Effects of Human Alpha-Synuclein A53T-A30P Mutations on SVZ and Local Olfactory Bulb Cell Proliferation in a Transgenic Rat Model of Parkinson Disease

A transgenic Sprague Dawley rat bearing the A30P and A53T α-synuclein (α-syn) human mutations under the control of the tyrosine hydroxylase promoter was generated in order to get a better understanding of the role of the human α-syn mutations on the neuropathological events involved in the progression of the Parkinson's disease (PD). This rat displayed olfactory deficits in the absence of motor impairments as observed in most early PD cases. In order to investigate the role of the mutated α-syn on cell proliferation, we focused on the subventricular zone (SVZ) and the olfactory bulbs (OB) as a change of the proliferation could affect OB function. The effect on OB dopaminergic innervation was investigated. The human α-syn co-localized in TH-positive OB neurons. No human α-syn was visualized in the SVZ. A significant increase in resident cell proliferation in the glomerular but not in the granular layers of the OB and in the SVZ was observed. TH innervation was significantly increased within the glomerular layer without an increase in the size of the glomeruli. Our rat could be a good model to investigate the role of human mutated α-syn on the development of olfactory deficits

Pathological lesions in colonic biopsies during Parkinson's disease.

International audiencePas de résum

Low-dose pesticide mixture induces senescence in normal mesenchymal stem cells (MSC) and promotes tumorigenic phenotype in premalignant MSC

Humans are chronically exposed to multiple environmental pollutants such as pesticides with no significant evidence about the safety of such poly-exposures. We exposed mesenchymal stem cells (MSC) to very low doses of mixture of seven pesticides frequently detected in food samples for 21 days in vitro. We observed a permanent phenotype modification with a specific induction of an oxidative stress-related senescence. Pesticide mixture also induced a shift in MSC differentiation towards adipogenesis but did not initiate a tumorigenic transformation. In modified MSC in which a premalignant phenotype wasinduced, the exposure to pesticide mixture promoted tumorigenic phenotype both in vitro andin vivo after cell implantation, in all nude mice. Our results suggest that a commoncombination of pesticides can induce a premature ageing of adult MSC, and as such couldaccelerate age-related diseases. Exposure to pesticide mixture may also promote thetumorigenic transformation in a predisposed stromal environment

Distinct Roles of Bcl-2 and Bcl-Xl in the Apoptosis of Human Bone Marrow Mesenchymal Stem Cells during Differentiation

Background: Adult mesenchymal stem cells (MSCs) can be maintained over extended periods of time before activation and differentiation. Little is known about the programs that sustain the survival of these cells. Principal Findings: Undifferentiated adult human MSCs (hMSCs) did not undergo apoptosis in response to different cell death inducers. Conversely, the same inducers can readily induce apoptosis when hMSCs are engaged in the early stages of differentiation. The survival of undifferentiated cells is linked to the expression of Bcl-Xl and Bcl-2 in completely opposite ways. Bcl-Xl is expressed at similar levels in undifferentiated and differentiated hMSCs while Bcl-2 is expressed only in differentiated cells. In undifferentiated hMSCs, the down-regulation of Bcl-Xl is associated with an increased sensitivity to apoptosis while the ectopic expression of Bcl-2 induced apoptosis. This apoptosis is linked to the presence of cytoplasmic Nur 77 in undifferentiated hMSCs. Significance: In hMSCs, the expression of Bcl-2 depends on cellular differentiation and can be either pro- or anti-apoptotic. Bcl-Xl, on the other hand, exhibits an anti-apoptotic activity under all conditions

Les réactions immunes dans les xénogreffes intracérébrales (mécanisme et prévention)

La transplantation de neurones fœtaux d origine humaine est une des stratégies thérapeutiques envisagées pour compenser les pertes cellulaires survenant lors de maladies neurodégénératives. Les problèmes éthiques et l accès limité à ces neurones limitent le développement de cette approche et font apparaître la transplantation de neuroblastes porcins (NBp) comme une alternative intéressante, même si les problèmes de rejet restent à maîtriser. Pour comprendre et tenter de contrôler ce phénomène, nous avons utilisé comme modèle la greffe de NBp dans le striatum de rat. Dans ce modèle, le rejet des NBp intervient 5 à 7 semaines post-greffe et est accompagné d une infiltration massive du greffon par des cellules microgliales/macrophages et des lymphocytes T (LT). La survie prolongée des NBp observée après un traitement par la minocycline suggère un rôle important du processus inflammatoire dans le rejet. Cet antibiotique pourrait notamment intervenir en limitant l activation de certaines cellules présentatrices d antigènes (CPA) dans les jours qui suivent la greffe. Outre les cellules microgliales activées, nos analyses ont révélé la présence d autres CPA au sein de la greffe. Ces cellules appelées cellules dendritiques (CD) sont présentes dès 3 jours après l opération. Les CD étant les seules cellules capables d initier une réponse immune spécifique via l activation de LT naïfs, elles pourraient jouer un rôle majeur dans l initiation du processus de rejet. De plus, la co-localisation des CD et des LT au sein de la greffe à J42 ouvre la possibilité d une restimulation locale des LT par les CD pour induire un rejet rapide des cellules greffées. Un des derniers points étudiés concerne l intérêt des précurseurs neuraux porcins (PNp) comme source cellulaire. La survie prolongée des PNp dans un striatum de rat comparativement à des NBp suggère que la greffe de ces précurseurs permettrait de minimiser la réaction de rejet lors d une xénogreffe dans le système nerveux central en plus de traitements immunosuppresseurs locaux ou systémiques.Transplantation of human fetal neurons is an interesting restorative approach for central nervous system (CNS) disorders such as Parkinson s disease. However, the limited accessibility to human embryonic neurons and the ethical problems regarding their use impair the development of this approach. In this context, porcine neuroblasts (pNB) appear as an interesting source of transplantable cells, even if the rejection problems remain to be solved. To understand and control the rejection mechanisms occurring in the CNS, we used as a model, the transplantation of pNB into the rat striatum. In this model, the pNB are rejected within 5 to 7 weeks and the phenomenon is accompanied by T cell infiltration and microglial cell activation. The long survival of pNB in case of treatment of the rats with minocycline suggests an important role of the inflammatory reaction in the rejection processes. This antibiotic may intervene by reducing the activation of antigen presenting cells (APC) just after cell grafting. Besides the activated microglial cells, our analyses reveal the presence of other APC inside the graft. These cells called dendritic cells (DC) are present as soon as 3 days post-grafting. Since they are the only cells capable of initiating a specific immune response by priming naïve T cells in deep cervical ganglia, they may have a major role in the initiation of rejection processes. In addition, the co-localisation of DC and T cells in the graft at day 42 raises the possibility of a local restimulation of T cells by DC, inducing a rapid rejection of xenogeneic cells. In the last point our studied concerns the advantage of using porcine neural precursor cells (pNPC) as a source of transplantable cells. The longer survival of pNPC in the rat striatum as compared to pNB suggests that the transplantation of these precursors might be one way to reduce the rejection process in case of intracerebral xenografting in addition to local or systemic immunosuppressive treatments.NANTES-BU Médecine pharmacie (441092101) / SudocSudocFranceF

Application de la cytométrie de flux aux cellules du système nerveux central et recherche de nouveaux marqueurs pour les cellules souches neurales

Les cellules souches neurales (CSN), cellules multipotentes ayant conservé une forte capacité proliférative, ouvrent de nouvelles perspectives dans le traitement des maladies neurodégénératives. Toutefois, leur meilleure caractérisation et la maîtrise de leur différenciation restent des problèmes fondamentaux. Cette thèse a eu pour objectif la recherche de marqueurs extracellulaires spécifiques des CSN permettant leur purification. Le premier point a été d'adapter l'analyse phénotypique par cytométrie de flux aux cellules du système nerveux central (SNC) puis de contrôler la fiabilité de marqueurs comme la nestine utilisée pour définir les CSN. Nous avons ensuite analysé l'expression de différents marqueurs type CD (cluster of differenciation) par les cellules présentes dans des cultures de CSN, d'astrocytes et de neurones. Nous avons montré que quelques marqueurs étaient majoritairement exprimés par ces cellules et qu'une sélection négative par CD3 permettrait un enrichissement en CSN.Neural stem cells (NSC) are of great interest for restorative strategies in neurodegenerative diseases because of their capacity to proliferate and to give rise to all of the major cell types of the central nervous system (CNS). However, major improvements in the characterisation and control of the differentiation of NSC are necessary. Our objectives were to search for specific extracellular markers of NSC in order to be able to identify and to sort them from amongst other cell types. First we investigated the possibility of using flow cytometry to characterise CNS-derived cells and then to control the specificity of markers such as nestin used to define NSC. Next, we evaluated the expression of a number of CD molecules (cluster of differentiation) by cells present in cultures of NSC, astrocytes and neurons. The results showed that some markers are differentially expressed by these cells and that a negative selection by CD3 would enable enrichment of NSC.NANTES-BU Médecine pharmacie (441092101) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

A theory that may explain the Hayflick limit — a means to delete one copy of a repeating sequence during each cell cycle in certain human cells such as fibroblasts

International audienceA model that may explain the limited division potential of certain cells such as human fibroblasts in culture is presented. The central postulate of this theory is that there exists, prior to certain key exons that code for materials needed for cell division, a unique sequence of specific repeating segments of DNA. One copy of such repeating segments is deleted during each cell cycle in cells that are not protected from such deletion through methylation of their cytosine residues. According to this theory, the means through which such repeated sequences are removed, one per cycle, is through the sequential action of enzymes that act much as bacterial restriction enzymes do--namely to produce scissions in both strands of DNA in areas that correspond to the DNA base sequence recognition specificities of such enzymes. After the first scission early in a replicative cycle, that enzyme becomes inhibited, but the cleavage of the first site exposes the closest site in the repetitive element to the action of a second restriction enzyme after which that enzyme also becomes inhibited. Then repair occurs, regenerating the original first site. Through this sequential activation and inhibition of two different restriction enzymes, only one copy of the repeating sequence is deleted during each cell cycle. In effect, the repeating sequence operates as a precise counter of the numbers of cell doubling that have occurred since the cells involved differentiated during development

Enteric glia at center stage of inflammatory bowel disease

International audienceAlthough our understanding of the pathophysiology of inflammatory bowel disease (IBD) is increasing, the expanding body of knowledge does not simplify the equation but rather reveals diverse, interconnected, and complex mechanisms in IBD. In addition to immune overactivation, defects in intestinal epithelial barrier (IEB) functioning, dysbiosis, and structural and functional abnormalities of the enteric nervous system are emerging as new elements contributing to the development of IBD. In addition to molecular changes in IBD, enteric glia from patients with Crohn's disease (CD) exhibits the inability to strengthen the IEB; these defects are not observed in patients with ulcerative colitis. In addition, there is a growing body of work describing that enteric glia interacts with not only enterocytes and enteric neurons but also other local cellular neighbours. Thus, because of their functions as connectors and regulators of immune cells, IEB, and microbiota, enteric glia could be the keystone of digestive homeostasis that is lacking in patients with CD

Cancer stem cells: Beyond Koch’s postulates

International audienceUntil the last century, infectious diseases were the leading cause of human mortality. Therefore, our current medical reasoning is profoundly influenced by views that originated from medical microbiology. The notion that cancer growth is sustained by a sub-population of particular cells, the cancer stem cells, is highly reminiscent of the germ theory of disease as exemplified by Koch's postulates in the XIX th century. However, accumulating data underscore the importance of cell-cell interactions and tumor environment. Hence it is essential to critically review the basic tenets of the cancer stem cell concept on the light of their relationships with Koch's postulates. Shifting the pathogenic element from a special cellular entity (cancer stem cell or microorganism) to a " pathogenic field " could be critical for curing both cancer and drug-resistant infectious diseases

Enteric glial cells have specific immunosuppressive properties

International audienceEnteric glial cells (EGC) have trophic and neuroregulatory functions in the enteric nervous system, but whether they exert a direct effect on immune cells is unknown. Here, we used co-cultures to show that human EGC can inhibit the proliferation of activated T lymphocytes. Interestingly, EGC from Crohn's patients were effective at one EGC for two T cells whereas EGC from control patients required a ratio of 1:1. These data suggest that EGC contribute to local immune homeostasis in the gastrointestinal wall. They also raise the possibility that EGC have particular immunosuppressive properties in inflammatory bowel diseases such as Crohn's disease