Biocompatibilité des complexes protéines–nanoparticules : perspectives sur la réponse cellulaire aux nanoparticules d’oxyde de fer fonctionnalisées, revêtues d’un corona

RÉSUMÉ Cette thèse rapporte l'étude de la biocompatibilité des nanoparticules (NPs) d‟oxyde de fer (Fe3O4) superparamagnetiques candidates à une livraison ciblée des molécules thérapeutiques. Nous nous sommes tout spécialement attachés à étudier l'impact de la composition de surface de ces NPs et l'adsorption des protéines à la surface de ces dernières sur les réponses cellulaires. Pour ce faire, nous avons tout d'abord examiné le potentiel toxique la magnétite avec divers fonctionnalisations : celle qui est préparée avec (1) une monocouche d'acide oléique (Fe3O4@OA), qui est ensuite converti en (2) une enveloppe de silane contenant une amine (Fe3O4@NH2), (3) un revêtement de silice (Fe3O4@SiO2), et (4) une enveloppe de silane contenant une amine sur un revêtement de silice (Fe3O4@SiO2@NH2). La présence de ces groupements à la surface des NPs a été confirmée par l'analyse XPS et la microscopie électronique à transmission (TEM). Nous avons pu prouver que le potentiel toxique des NPs est dose-dépendant et ainsi déterminer les doses biocompatibles pour chaque fonctionnalisation de surface. L'observation au microscope de la morphologie des cellules exposées aux NPs, leur activité mitochondriale et proinflammatoire ont montré que, en plus des caractéristiques de surface, le milieu de culture cellulaire influence également la cytotoxicité des NPs. Ces résultats montrent clairement que pour pouvoir utiliser nos NPs comme nanovecteur pharmaceutique de façon sécuritaire, nous devons contrôler la fonctionnalisation de surface et l'interaction dynamique entre la NP et le milieu physiologique dans lequel elle est suspendue. Pour comprendre l'interaction entre la NP et le milieu de culture, dans une première étape, nous avons utilisé trois milieux de culture différents à savoir : DMEM, F-12K et DMEM/F12 (voir annexe A) et la magnétite sans revêtement (Fe3O4). Ces milieux ont été enrichis soit avec le sérum bovin (voir annexe B) soit avec un sérum synthétique (SFMS). Nous avons prouvé la présence d'une couronne protéique (corona) sur les NPs suspendues dans les milieux de culture enrichis de sérum bovin. Nous avons également démontré que la formation du corona sur les nanoparticules d'oxyde de fer superparamagnétiques (SPIONs) dépend de la composition du milieu de culture et que le potentiel cytotoxique des NPs est influencé par l'interaction NP-protéines.----------ABSTRACT This thesis presents the study of the biocompatibility of nanoparticles (NPs) of iron oxide (Fe3O4) candidates for targeted delivery of therapeutic molecules. We especially devoted to study the impact of the surface composition of the NPs and protein adsorption at the surface thereof on the cellular responses. To do this, we first examined the toxic potential of magnetite with various functionalizations: one that is prepared with (1) a monolayer of oleic acid (Fe3O4@OA), which is then converted to (2) an envelope silane containing an amine (Fe3O4@NH2), (3) a coating of silica (Fe3O4@SiO2), and (4) an envelope containing a silane coating on amine silica (Fe3O4@SiO2@NH2). The presence of these groups at the surface of the NPs was confirmed by XPS and transmission electron microscopy (TEM) analysis. We were able to prove that the toxic potential of NPs is dose-dependent and we determine the biocompatible doses for each surface functionalization. Microscopic observation of the morphology of the cells exposed to NPs, and their proinflammatory and mitochondrial activity showed that, in addition to surface features, the cell culture medium also affect the cytotoxicity of the NPs. These results clearly show that in order to use our NPs as pharmaceutical nanocarrier safely, we need to control the surface functionalization and the dynamic interaction between the NP and the physiological environment in which it is suspended. To understand the interaction between the NP and the culture medium, as a first step, we used three different culture media namely: DMEM, F-12K and DMEM / F12 (see Appendix A) and uncoated magnetite (Fe3O4). These media were enriched with either fetal bovine serum (see Appendix B) or with a synthetic serum (SFMS). We have proved the presence of a protein corona on NPs suspended in culture media enriched with bovine serum. We also demonstrated that the formation of the corona depends on the composition of the culture medium and that the cytotoxic potential of the NPs is influenced by NP-protein interaction. In a second step, we used one culture medium (DMEM / F12) and the magnetite with three different surface compositions: uncoated SPIONs with hydroxyl groups (OH) on the surface; coated SPIONs with an amine group (NH2) on the surface and the last one with a carboxylic group (COOH) on the surface. The results show that the composition of the corona dependst o

The effect of micro-pulsatile electrical and ultrasound stimulation on cellular biosynthetic activities such as cellular proliferation, endogenous nitrogen oxide and collagen synthesis

ABSTRACT: The skin barrier poses an ongoing challenge for the cosmetics industry. Its penetration, by non-invasive means, can readily be achieved with currents and ultrasound or radiofrequency devices through electroporation, sonophoresis, iontophoresis or cavitation. When several types of energy are applied simultaneously, we expect the effects to be magnified and all the more effective. Although the mechanism of action of each technology on the skin is not entirely controlled, and is even less so when multiple technologies are applied concurrently, some studies demonstrate that nitric oxide (NO) plays a pivotal role in skin wound-healing and regeneration. With regard to wound healing, one of the key functions of NO appears to be its permissive effect on keratinocyte and fibroblast proliferation, which helps promote wound re-epithelialization. The objective of the actual research is to gain an in-depth understanding of the mechanisms generated by NO through the application of a specific combination of technologies

Effect of ceria on the organization and bio-ability of anatase fullerene-like crystals

The nanostructure and the oxygen storage capacity against reactive oxygen species (ROS) are essential features to take into account during the design of a new material that will be used as the basis of novel therapeutic technologies. Here we evaluate the incorporation of nano-ceria, which has a demonstrated ability to scavenge free radicals under physiological conditions comparable to those observed for superoxide dismutase and catalase, to TiO2 crystalline assemblies. The material was planned to merge the scavenging properties of CeO2 on a specifically designed structured TiO2 substrate. The presence of Ce atoms has a clear influence in the materials' morphologies, distorting anatase crystal lattice and inducing the formation of fullerene-like structures. The cytotoxicity of the materials against L929 fibroblasts after 24 hours of cell culture was evaluated. Both structural and oxidative properties of the materials have a clear effect on fibroblast viability; in fact it was demonstrated that cellular proliferation can be modulated varying the Ce3+/Ti4+ molar ratio.Fil: Gravina, Noel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Bahía Blanca. Instituto de Química del Sur; Argentina. Universidad Nacional del Sur; ArgentinaFil: Ruso, Juan Manuel. Universidad de Santiago de Compostela; EspañaFil: Mbeh, Doris Antoinette. École Polytechnique De Montréal; CanadáFil: Yahia, L'Hocine. École Polytechnique De Montréal; CanadáFil: Merhi Yahye. University Of Montreal; CanadáFil: Sartuqui, Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Bahía Blanca. Instituto de Química del Sur; Argentina. Universidad Nacional del Sur; ArgentinaFil: Messina, Paula Veronica. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Bahía Blanca. Instituto de Química del Sur; Argentina. Universidad Nacional del Sur; Argentin

Protective role against hydrogen peroxide and fibroblast stimulation via Ce-doped TiO2 nanostructured materials

Background: Cerium oxide (CeO2) and Ce-doped nanostructured materials (NMs) are being seen as innovative therapeutic tools due to their exceptional antioxidant effects; nevertheless their bio-applications are still in their infancy. Methods: TiO2, Ce–TiO2 and CeO2–TiO2 NMs were synthesized by a bottom-up microemulsion-mediated strategy and calcined during 7 h at 650 °C under air flux. The samples were compared to elucidate the physicochemical characteristics that determine cellular uptake, toxicity and the influence of redox balance between the Ce3 +/Ce4 + on the cytoprotective role against an exogenous ROS source: H2O2. Fibroblasts were selected as a cell model because of their participation in wound healing and fibrotic diseases. Results: Ce–TiO2 NM obtained via sol–gel reaction chemistry of metallic organic precursors exerts a real cytoprotective effect against H2O2 over fibroblast proliferation, while CeO2 pre-formed nanoparticles incorporated to TiO2 crystalline matrix lead to a harmful CeO2–TiO2 material. TiO2 was processed by the same pathways as Ce–TiO2 and CeO2–TiO2 NM but did not elicit any adverse or protective influence compared to controls. Conclusions: It was found that the Ce atoms source and its concentration have a clear effect on material's physicochemical properties and its subsequent influence in the cellular response. It can induce a range of biological reactions that vary from cytotoxic to cytoprotective. General significance: Even though there are still some unresolved issues and challenges, the unique physical and chemical properties of Ce-based NMs are fascinating and versatile resources for different biomedical applications.Fil: Gravina, Noel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Química del Sur. Universidad Nacional del Sur. Departamento de Química. Instituto de Química del Sur; ArgentinaFil: Maghni, Karim. Hôpital du Sacré-Coeur-de-Montréal; CanadáFil: Welman, Mélanie. Hôpital du Sacré-Coeur-de-Montréal; CanadáFil: Yahia, L'Hocine. École Polytechnique de Montréal; CanadáFil: Mbeh, Doris Antoinette. École Polytechnique de Montréal; CanadáFil: Messina, Paula Verónica. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Química del Sur. Universidad Nacional del Sur. Departamento de Química. Instituto de Química del Sur; Argentin