The formation of neurospheres was important in demonstrating that neurogenesis in the adult brain may be fuelled by a stem cell population. The olfactory mucosa is another site of neurogenesis which, in humans, has been observed to contain a stem cell population through the formation of neurospheres (Murrell et al., 2005). Stem cells can be defined as cells capable of self-renewal and multipotency. The aim of this study was to investigate the potential of rat olfactory stem cells growing as neurospheres. The hypothesis is that the rat olfactory mucosa contains a 'true' stem cell population that can be cultured as neurospheres and that will demonstrate multipotency by differentiating into 'non-olfactory' cell types and possess the capacity for self-renewal, if provided with the appropriate environmental niche. Here it was found that adult rat olfactory mucosa is capable of generating neurospheres when cultured in EGF and bFGF. Evidence of self-renewal was provided by the formation of six generations of neurospheres, the formation of neurospheres from single cells and the expression of markers associated with self-renewal by neurosphere cells. The multipotency of olfactory neurosphere cells was demonstrated through manipulation of the stem cell niche. In defined culture conditions, extracellular matrix molecules and growth factors were able to induce the differentiation of neurosphere cells down the dopaminergic lineage pathway. When co-cultured with differentiating cells, neonatal myoblasts and 3T3-L1 cells, olfactory neurosphere cells were able to differentiate and incorporate into a skeletal muscle myotube and differentiate into adipocytes, respectively. In conclusion it was found that the adult rat olfactory mucosa is capable of generating neurospheres. When presented with an appropriate niche neurosphere cells are able to self-renew and demonstrate multipotency.

Losing the sense of smell because of aging compromises health and quality of life. In the mouse olfactory epithelium, aging reduces the capacity for tissue homeostasis and regeneration. The microvillous cell subtype that expresses both inositol trisphosphate receptor type 3 (IP3R3) and the neuroproliferative factor neuropeptide Y (NPY) is critical for regulation of homeostasis, yet its role in aging is undefined. We hypothesized that an age-related decline in IP3R3 expression and NPY signaling underlie age-related homeostatic changes and olfactory dysfunction. We found a decrease in IP3R3+ and NPY+ microvillous cell numbers and NPY protein and a reduced sensitivity to NPY-mediated proliferation over 24months. However, in IP3R3-deficient mice, there was no further age-related reduction in cell numbers, proliferation, or olfactory function compared with wild type. The proliferative response was impaired in aged IP3R3-deficient mice when injury was caused by satratoxin G, which induces IP3R3-mediated NPY release, but not by bulbectomy, which does not evoke NPY release. These data identify IP3R3 and NPY signaling as targets for improving recovery following olfactotoxicant exposure.

Disorders of the autonomic nervous system constitute serious complications of spinal cord injury (SCI) and their treatment is usually highly prioritised by spinal patients. Among these, autonomic dysreflexia and impaired thermoregulation are potentially life threatening conditions and require effective management. Olfactory ensheathing cells (OECs), progenitor cells and polymeric scaffolds have been tested in animal models of SCI and some of them have been considered for clinical trials. However, evaluation of the effect of such interventions on autonomic functions has received only rudimentary attention and would require a more thorough experimental assessment before the methods are utilised in human patients. This thesis tested two potential therapeutic strategies for autonomic dysreflexia and examined disorders of thermoregulatory functions in a rat model of spinal cord transection. Magnitude and duration of autonomic dysreflexia were evaluated with radio telemetry in spinalised animals treated with (i) implants of OECs and olfactory neurosphere-derived cells seeded in poly(lactic co glycolic) porous scaffolds or with (ii) transplants of OECs alone. (iii) Effects of SCI and of OECs on the morphology of sympathetic preganglionic neurons (SPNs; which are involved in pathogenesis of autonomic dysreflexia) stained for NADPH diaphorase were examined. (iv) Doppler ultrasonography and infrared thermography were used to assess responses of tail blood flow and surface temperature to cold. Transplants of OECs alone, but not in combination with olfactory neurosphere-derived cells and polymeric scaffolds, resulted in significantly shortened episodes of autonomic dysreflexia. This may be attributed to the alterations to the morphology of SPNs adjacent to the lesion: a transient increase in the morphometric features of the SPNs was evoked by spinal cord transection and this was further altered by transplantation of OECs. The thesis also showed that local responses of tail blood flow and temperature to cold were not abolished by complete SCI suggesting that temperature homeostasis could still be maintained in response to cold. It is hypothesised that OECs facilitate improved recovery from autonomic dysreflexia through alteration of the morphology of SPNs. Furthermore, it is suggested that the role of the tail in heat conservation can be regulated by mechanisms that are independent of the descendent neural control from supraspinal centres.

Disorders of the autonomic nervous system constitute serious complications of spinal cord injury (SCI) and their treatment is usually highly prioritised by spinal patients. Among these, autonomic dysreflexia and impaired thermoregulation are potentially life threatening conditions and require effective management. Olfactory ensheathing cells (OECs), progenitor cells and polymeric scaffolds have been tested in animal models of SCI and some of them have been considered for clinical trials. However, evaluation of the effect of such interventions on autonomic functions has received only rudimentary attention and would require a more thorough experimental assessment before the methods are utilised in human patients. This thesis tested two potential therapeutic strategies for autonomic dysreflexia and examined disorders of thermoregulatory functions in a rat model of spinal cord transection. Magnitude and duration of autonomic dysreflexia were evaluated with radio telemetry in spinalised animals treated with (i) implants of OECs and olfactory neurosphere-derived cells seeded in poly(lactic co glycolic) porous scaffolds or with (ii) transplants of OECs alone. (iii) Effects of SCI and of OECs on the morphology of sympathetic preganglionic neurons (SPNs; which are involved in pathogenesis of autonomic dysreflexia) stained for NADPH diaphorase were examined. (iv) Doppler ultrasonography and infrared thermography were used to assess responses of tail blood flow and surface temperature to cold. Transplants of OECs alone, but not in combination with olfactory neurosphere-derived cells and polymeric scaffolds, resulted in significantly shortened episodes of autonomic dysreflexia. This may be attributed to the alterations to the morphology of SPNs adjacent to the lesion: a transient increase in the morphometric features of the SPNs was evoked by spinal cord transection and this was further altered by transplantation of OECs. The thesis also showed that local responses of tail blood flow and temperature to cold were not abolished by complete SCI suggesting that temperature homeostasis could still be maintained in response to cold. It is hypothesised that OECs facilitate improved recovery from autonomic dysreflexia through alteration of the morphology of SPNs. Furthermore, it is suggested that the role of the tail in heat conservation can be regulated by mechanisms that are independent of the descendent neural control from supraspinal centres.

Les faibles capacités régénératives intrinsèques du système nerveux central, après la survenue de lésions traumatiques ou l'apparition de maladies neuro-dégénératives, ont orienté les recherches vers des thérapies basées sur l'utilisation de cellules souches dans le but de régénérer le tissu cérébral. Cependant, des limitations éthiques et techniques associées aux cellules souches embryonnaires, fœtales ou neurales chez l'adulte restreignent leur utilisation en clinique humaine. À la recherche d'une source alternative, nous nous sommes intéressés à des cellules souches adultes peu connues, provenant du chorion d'un tissu nerveux périphérique en perpétuel renouvellement : les cellules souches de la lamina propria olfactive, localisée dans la cavité nasale. Ces cellules multipotentes ont été décrites comme un sous-type de cellules souches mésenchymateuses, présentant de fortes capacités prolifératives et neurogéniques. En utilisant un premier modèle murin d'amnésie induit par lésion excito-toxique des hippocampes, nous avons montré que des cellules souches olfactives humaines, greffées dans les zones lésées ou dans le liquide céphalo-rachidien, i) s'installent et adoptent un phénotype neuronal, ii) rétablissent la circulation d'informations au sein des réseaux neuronaux défectueux et iii) permettent une récupération des capacités d'apprentissage et de mémorisation. Suite à ces résultats très encourageants, le premier objectif de cette thèse a été de mieux faire connaître ces cellules souches adultes auprès de la communauté scientifique. / The brain displaying poor regenerative capacities, exogenous stem cell-based therapy has been proposed as an attractive strategy to regenerate cerebral tissue after acute injuries or neurodegenerative disorders. However, ethical and technical issues, associated with embryonic, fetal or adult neural stem cells, limit their use in human medicine. In search of alternative candidates, we focused our attention on adult stem cells, located in a peripheral nervous tissue: the nasal stem cells sited in the olfactory lamina propria. These multipotent stem cells have been characterized as a member of the mesenchymal stem cell superfamily, displaying strong proliferative and neurogenic properties. Recently, using a mouse model of amnesia induced by excito-toxic lesions of hippocampal neurons, we demonstrated that olfactory stem cells, grafted in lesioned areas or into the cerebrospinal fluid, i) migrate and differentiate into neuron-like cells, ii) contribute to the restoration of local neuronal networks and iii) promote recovery of learning and memory abilities. In line with these promising results, the first aim of the current thesis was to promote the use of these adult stem cells by the scientific community. For this purpose, we published an article and a book chapter in which we demonstrated that they are suitable for autologous cell therapy in humans. Using an audiovisual document, we showed that these cells i) can be safely obtained in humans, under local anesthesia, without any loss of smell and ii) are easily and quickly amplifiable in vitro.