxin-3 deposition. More broadly, we show that the beneficial effect of CYP46A1 is also observed with mutant ataxin-2 aggregates. Altogether, our results confirm a pivotal role for CYP46A1 and brain cholesterol metabolism in neuronal function, pointing to a key contribution of the neuronal cholesterol pathway in mechanisms mediating clearance of aggregate-prone proteins. This study identifies CYP46A1 as a relevant therapeutic target not only for SCA3 but also for other SCAs.

Inhibition of DYRK1A proteolysis modifies its kinase specificity and rescues Alzheimer phenotype in APP/PS1 mice.

Recent evidences suggest the involvement of DYRK1A (dual specificity tyrosine phosphorylation-regulated kinase 1 A) in Alzheimer's disease (AD). Here we showed that DYRK1A undergoes a proteolytic processing in AD patients hippocampus without consequences on its kinase activity. Resulting truncated forms accumulate in astrocytes and exhibit increased affinity towards STAT3ɑ, a regulator of inflammatory process. These findings were confirmed in APP/PS1 mice, an amyloid model of AD, suggesting that this DYRK1A cleavage is a consequence of the amyloid pathology. We identified in vitro the Leucettine L41 as a compound able to prevent DYRK1A proteolysis in both human and mouse protein extracts. We then showed that intraperitoneal injections of L41 in aged APP/PS1 mice inhibit STAT3ɑ phosphorylation and reduce pro-inflammatory cytokines levels (IL1- ß, TNF-ɑ and IL-12) associated to an increased microglial recruitment around amyloid plaques and decreased amyloid-ß plaque burden. Importantly, L41 treatment improved synaptic plasticity and rescued memory functions in APP/PS1 mice. Collectively, our results suggest that DYRK1A may contribute to AD pathology through its proteolytic process, reducing its kinase specificity. Further evaluation of inhibitors of DYRK1A truncation promises a new therapeutic approach for AD.

Transient increase in sAPPα secretion in response to Aß1-42 oligomers: an attempt of neuronal self-defense?

Amyloid precursor protein (APP), a key molecule of Alzheimer disease, is metabolized in 2 antagonist pathways generating the soluble APP alpha (sAPPα) having neuroprotective properties and the beta amyloid (Aß) peptide at the origin of neurotoxic oligomers, particularly Aß1-42. Whether extracellular Aß1-42 oligomers modulate the formation and secretion of sAPPα is not known. We report here that the addition of Aß1-42 oligomers to primary cortical neurons induced a transient increase in α-secretase activity and secreted sAPPα 6-9 hours later. Preventing the generation of sAPPα by using small interfering RNAs (siRNAs) for the α-secretases ADAM10 and ADAM17 or for APP led to increased Aß1-42 oligomer-induced cell death after 24 hours. Neuronal injuries due to oxidative stress or growth factor deprivation also generated sAPPα 7 hours later. Finally, acute injection of Aß1-42 oligomers into wild-type mouse hippocampi induced transient secretion of sAPPα 48-72 hours later. Altogether, these data suggest that neurons respond to stress by generating sAPPα for their survival. These data must be taken into account when interpreting sAPPα levels as a biomarker in neurological disorders.

ßAPP Processing Drives Gradual Tau Pathology in an Age-Dependent Amyloid Rat Model of Alzheimer's Disease.

The treatment of Alzheimer's disease (AD) remains challenging and requires a better in depth understanding of AD progression. Particularly, the link between amyloid protein precursor (APP) processing and Tau pathology development remains poorly understood. Growing evidences suggest that APP processing and amyloid-ß (Aß) release are upstream of Tau pathology but the lack of animal models mimicking the slow progression of human AD raised questions around this mechanism. Here, we described that an AD-like ßAPP processing in adults wild-type rats, yielding to human APP, ßCTF and Aß levels similar to those observed in AD patients, is sufficient to trigger gradual Tauopathy. The Tau hyperphosphorylation begins several months before the formation of both amyloid plaques and tangle-like aggregates in aged rats and without associated inflammation. Based on a longitudinal characterization over 30 months, we showed that extrasynaptic and emotional impairments appear before long-term potentiation deficits and memory decline and so before Aß and Tau aggregations. These compelling data allowed us to (1) experimentally confirm the causal relationship between ßAPP processing and Tau pathology in vivo and without Tau transgene overexpression, (2) support the amyloidogenic cascade and (3) propose a 4-step hypothesis of prodromal AD progression.

Clinical Gene Therapy for Neurodegenerative Diseases: Past, Present, and Future.

Clinical gene therapy has made important advances over the last decade. Among neurological diseases, severe genetic neurodegenerative conditions have been the focus of initial clinical applications. Gene therapy has also addressed complex neurodegenerative diseases, particularly Parkinson's disease, with encouraging results in human patients, demonstrating that specific targeting of central nervous system (CNS) cells is a relevant strategy for severe pathologies and that efficient access to the CNS with viral vectors is an achievable goal. The purpose of this review is to summarize the gene therapy clinical applications that have been conducted for neurodegenerative diseases. Limitations and hurdles to obtain and demonstrate benefit in patients, and the new developments that should allow new clinical applications with high beneficial potential are discussed.

Prevalence of pulmonary tuberculosis among prison inmates: A cross-sectional survey at the Correctional and Detention Facility of Abidjan, Côte d'Ivoire.

BACKGROUND: In Côte d'Ivoire, a TB prison program has been developed since 1999. This program includes offering TB screening to prisoners who show up with TB symptoms at the infirmary. Our objective was to estimate the prevalence of pulmonary TB among inmates at the Correctional and Detention Facility of Abidjan, the largest prison of Côte d'Ivoire, 16 years after this TB program was implemented. METHODS: Between March and September 2015, inmates, were screened for pulmonary TB using systematic direct smear microscopy, culture and chest X-ray. All participants were also proposed HIV testing. TB was defined as either confirmed (positive culture), probable (positive microscopy and/or chest X-ray findings suggestive of TB) or possible (signs or symptoms suggestive of TB, no X-Ray or microbiological evidence). Factors associated with confirmed tuberculosis were analysed using multivariable logistic regression. RESULTS: Among the 943 inmates screened, 88 (9.3%) met the TB case definition, including 19 (2.0%) with confirmed TB, 40 (4.2%) with probable TB and 29 (3.1%) with possible TB. Of the 19 isolated TB strains, 10 (53%) were TB drug resistant, including 7 (37%) with multi-resistance. Of the 10 patients with TB resistant strain, only one had a past history of TB treatment. HIV prevalence was 3.1% overall, and 9.6%among TB cases. Factors associated with confirmed TB were age ≥30 years (Odds Ratio 3.8; 95% CI 1.1-13.3), prolonged cough (Odds Ratio 3.6; 95% CI 1.3-9.5) and fever (Odds Ratio 2.7; 95% CI 1.0-7.5). CONCLUSION: In the country largest prison, pulmonary TB is still 10 (confirmed) to 44 times (confirmed, probable or possible) as frequent as in the Côte d'Ivoire general population, despite a long-time running symptom-based program of TB detection. Decreasing TB prevalence and limiting the risk of MDR may require the implementation of annual in-cell TB screening campaigns that systematically target all prison inmates.

Neuronal Cholesterol Accumulation Induced by Cyp46a1 Down-Regulation in Mouse Hippocampus Disrupts Brain Lipid Homeostasis.

Impairment in cholesterol metabolism is associated with many neurodegenerative disorders including Alzheimer's disease (AD). However, the lipid alterations underlying neurodegeneration and the connection between altered cholesterol levels and AD remains not fully understood. We recently showed that cholesterol accumulation in hippocampal neurons, induced by silencing Cyp46a1 gene expression, leads to neurodegeneration with a progressive neuronal loss associated with AD-like phenotype in wild-type mice. We used a targeted and non-targeted lipidomics approach by liquid chromatography coupled to high-resolution mass spectrometry to further characterize lipid modifications associated to neurodegeneration and cholesterol accumulation induced by CYP46A1 inhibition. Hippocampus lipidome of normal mice was profiled 4 weeks after cholesterol accumulation due to Cyp46a1 gene expression down-regulation at the onset of neurodegeneration. We showed that major membrane lipids, sphingolipids and specific enzymes involved in phosphatidylcholine and sphingolipid metabolism, were rapidly increased in the hippocampus of AAV-shCYP46A1 injected mice. This lipid accumulation was associated with alterations in the lysosomal cargoe, accumulation of phagolysosomes and impairment of endosome-lysosome trafficking. Altogether, we demonstrated that inhibition of cholesterol 24-hydroxylase, key enzyme of cholesterol metabolism leads to a complex dysregulation of lipid homeostasis. Our results contribute to dissect the potential role of lipids in severe neurodegenerative diseases like AD.