Contribution à l'étude de la biosynthèse et des effets pro-remyelinisants de la progesterone dans le système nerveux central

PARIS-BIUP (751062107) / SudocPARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Contribution à l'étude de la biosynthèse et des effets pro-remyelinisants de la progesterone dans le système nerveux central

PARIS-BIUP (751062107) / SudocPARIS-BIUSJ-Thèses (751052125) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Effect of uranium on multipotency of neural stem cells in a primary neurosphere culture model

International audienceUranium exposure situations are diverse and originate from its natural presence in the environment, and from its use in specific professional activities in relation with the nuclear industry (extraction, nuclear fuel cycles, and dismantling operations). Uranium internal contamination can occur via ingestion of contaminated food and drinking water or via inhalation of particulate aerosols containing uranium dust. This latest situation is the main cause of contamination in nuclear occupational activities. These contaminations raise concern in terms of potential consequences on human health. They appear to have negative impact on the brain as experimental studies have shown that uranium exposure via ingestion or inhalation can lead to cognitive impairments in rats. Neurogenesis disruption has been proposed to underlie these effects. To address this question, we used in vitro neurosphere primary cultures from rat embryo’s telencephalon at embryonic day 13. We studied uranium impact on multipotency of neural stem cell within a range of concentrations (10, 50, 100 μM) versus control over 7 days of contamination. Our results show a significant effect on cell survival via a decrease of the absolute number of all cell types: neurons, astrocytes and mature oligodendrocytes at 50 and 100 μM. Among cells surviving after 7 days of contamination, analysis of apoptotic gene expression tend to suggest an adaptive response via Bax/Bcl2 balance in favour of cell survival at 100 μM condition, that will need further investigations. In this condition (100 μM), neurons exhibit an aborted morphology with a reduction of the axon and dendrite length correlated with a significant decrease of gene expression GAP43 known to be involved in dendritic arborization development. Regarding gliogenesis, uranium seems to have a direct action on the maintenance of a population of glial progenitors Olig2 positive, linked with a significant increase of NeuroG3 gene expression at 100 μM. All together, these results suggest that uranium exerts a specific action on late cell maturation phases rather than on early determination stages.Keywords: neurotoxicity, stem cell, multipotency, metals, uraniu

Effect of uranium on multipotency of neural stem cells in a primary neurosphere culture model

International audienceUranium exposure situations are diverse and originate from its natural presence in the environment, and from its use in specific professional activities in relation with the nuclear industry (extraction, nuclear fuel cycles, and dismantling operations). Uranium internal contamination can occur via ingestion of contaminated food and drinking water or via inhalation of particulate aerosols containing uranium dust. This latest situation is the main cause of contamination in nuclear occupational activities. These contaminations raise concern in terms of potential consequences on human health. They appear to have negative impact on the brain as experimental studies have shown that uranium exposure via ingestion or inhalation can lead to cognitive impairments in rats. Neurogenesis disruption has been proposed to underlie these effects. To address this question, we used in vitro neurosphere primary cultures from rat embryo’s telencephalon at embryonic day 13. We studied uranium impact on multipotency of neural stem cell within a range of concentrations (10, 50, 100 μM) versus control over 7 days of contamination. Our results show a significant effect on cell survival via a decrease of the absolute number of all cell types: neurons, astrocytes and mature oligodendrocytes at 50 and 100 μM. Among cells surviving after 7 days of contamination, analysis of apoptotic gene expression tend to suggest an adaptive response via Bax/Bcl2 balance in favour of cell survival at 100 μM condition, that will need further investigations. In this condition (100 μM), neurons exhibit an aborted morphology with a reduction of the axon and dendrite length correlated with a significant decrease of gene expression GAP43 known to be involved in dendritic arborization development. Regarding gliogenesis, uranium seems to have a direct action on the maintenance of a population of glial progenitors Olig2 positive, linked with a significant increase of NeuroG3 gene expression at 100 μM. All together, these results suggest that uranium exerts a specific action on late cell maturation phases rather than on early determination stages.Keywords: neurotoxicity, stem cell, multipotency, metals, uraniu

Design of an Inhalation Chamber and Metrology Assessment to Study Tungsten Aerosol Neurotoxic Effects

International audienceTo evaluate the neurotoxic effects from exposure to airborne tungsten, we developed a method of generating mass concentrations of this element between 5 and 10 mg m-3, the time weighted average occupational exposure limits. We then conducted measurements of the aerosol—a challenge due to the high particle density—that enabled us to calculate the deposition in the upper airway and lungs.First, we fed a mixture of coarse tungsten bead powder and aerosolizable tungsten powder, which had been combined in specific mass proportions, to an RBG 1000 (Palas®) equipped with a cyclone at the outlet that filtered out the coarse particles. Then, we simultaneously measured the resultant aerosol, which was generated in an inhalation chamber, using three pairs of instruments—a Dekati® Low Pressure Impactor (DLPI; 30 L min-1) and a gravimetric filter holder, a DLPI and a TSI® Aerodynamic Particle Sizer (APS; Model 3321), a TSI Engine Exhaust Particle Sizer (EEPS; Model 3090) and an APS—and symmetrical sampling lines. The mass concentrations obtained with the DLPI and the filter holder were extremely consistent with each other, and the mass median aerodynamic diameters based on the DLPI and the APS data (with the Stokes correction applied to the latter) were also fairly close (1.77 and 1.89 ^m, respectively). Additionally, the count median diameter determined from the electrical mobility measured by the EEPS equaled 0.17 ^m, which falls beyond both the intended range of the instrument and the range of previously studied aerodynamic sizes.Overall, the results from the DLPI, the APS, and the EEPS showed very good agreement. Computational fluid dynamics (CFD) simulations of the airflows and aerosol dispersion in the inhalation chamber verified that the test aerosol was homogeneous and representative

Brain accumulation of inhaled uranium in the rat depends on aerosol concentration, exposure repetitions, particle size and solubility

International audienceA rostro-caudal gradient of uranium (U) in the brain has been suggested after its inhalation. To study thefactors influencing this mapping, we first used 30-min acute inhalation at 56 mg/m3 of the relatively soluble form UO4 in the rat. These exposure parameterswere then used as a reference in comparison with the other experimental conditions. Other groups received acute inhalation at different concentrations, repeated low dose inhalation of UO4 (10 exposures) or acute low dose inhalation of the insoluble form UO2. At 24 hafter the last exposure, all rats showed a brain U accumulation with a rostro-caudal gradient as compared to controls. However, the total concentration to the brain was greater after repeated exposure than acute exposure, demonstrating an accumulative effect. In comparison with the low dose soluble U exposure, a higher accumulation in the front of the brain was observed after exposure to higher dose, to insoluble particles and following repetition of exposures, thus demonstrating a dose effect and influences of solubility and repetition of exposures. In the last part, exposure to ultrafine U particles made it possible to show 24 h after exposure the presence of U in the brain according to a rostro-caudal gradient. Finally, the time-course after exposure to micronic or nanometric U particles has revealed greater residence times for nanoparticles

Effets de l’inhalation de particules de tungstène sur le cerveau de rat : métrologie et étude pilote

International audienc

Neurotoxicological outcome of a chemical and radiological co-exposure in rat: low dose gamma irradiation and inhalation of tungsten particles

International audienceBackground and PurposeToxicology studies gradually tend to consider the concept of exposome in their experimental design. Each individual is indeed exposed to a unique and wide range of stressors simultaneously, and these stressors might interact. Combining them therefore provides a more realistic representation of an individual’s exposome and the toxicity they experience. Nuclear industry workers integrate specific stressors in their occupational exposome, including external gamma irradiation. It is of particular interest due to uncertainties regarding its dose-response curve in the low dose range, some studies even reporting beneficial neuroprotective effects. Nuclear industry workers are also subject to particulate aerosol inhalation, a major cause of contamination. Our chemical stressor, Tungsten (W), is an emerging contaminant in the environment, used in various industrial or military settings due to its remarkable physical properties. More specifically, nuclear fusion reactors integrate a W shielding which is eroded by the plasma stream during normal operation. The W particles produced pose a risk of exposure of nuclear workers during maintenance, dismantling operations, or loss of vacuum accidents. Although W was initially considered an inert metal, recent national authorities report raise questions about its effective toxicity.Our aim is to explore the differential neurotoxicological effects of a radiological and chemical co-exposure combining W particles inhalation and low dose gamma irradiation and study the mechanisms involved.Methods12-week-old male Sprague-Dawley rats were exposed to a polydisperse aerosol of metal W particles in a nose-only setting (80 mg.m-3, 30 minutes) and/or to a full body low dose gamma irradiation (50 mGy, 50 mGy.min-1). 24 hours (24h) and 28 days (28d) post-exposure, biological samples were collected: whole brains, brain structures, olfactory epithelium, lung, kidney, and plasma. Processes of interest included neuronal integrity, cell survival, oxidative stress, and inflammation in the frontal cortex (FC) and olfactory bulb (OB).ResultsWhile no changes could be observed in OB or FC in apoptosis or proliferation with the markers used, histological studies did unveil statistically significant differences more often between co-exposed and control groups for several parameters, than groups exposed to stressors alone. We observed an increase at 24h and 28d of total cellular density in FC. Microglial density increased at 24h and decreased at 28d in the FC, while activation phenotypes remained unchanged. In OB, both microglial density and activation increased after 28d in the co-exposed group. Analyzing a specific suffering phenotype in FC, we observed a decrease of the density of donut-like neurons at 24h, and an increase after 28d.Gene expression analysis via RT-qPCR revealed a strong heterogeneity of response between FC and OB and the main significant differences were observed for the co-exposed group compared to control. A significant increase of the expression of antioxidant genes in the FC at 28d was observed in parallel with a decrease of these genes in OB. In terms of inflammation, we measured an increased expression of MCP1 in FC at 24h and a decreased expression of TNFɑ in OB at 28d. Conclusions Our goal was to determine whether differential effects could be observed on target processes involved in cerebral toxicity. Co-exposure appears to cause significant effects when compared to control and single stressor groups, hence our discussion and conclusion will focus on the co-exposed groups.Although we see modulations of cell density for microglia and donut-like neurons, our findings do not suggest an increase in cell death or proliferation. The temporal concordance between the microglial depletion in FC and the increase in density in OB at 28d suggests a potential migration of these cells from the FC to the OB in the time window considered. Similarly, the increase in microglial density in FC at 24h is probably caused by a microglial migration from deeper layers of the FC or neighboring microstructures. The changes observed in terms of cytokines gene analysis support the migratory hypothesis as MCP1 expression increases in FC at 24h. The decrease in TNFɑ gene expression in OB at 28d could reflect microglial activation towards anti-inflammatory phenotypes.The antioxidant response appearing in FC at 28d could be the consequence of a disruption in the cerebral microenvironment caused by the microglial depletion. This unbalance in microenvironment could also be increasing the density of donut-like neurons at 28d in FC.Overall, our results suggest heterogeneous responses among structures and an increased effect when combining our stressors. Ongoing experiments are investigating antioxidative mechanisms (transcription factor translocation) and hypoxia contribution. ICP-MS studies will determine the impact of irradiation on the biodistribution of W in our target organs. Additionally, the increase in total cell density observed in FC at both time points cannot be explained entirely by microglial density modulations. Other cell types will be considered in this process (progenitor cells, peripheral cell intrusion). As the modulations are still visible or reversed at 28d, studies should be undertaken beyond this time point to question their persistence

Cell proliferation and cell death are disturbed during prenatal and postnatal brain development after uranium exposure

International audienceThe developing brain is more susceptible to neurotoxic compounds than adult brain. It is also well known that disturbances during brain development cause neurological disorders in adulthood. The brain is known to be a target organ of uranium (U) exposure and previous studies have noted that internal U contamination of adult rats induces behavioral disorders as well as affects neurochemistry and neurophysiological properties. In this study, we investigated whether depleted uranium (DU) exposure affects neurogenesis during prenatal and postnatal brain development. We examined the structural morphology of the brain, cell death and finally cell proliferation in animals exposed to DU during gestation and lactation compared to control animals. Our results showed that DU decreases cell death in the cortical neuroepithelium of gestational day (GD) 13 embryos exposed at 40. mg/L and 120. mg/L and of GD18 fetuses exposed at 120. mg/L without modification of the number of apoptotic cells. Cell proliferation analysis showed an increase of BrdU labeling in the dentate neuroepithelium of fetuses from GD18 at 120. mg/L. Postnatally, cell death is increased in the dentate gyrus of postnatal day (PND) 0 and PND5 exposed pups at 120. mg/L and is associated with an increase of apoptotic cell number only at PND5. Finally, a decrease in dividing cells is observed in the dentate gyrus of PND21 rats developmentally exposed to 120. mg/L DU, but not at PND0 and PND5. These results show that DU exposure during brain development causes opposite effects on cell proliferation and cell death processes between prenatal and postnatal development mainly at the highest dose. Although these modifications do not have a major impact in brain morphology, they could affect the next steps of neurogenesis and thus might disrupt the fine organization of the neuronal network