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International audience; Background: The 5XFAD model of Alzheimer’s disease (AD) bearing five familial mutations of Alzheimer’s disease on human APP and PSEN1 transgenes shows deposits of amyloid-β peptide (Aβ) as early as 2 months, while deficits in long-term memory can be detected at 4 months using the highly sensitive olfactory-dependent tests that we previously reported. Objective: Given that detecting early dysfunctions in AD prior to overt pathology is of major interest in the field, we sought to detect memory deficits at earlier stages of the disease in 3-month-old male 5XFAD mice. Methods: To this end, we used the Helico Maze, a behavioral task that was recently developed and patented. This device allows deeper analysis of learning and subcategories of hippocampal-dependent long-term memory using olfactory cues. Results: Eight male 5XFAD and 6 male wild-type (WT: C57Bl6 background) mice of 3 months of age were tested in the Helico Maze. The results demonstrated, for the first time, a starting deficit of pure reference long-term memory. Interestingly, memory impairment was clearly correlated with Aβ deposits in the hippocampus. While we also found significant differences in astrogliosis between 5XFAD and WT mice, this was not correlated with memory abilities. Conclusion: Our results underline the efficiency of this new olfactory-dependent behavioral task, which is easy to use, with a small cohort of mice. Using the Helico Maze may open new avenues to validate the efficacy of treatments that target early events related to the amyloid-dependent pathway of the disease and AD progression.

International audience; Global Cerebral Ischemia (GCI) occurs following cardiac arrest or neonatal asphyxia and leads to harmful neurological consequences. In most cases, patients who survive cardiac arrest develop severe cognitive and motor impairments. This study focused on learning and memory deficits associated with brain neuroanatomical reorganization that appears after GCI. The four-vessel occlusion (4VO) model was performed to produce a transient GCI. Hippocampal lesions in ischemic rats were visualized using anatomical Magnetic Resonance Imaging (aMRI). Then, the learning and memory abilities of control and ischemic (bilaterally or unilaterally) rats were assessed through the olfactory associated learning task. Finally, a "longitudinal" histological study was carried out to highlight the cellular reorganizations occurring after GCI. We demonstrated that the imaging, behavioral and histological results are closely related. In fact, aMRI revealed the appearance of hyper-intense signals in the dorsal hippocampus at day 3 post-GCI. Consequently, we showed a rise in cell proliferation (Ki 67+cells) and endogenous neurogenesis especially in the dentate gyrus (DG) at day 3 post-GCI. Then, hyper-intense signals in the dorsal hippocampus were confirmed by strong neuronal losses in the CA1 layer at day 7 post-GCI. These results were linked with severe learning and memory impairments only in bilaterally ischemic rats at day 14 post-GCI. This amnesia was accompanied by huge astroglial and microglial hyperactivity at day 30 post-GCI. Finally, Nestin+cells and astrocytes gave rise to astroglial scars, which persisted 60days post-GCI. In the light of these results, the 4VO model appears a reliable method to produce amnesia in order to study and develop new therapeutic strategies.

International audience; Background: Stem cell-based therapies are an attractive option to promote regeneration and repair defective tissues and organs. Thanks to their multipotency, high proliferation rate and the lack of major ethical limitations, "olfactory ecto-mesenchymal stem cells" (OE-MSCs) have been described as a promising candidate to treat a variety of damaged tissues. Easily accessible in the nasal cavity of most mammals, these cells are highly suitable for autologous cell-based therapies and do not face issues associated with other stem cells. However, their clinical use in humans and animals is limited due to a lack of preclinical studies on autologous transplantation and because no well-established methods currently exist to cultivate these cells. Here we evaluated the feasibility of collecting, purifying and amplifying OE-MSCs from different mammalian genera with the goal of promoting their interest in veterinary regenerative medicine. Biopsies of olfactory mucosa from eight mammalian genera (mouse, rat, rabbit, sheep, dog, horse, gray mouse lemur and macaque) were collected, using techniques derived from those previously used in humans and rats. The possibility of amplifying these cells and their stemness features and differentiation capability were then evaluated. Results: Biopsies were successfully performed on olfactory mucosa without requiring the sacrifice of the donor animal, except mice. Cell populations were rapidly generated from olfactory mucosa explants. These cells displayed similar key features of their human counterparts: a fibroblastic morphology, a robust expression of nestin, an ability to form spheres and similar expression of surface markers (CD44, CD73). Moreover, most of them also exhibited high proliferation rates and clonogenicity with genus-specific properties. Finally, OE-MSCs also showed the ability to differentiate into mesodermal lineages. Conclusions: This article describes for the first time how millions of OE-MSCs can be quickly and easily obtained from different mammalian genera through protocols that are well-suited for autologous transplantations. Moreover, their multipotency makes them relevant to evaluate therapeutic application in a wide variety of tissue injury models. This study paves the way for the development of new fundamental and clinical studies based on OE-MSCs transplantation and suggests their interest in veterinary medicine.

International audience; The transplantation of Olfactory Ecto-Mesenchymal Stem Cells (OEMSCs) could be a helpful therapeutic strategy for spinal cord repair. Using an acute rat model of high cervical contusion that provokes a persistent hemi-diaphragmatic and foreleg paralysis, we evaluated the therapeutic effect of a delayed syngeneic transplantation (two days post-contusion) of OEMSCs within the injured spinal cord. Respiratory function was assessed using diaphragmatic electromyography and neuro-electrophysiological recordings of phrenic nerves (innervating the diaphragm). Locomotor function was evaluated using the ladder-walking locomotor test. Cellular reorganization in the injured area was also studied using immunohistochemical and microscopic techniques. We report a substantial improvement in breathing movements, in activities of the ipsilateral phrenic nerve and ipsilateral diaphragm and also in locomotor abilities four months post-transplantation with nasal OEMSCs. Moreover, in the grafted spinal cord, lesioned areas and inflammation were reduced. Some grafted stem cells adopted a neuronal phenotype and axogenesis was observed in the injury site. The therapeutic effect on the supraspinal command is presumably due to both neuronal replacements and beneficial paracrine effects on the injury area. Our study provides evidence that nasal OEMSCs could be a first step in clinical application, particularly in patients with reduced breathing/locomotor movements.

Alzheimer's disease (AD) is the main cause of dementia and a major health issue worldwide. The complexity of the pathology continues to challenge its comprehension and the implementation of effective treatments. In the last decade, a number of possible targets of intervention have been pointed out, among which the stimulation of 5-HT

International audience; The frontal cortex is a brain structure that plays an important role in cognition and is known to be affected in Alzheimer's disease (AD) in humans. Over the past years, transgenic mouse models have been generated to recapitulate the main features of this disease, including cognitive impairments. This study investigates frontal cortex dependent learning abilities in one of the most early-onset transgenic murine model of AD, the 5XFAD mice. We compared frontal performance of 2-, 4-, and 6-month-old 5XFAD mice with their wild-type littermates using a newly developed automated device, the olfactory H-maze, in which mice have to discover three different rules consecutively according to the delayed reaction paradigm. We report early cognitive deficits related to frontal cortex appearing in 4-month-old 5XFAD mice before hippocampal-dependent learning and memory impairment, in relation with neuropathologic processes such as strong gliosis and emerging amyloid plaques. The present results demonstrate that the olfactory H-maze is a very sensitive and simple experimental paradigm that allows assessment of frontal functions in transgenic mice and should be useful to test pre-clinical therapeutic approaches to alter the course of AD.

International audience; The FVB/N mice are well suited to generate transgenic animals. These ă mice are also particularly sensitive to seizures and neurodegeneration ă induced by systemic administration of chemoconvulsants and are very ă useful to model epilepsy. However, previous studies report strong ă cognitive and visual impairments suggesting this background unsuitable ă for behavioral analysis. In this study, we assessed and compared ă learning abilities of FVB/N mice to the well characterized C57BL/6 ă strain using the olfactory tubing maze, a non-visual ă hippocampus-dependent task in which the mice were trained to learn ă odor-reward associations. Exploratory behavior and spontaneous locomotor ă activity were then compared using the open field test. We demonstrated ă that FVB/N mice were able to learn the task, reaching at the end of the ă test a high percentage of correct responses. Interestingly, the ă performance of the FVB/N mice was at least similar to that of the ă C57BL/6 mice. Moreover, in contrast to previous reports, the FVB/N mice ă displayed a spontaneous locomotor activity lower than C57BL/6 mice. Our ă study demonstrated that FVB/N mice are not cognitively impaired and that ă their learning and memory performance can be assessed when the task is ă based on olfaction rather than vision. (C) 2015 Elsevier B.V. All rights ă reserved.

International audience; Stem cells are attractive tools to develop new therapeutic strategies for a variety of disorders. While ethical and technical issues, associated with embryonic, fetal and neural stem cells, limit the translation to clinical applications, the nasal stem cells identified in the human olfactory mucosa stand as a promising candidate for stem cell-based therapies. Located in the back of the nose, this multipotent stem cell type is readily accessible in humans, a feature that makes these cells highly suitable for the development of autologous cell-based therapies. However, preclinical studies based on autologous transplantation of rodent olfactory stem cells are impeded because of the narrow opening of the nasal cavity. In this study, we report the development of a unique method permitting to quickly and safely biopsy olfactory mucosa in rats. Using this newly developed technique, rat stem cells expressing the stem cell marker Nestin were successfully isolated without requiring the sacrifice of the donor animal. As an evidence of the self-renewal capacity of the isolated cells, several millions of rat cells were amplified from a single biopsy within four weeks. Using an olfactory discrimination test, we additionally showed that this novel biopsy method does not affect the sense of smell and the learning and memory abilities of the operated animals. This study describes for the first time a methodology allowing the derivation of rat nasal cells in a way that is suitable for studying the effects of autologous transplantation of any cell type present in the olfactory mucosa in a wide variety of rat models.

International audience; Adult olfactory ectomesenchymal stem cells (OE-MSCs) and olfactory ensheathing cells (OECs), both from the nasal olfactory lamina propria, display robust regenerative properties when transplanted into the nervous system, but the mechanisms supporting such therapeutic effects remain unknown. Matrix metalloproteinases (MMPs) are an important family of proteinases contributing to cell motility and axonal outgrowth across the extracellular matrix (ECM) in physiological and pathological conditions. In this study, we have characterized for the first time in nasal human OE-MSCs the expression profile of some MMPs currently associated with cell migration and invasiveness. We demonstrate different patterns of expression for MMP-1, MMP-2, MMP-9, and MT1-MMP upon cell migration when compared with nonmigrating cells. Our results establish a correspondence between the localization of these proteinases in the migration front with the ability of cells to migrate. Using various modulators of MMP activity, we also show that at least MMP-2, MMP-9, and MT1-MMP contribute to OE-MSC migration in an in vitro 3D test. Furthermore, we demonstrate under the same conditions of culture used for in vivo transplantation that OE-MSCs and OECs secrete neurotrophic factors that promote neurite outgrowth of cortical and dorsal root ganglia (DRG) neurons, as well as axo-dendritic differentiation of cortical neurons. These effects were abolished by the depletion of MMP-2 and MMP-9 from the culture conditioned media. Altogether, our results provide the first evidence that MMPs may contribute to the therapeutic features of OE-MSCs and OECs through the control of their motility and/or their neurotrophic properties. Our data provide new insight into the mechanisms of neuroregeneration and will contribute to optimization of cell therapy strategies.

International audience; The olfactory mucosa, located in the nasal cavity, is in charge of detecting odours. It is also the only nervous tissue that is exposed to the external environment and easily accessible in every living individual. As a result, this tissue is unique for anyone aiming to identify molecular anomalies in the pathological brain or isolate adult stem cells for cell therapy. Molecular abnormalities in brain diseases are often studied using nervous tissue samples collected post-mortem. However, this material has numerous limitations. In contrast, the olfactory mucosa is readily accessible and can be biopsied safely without any loss of sense of smell(1). Accordingly, the olfactory mucosa provides an "open window" in the adult human through which one can study developmental (e.g. autism, schizophrenia)(2-4) or neurodegenerative (e.g. Parkinson, Alzheimer) diseases(4,5). Olfactory mucosa can be used for either comparative molecular studies(4,6) or in vitro experiments on neurogenesis(3,7). The olfactory epithelium is also a nervous tissue that produces new neurons every day to replace those that are damaged by pollution, bacterial of viral infections. This permanent neurogenesis is sustained by progenitors but also stem cells residing within both compartments of the mucosa, namely the neuroepithelium and the underlying lamina propria(8-10). We recently developed a method to purify the adult stem cells located in the lamina propria and, after having demonstrated that they are closely related to bone marrow mesenchymal stem cells (BM-MSC), we named them olfactory ecto-mesenchymal stem cells (OE-MSC)(11). Interestingly, when compared to BM-MSCs, OE-MSCs display a high proliferation rate, an elevated clonogenicity and an inclination to differentiate into neural cells. We took advantage of these characteristics to perform studies dedicated to unveil new candidate genes in schizophrenia and Parkinson's disease(4). We and others have also shown that OE-MSCs are promising candidates for cell therapy, after a spinal cord trauma(12,13), a cochlear damage(14) or in an animal models of Parkinson's disease(15) or amnesia(16). In this study, we present methods to biopsy olfactory mucosa in rats and humans. After collection, the lamina propria is enzymatically separated from the epithelium and stem cells are purified using an enzymatic or a non-enzymatic method. Purified olfactory stem cells can then be either grown in large numbers and banked in liquid nitrogen or induced to form spheres or differentiated into neural cells. These stem cells can also be used for comparative omics (genomic, transcriptomic, epigenomic, proteomic) studies.

International audience; Stem cell-based therapy has been proposed as a potential means of treatment for a variety of brain disorders. Because ethical and technical issues have so far limited the clinical translation of research using embryonic/fetal cells and neural tissue, respectively, the search for alternative sources of therapeutic stem cells remains ongoing. Here, we report that upon transplantation into mice with chemically induced hippocampal lesions, human olfactory ecto-mesenchymal stem cells (OE-MSCs) - adult stem cells from human nasal olfactory lamina propria - migrated toward the sites of neural damage, where they differentiated into neurons. Additionally, transplanted OE-MSCs stimulated endogenous neurogenesis, restored synaptic transmission, and enhanced long-term potentiation. Mice that received transplanted OE-MSCs exhibited restoration of learning and memory on behavioral tests compared with lesioned, nontransplanted control mice. Similar results were obtained when OE-MSCs were injected into the cerebrospinal fluid. These data show that OE-MSCs can induce neurogenesis and contribute to restoration of hippocampal neuronal networks via trophic actions. They provide evidence that human olfactory tissue is a conceivable source of nervous system replacement cells. This stem cell subtype may be useful for a broad range of stem cell-related studies.