Thèses sur le sujet « Cellules stromales mésenchymateuses – Cultures cellulaires »
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Magne, Brice. « Évaluation d’une stratégie de préconditionnement de cellules stromales mésenchymateuses pour le traitement des grands-brulés ». Thesis, Université Paris-Saclay (ComUE), 2018. http://www.theses.fr/2018SACLS558/document.
Texte intégralSince the 1980’s, little progress has been made in the management of major burns, in spite of several research advances in the field of skin tissue engineering and regenerative medicine. Developed in 1975, Cultured Epithelial Autografts (CEA) are the last-in-date significant breakthrough, allowing patient survival in most critical cases. However, patients still have to cope with debilitating sequelae including hypertrophic scars, skin fragility, immunometabolic dysfunctions, chronic pain and post-traumatic stress disorder. Mesenchymal Stromal Cells (MSC) have raised an increasing interest during the past 50 years due to their trophic and immunomodulatory properties. Recent findings about their high plasticity to external stimuli have fostered the development of new targeted therapies known as “preconditioning strategies”. This PhD work thus aimed to assess a MSC preconditioning strategy for the treatment of major burns using CEA. The present work was divided into three main experimental parts. First, in vitro experiments were developed in order to set up the preconditioning strategy, and revealed the interesting role of both interleukin-1β (IL-1β) and substance P (SP). Then, both effectiveness and mechanism of action of these preconditioning modalities were assessed in vitro and in vivo, either using MSC or their secretory products. It was thus shown that IL-1β could potentiate MSC effectiveness through the promotion of pro-migratory, anti-inflammatory and pro-remodeling activities. This effect was shown to be partly mediated by the TGF-β1 signaling pathway. At last, this preconditioning strategy was evaluated in a third degree burn rat model mimicking the surgical treatment applied to severe burn patients. Despite technical hurdles limiting CEA engraftment, anti-inflammatory and pro-angiogenic properties of MSCs were confirmed in this model. These preliminary results underline the potential of a preconditioned MSC therapy in wound healing. Additional preclinical studies are now required to corroborate the benefit of such a therapy in major burns
Ishac, Nicole. « Comment deux lignées cellulaires stromales mésenchymateuses humaines récapitulent in vitro le microenvironnement hématopoïétique ? : Intérêt en ingénierie ». Thesis, Tours, 2015. http://www.theses.fr/2015TOUR4038/document.
Texte intégralHematopoiesis occurs in a hypoxic microenvironment or niche in which hematopoietic stem cells (HSCs) are in close contact with mesenchymal stromal cells. Cellular interactions as well as microenvironmental factors such as reactive oxygen species are crucial for the maintenance of normal and leukemic HSCs. Developing an in vitro human culture system that closely mimcs marrow physiology is therefore essential to study the niche. Here, we present a model using two human stromal cell lines, HS-27a and HS-5. Previously poorly described in the literature, we have further characterized both of these cell lines. The first objective was to assess the quality of HS-27a and HS-5 niches by investigating their cellular, molecular and functional characteristics. Our results clearly show that HS-27a cells display features of a “quiescent” niche whereas HS-5 cells rather represent a “proliferative” niche. The second objective was to engineer a hematopoietic niche where the oxidative metabolism is optimized for the expression of an antioxidant protein, glutathione peroxidase 3 (GPx3). The originality of this work is the use of a non-viral gene transfer system by using the transposon piggyBac. This strategy was achieved by delivering a DNA plasmid carrying the gene of interest, and an mRNA source of transposase, the enzyme which catalyzes the transgene integration. Functionally, GPx3 was shown to be a key regulator for sustaining hematopoietic homeostasis by maintaining immature progenitor cells. For the first time, an original non-viral gene transfer has been used to create an in vitro hematopoietic niche that recapitulates the complexity of normal and leukemic microenvironment. This niche not only provides a platform to identify regulatory factors controlling medullary cells, but may also help in the development of targeted therapeutic strategies
Maillot, Charlotte. « Quantification and impact of microcarrier collisions during mesenchymal stem cell culture in bioreactors ». Electronic Thesis or Diss., Université de Lorraine, 2022. http://www.theses.fr/2022LORR0314.
Texte intégralTo date, bottlenecks persist concerning deep scientific understanding of how various process parameters will impact the Mesenchymal Stem Cell production. Specifically applied to microcarrier-based expansion processes of WJ-MSC's, very little information is available to characterize the impact of microcarrier concentration on MSC growth and death rates or on critical quality attributes which may have crucial and possibly dangerous clinical impacts. As a result, the following work proposes to rationally describe the impact of particle concentration on MSC growth through a pluri-disciplinary characterization of microcarrier-microcarrier interactions in agitated conditions. In order to do so the biological and physical aspects of this work will be presented. To begin with, a quantitative approach to estimate cell growth and death kinetics caused by microcarrier-microcarrier collisions in both Erlenmeyer Flasks and Spinner Flasks is described. For this, cells were grown at various microcarrier concentrations using two microcarrier types : Cytodex-1 and Synthemax II. Complementary cultures were performed by adding various concentrations of particles with the same size and density as microcarriers in view of providing specific information on how additional particles may impact MSC growth on microcarriers. In addition, elements of MSC characterization were performed for these experiments to understand not only the impact of microcarrier-microcarrier interactions on growth but also on defined elements of cell quality. In parallel, in order to estimate the amount and intensity of microcarrier-microcarrier collisions in a specific tank geometry, experiments were performed using both the attenuation of light by Cytodex-1 microcarriers (to estimate local microcarrier concentration) and the acoustic signal which comes from particles colliding with a hydrophone (to estimate microcarrier-sensor collision frequency and intensity). These experiments provided elements to estimate the amount of particle collisions that MSC's may perceive during specific dynamic and steady phases of cell culture in STR's. Lastly, a bioreactor-based approach to MSC manufacturing will be presented focusing on biological aspects of how particle concentration and agitation impacts MSC growth and quality attributes. For this, various MSC cultures were performed in STRs with varying particle concentrations and agitation strategies. The MSC's produced in these conditions were then characterized to define if certain critical quality attributes could be affected by parameters such as microcarrier concentration and/or agitation
Loncaric, Darija. « Atténuation des oxydations phosphorylantes et induction d'une réponse cellulaire hypoxique : effêt de l'[alpha]-tocophérol-acétate et de miR-210 sur les cellules stromales mesenchymateuses ». Thesis, Bordeaux, 2019. http://www.theses.fr/2019BORD0198.
Texte intégralIn this thesis, we combined approaches of single-cell cultures, flow-cytometry, energetic metabolismanalysis and molecular genetics in order to get insight in the effects of α-Tocopherol-Acetate (α-TOA)on Mesenchymal Stromal Cells (MStroC) and their functional subpopulations (mesenchymal stem and progenitor cells). The other aim was to test a miR-210 molecule with respect to its potential use as hypoxia mimicking molecule. After defining MStroC population heterogeneity and concluding that the first passage population is convenient for further experiments, we demonstrated that α-TOA exhibits a positive effect on the maintenance of high proliferative capacity of mesenchymal stem cells. This effect could be associated with an attenuation of electron transport chain (ETC) activity, which, on the other hand could explain moderate increase in the level of mitochondrial Reactive Oxygen Species (mtROS) we detected. The increase in mtROS level could be associated with a decreased HIF-1 alpha protein degradation in MStroC exposed to α-TOA. Although we did not detect a compensatory increase in glycolysis, the observed phenomena depict part of a complex cellular response to the low O2 that is demonstrated to be related with maintenance of stem cell primitiveness. The exact mechanism remains to be elucidated as well as its translational potential. In addition, we provided new evidences that miR-210 is integral part in MStroC response to low O2. In the study, we showed increased in miR-210 expression in a short-term (up to 24 hours) and after extended (up to 72 hours) MStroC exposed to low O2. Moreover, we demonstrated that this micro- RNA could be regulated by both HIF-1 and HIF-2 transcriptional factors, suggesting it as integral part of MStroC response to low O2. So far, our data suggest that miR-210 is worthy to be considered as good hypoxia mimicking molecule
Martin, Céline. « Étude des procédés d’amplification de cellules souches mésenchymateuses humaines ». Thesis, Université de Lorraine, 2016. http://www.theses.fr/2016LORR0261/document.
Texte intégralProgress in regenerative medicines over the past ten years have led to an important research mobilisation, but obtaining a sufficient amount of human stem cells remains nonetheless problematic, especially for mesenchymal stem cells (MSC). Hence, this work developed an approach coupling biology and process engineering to identify barriers limiting MSC growth. The study of scaled-up amplification methods was performed using microcarriers and a 200~mL minibioreactors platform. In order to maximise MSC growth in a biochemically controlled environment, a serum free medium development was tested as well. Human MSC as model cell type for cellular therapies have thus been demonstrated as extremely sensitive to freeze/thaw cycles, temperature variations, subject to premature aging and needing a complex medium enriched in multiple growth and adherence factors. Following this study, several pitfalls might be avoided during MSC process scale-up by integrating the cells biology into the bioreactors' process engineering parameters (heat transfer, hydrodamic stress, adhesion surface)
Sion, Caroline. « Development of an optimized perfused-continuous process of culture of human umbilical cord mesenchymal stem cells (hMSC) grown on innovative adhesion supports ». Electronic Thesis or Diss., Université de Lorraine, 2020. http://www.theses.fr/2020LORR0113.
Texte intégralMesenchymal stem cells (MSCs) show great interest in cellular therapies. Their various characteristics such as their immuno-modulatory properties, their ability to differentiate, and also the secretion of factors, are numerous and promising for new clinical treatments for diseases where few therapies are proposed or have few efficiencies. The doses to be injected for significant results must be repeated and generally contain high quantities of cells (106 cells kg-1 per patient approx). Large scale production methods must be implemented to meet the demand, and in the least costly way possible. In this PhD work, the main objective was to develop a scalable process adapted to these support-dependent cells. For this end, a first study allowed to understand part of the mechanisms of interaction of cells with their growth supports, the microcarriers. The adhesion time but also the cell migrations between microcarriers were characterized and evaluated. A strategy of fed-batch mode strategy with microcarriers addition at specific times in the culture was also proposed. Following this, the second part of the study of this work was to determine the efficiency on larger scale expansion process (1.5 L), using of innovative microcarriers developed by the partner teams of the ‘ImprovesStem’ European project. Several microcarriers candidates with chemically modified surface proved to be promising for the expansion of Wharton’s jelly stem cells. Finally, in the last part of the thesis, an innovative process based on the removal of empty microcarriers, avoiding the risk of deleterious frictions between highly concentrated microcarriers was proposed. Moreover, an on-line monitoring of viable cell concentration was carried-out in the stirred tank bioreactor. Innovative commercial microcarriers, soluble under the action of enzymes, were used in this last part of the study. An improvement of the expansion factor (by a factor of 1.5) was obtained in this continuous-perfused mode of culture in the stirred bioreactor. In addition, these enzymatically-soluble commercial microcarriers allowed for an excellent detachment yield, essential to consider their use in cell therapy
Le, Pape Fiona. « Evaluation de la contribution d'une hémoglobine marine dans la culture cellulaire et dans la cellularisation de substituts osseux et méniscaux par des cellules souches mésenchymateuses ». Thesis, Brest, 2016. http://www.theses.fr/2016BRES0002/document.
Texte intégralThis work aimed to develop cell culture systems, in 2D and 3D, based on the properties of HEMOXCell®, a marine oxygen carrier. Our approach was articulated in two main parts: the first one dealing with the assessment of the use of HEMOXCell® in the culture of two cellular models, and the second one, exploiting the results obtained for tissue engineering purposes. In this first axis, the dose-response effect of HEMOXCell® in the CHO-S cells and mesenchymal stem cells (MSC) in vitro culture, allowed the identification of optimal working concentrations, which can promote cell viability and proliferation. The CHO-S model has contributed to the establishment of a performance test of the molecule, and encouraged its use for bioproduction stimulation. The tests performed on MSCs were used to validate the harmlessness of the molecule at low doses and the maintenance of "stemness". The idea to associate MSCs with porous scaffolds is a promising approach for tissue engineering applications, but it is confronted to the lack of oxygen in the depth of the substitutes. In the second part of this project, we worked at improving the cellularization of bone and meniscal substitutes, under static and dynamic culture systems, w/ and w/o HEMOXCell®. In parallel, a study was conducted to attempt to characterize the meniscal cells. Analyses of cellularized biomaterials suggest a beneficial effect of HEMOXCell® when used as a differentiation media supplement. This work contributed to improve this oxygen carrier understanding and to extend the field of its potential uses particularly for therapeutic applications
Albert, Philippe. « Contribution de la culture de cellules à l'étude du contrôle de la prolifération cellulaire au cours de la régénération du membre d'Axolotl (Amphibien Urodèle) ». Lille 1, 1987. http://www.theses.fr/1987LIL10038.
Texte intégralRoger, Mathilde. « Cellules stromales mésenchymateuses comme vecteurs cellulaires de nanoparticules : un nouvel outil thérapeutique des tumeurs cérébrales ». Phd thesis, Université d'Angers, 2010. http://tel.archives-ouvertes.fr/tel-00577731.
Texte intégralMazurier, Christelle. « Perspectives de thérapies cellulaires dans le myélome multiple ». Versailles-St Quentin en Yvelines, 2013. http://www.theses.fr/2013VERS0054.
Texte intégralMultiple Myeloma (MM) is a frequent haematological disorder, often incurable, characterized by a clonal plasma cell proliferation, anemia, kidney insufficiency and hypercalcemy. Current treatments are based on the administration of chemotherapies followed by autologous or allogenic hematopoietic stem cells transplantation. My work is a translational research work which finally aims at developing new cell therapy protocols. First, I have studied the feasability of developing autologous anti-tumoral vaccination to eradicate the plasma cell development. Secondly I have analyzed the putative role of Mesenchymal stem cells (MSC) in osteolysis. Finally, I have focused on mechanisms responsible of immunosuppressive properties of MSC to use them as accessory cells to attenuate the toxicity of the allograft
Guerrero, Julien. « Devenir des cellules souches mésenchymateuses humaines dans un environnement tridimensionnel : application à l’ingénierie du tissu osseux ». Thesis, Bordeaux, 2014. http://www.theses.fr/2014BORD0200/document.
Texte intégralBone tissue engineering aims to resolve the existing limitations of boneregeneration methods. One of the proposed strategies consists on the association,within a three-dimensional (3D) matrix, with autologous cells able to regenerate afunctional 3D tissue. The purpose of this study was therefore to investigate theimpact of cellular communication, between cells of the stromal compartment andendothelial cells, within the three-dimensional porous matrix made of biodegradablenatural polysaccharides, focusing on bone repair. Our results show that thearchitecture and the nature of the 3D macroporous matrix promotes the guidance ofmesenchymal stems cells, derived from human bone marrow, towards theosteoblastic lineage. Also, that the organization in aggregates, promoted by the 3Dmatrices, stimulated cell communication, evidenced by the formation of GAPjunctions and activity of Connexins 43. We also focused on the function ofPannexines 1 and 3 for the 3D culture in these matrices of polysaccharides. Inconclusion, this work shows that cell-cell interactions play a major role in order toimprove bone tissue regeneration. Also, cellular and experimental data demonstratesthe advantage of using a total fraction of bone marrow cells to promote both boneformation and vascularization
Leménager, Hélène. « Mécanismes moléculaires et cellulaires des processus de différenciation et de plasticité cellulaire pour la formation des adipocytes ». Thesis, Toulouse 3, 2019. http://www.theses.fr/2019TOU30227.
Texte intégralAdipocytes are the functional units of adipose tissue (AT). Within white AT, white adipocytes contribute to both storage and release of energy within the organism, mainly in the form of fatty acids1. On the other hand, brown adipocytes, from brown AT, have a high capacity to consume fatty acids. This results from the activity of the UnCoupling Protein 1 (UCP1)2. Finally, UCP1+ adipocytes have been described in white AT, notably in response to cold exposure3. These adipocytes are named beige adipocytes and are generated through two pathways: on one hand via adipogenesis from adipose-derived mesenchymal stem/stromal cells (ASC), and on the other hand by conversion of white-to-beige adipocytes4. Being a reversible process, beige conversion highlights the plasticity of these cells. The aim of the thesis was to characterize the molecular mechanisms involved in both processes, by using culture models of human ASC and in vivo mice models. Given the perivascular and pericyte localization of ASC in vivo5,6, we investigated the use of Endothelial Growth Medium 2 (EGM2) for their in vitro expansion as an alternative to Standard culture conditions (Eagle's medium supplemented with fetal calf serum). Our results showed that the TGFß1 contained in serum of culture medium altered the relative immature state of ASC. Indeed, TGFß1 induces their commitment toward osteoblastic, chondroblastic or vascular smooth muscle lineage. Also, the small amount of serum in EGM2 medium, and thus low TGFß1 concentration, preserves ASC immaturity in culture, as well as their strong capacities to differentiate into adipocytes, including beige phenotype. We showed that ASC with high potential to generate beige adipocytes over-expressed SOX2 protein. Our results also showed that expression of SOX2 was positively correlated to both formation of beige adipocytes and to brown adipocytes activation in vivo in cold-exposed mice. In addition, using two types of human ASC models in vitro, we observed that SOX2 was overexpressed during adipogenesis, and even more when cells were differentiated into beige adipocytes. Thus, SOX2 appears to be a key factor involved in AT browning potential and adipocyte plasticity in vivo and in vitro. This thesis has allowed the access to a better understanding of the impact of culture conditions on the biology of ASC and highlighted molecules involved in the plasticity of adipocytes
Keller, Laetitia. « Ressources cellulaires mésenchymateuses pour l'ingénierie de l'organe dentaire ». Phd thesis, Université de Strasbourg, 2012. http://tel.archives-ouvertes.fr/tel-00766865.
Texte intégralHammoud, Mohammad. « Effet de l’association des basses concentrations d’O2 et des cellules stromales mésenchymateuses sur l’expansion ex vivo des cellules souches et progénitrices hématopoïétiques ». Thesis, Besançon, 2012. http://www.theses.fr/2012BESA3008/document.
Texte intégralTo optimize at best the hematopoietic engraftment, we suggest in this work to improve the ex vivo expansion conditions by moving them closer to physiology. Indeed, we propose to culture placental CD34+ (HSC/PH) on MSC layer in combination with LO2-C to ensure the amplification of HP together with the maintenance/expansion of HSC. Compared to the single culture and/or atmospheric oxygenation, our experimental model allows a better maintenance of primitive HP (Pre-CFC) and HSC together with a quite good amplification of total cells, CD34+ cells and committed HP despite of lower than control condition. Moreover, exogenous IL-3 shows crucial effect in co-culture at LO2-C (1.5% O2) since its addition better preserves and even increases the number of HSC compared to the CD34+ cells control from D0. We then studied the secretion of soluble factors in culture supernatants and found that IL-6, VEGF and IL-8 were present in larger quantities at LO2-C in both co-culture and MSC culture. Finally, the CD146, CD49a, CD54, CD200 and CD105 membrane antigens appear to be up-regulated in MSCs when incubated at 5% O2. However, the involvement of these factors and antigens in paracrine effect and/or direct cell to cell contact mechanisms at LO2-C requires further investigations. In conclusion, the combination of LO2-C and MSC would be promising in the field of HSC/PH grafts expansion to achieve its main objective of reducing the post-transplant cytopenia period together with maintaining the long-term graft potential
Al, Tawil Elias. « Conception de biomatériaux tridimensionnels appropriés au développement de cellules humaines et leur application dans l'ingénierie tissulaire du cartilage et dans l'ingénierie tumorale ». Rouen, 2014. http://www.theses.fr/2014ROUES062.
Texte intégralReppel, Loïc. « Potentialité des cellules stromales de la gelée de Wharton en ingénierie du cartillage ». Thesis, Université de Lorraine, 2014. http://www.theses.fr/2014LORR0164/document.
Texte intégralMesenchymal Stromal/Stem Cells from human Wharton’s jelly (WJ-MSC) are an abundant and interesting source of stem cells for applications in cell and tissue engineering. Their fetal origin confers specific characteristics compared to Mesenchymal Stromal/Stem Cells isolated from human bone marrow (BM-MSC). First, the aim of this work is to optimize WJ-MSC culture conditions for their subsequent clinical use. We focus on the influence of oxygen concentration during monolayer expansion on several parameters to characterize MSC. The results are compared to those obtained with BM-MSC. Our work distinguishes WJ-MSC from BM-MSC in terms of proliferation and adipogenic differentiation. Considering our results, hypoxia during cell expansion is an important parameter to take into account regarding proliferation potential but also chondrogenic differentiation potential. The influence of obstetric factors on WJ-MSC characteristics is also explored. In cartilage tissue engineering context, the second phase of the project is to induce cell differentiation into chondrocytes by seeding them in Alginate/Hyaluronic Acid hydrogel scaffold, and during 28 days. The results obtained are compared to those obtained with BM-MSC. After 4 weeks of culture, WJ-MSC are able to adapt to their environment and express specific cartilage-Related genes and matrix proteins such as type 2 collagen, which is found more expressed after differentiation fromWJ-MSC, than from BM-MSC
Loison-Robert, Ludwig. « Cellule souche gingivale : origine et multipotence ». Thesis, Paris Est, 2016. http://www.theses.fr/2016PESC0083/document.
Texte intégralGingiva is a natural regeneration model thanks to its "ad integrum" healing capability. Gingival fibroblasts are the main actors of this property. These cells, the main cellular component of the gingival connective tissue, regulate the inflammatory responses and healing process. This tissue contains, like many others, mesenchymal stem cells; which also partly explain these regenerative abilities. Moreover, as the gingiva is abundant and easily accessible, the use of these stem cells may interest cell therapy or in vitro model tissues responses. In this work, we demonstrated that Stem Cells Derived from Human Gingiva (SCHG) have common properties with neural crest adult stem cells. These cells can be called "stem cells" for their ability to self-renew, adhere to plastic and to differentiate. First, we have shown that the method and the culture products used for isolation of gingival fibroblasts from gingival biopsy had an influence on the obtained cells. Secondly, an analysis of in vitro clonal populations of gingival fibroblasts has shown that gingival fibroblasts are composed of subpopulations that express specific markers of stem cells and neural crests. In addition to their embryological origin, the study of their multipotency was also characterized after expansion and depending on the used additives. Finally, two examples of using these cells and dental pulp stem cells as a model to study the in vitro biocompatibility of biomaterials have been developed, mimicking oral mucosa or dentin reactions (reparative or reactional)
Basciano, Leticia. « L'effet de l'hypoxie sur les conditions de culture des cellules souches mésenchymateuses de la moelle osseuse ». Thesis, Nancy 1, 2011. http://www.theses.fr/2011NAN10100/document.
Texte intégralIt is now settled that mesenchymal stem cells (MSC), reside in the same microenvironment or niche than hematopoietic stem cells (HSC), within the bone marrow (BM). It is also known that the O2 tension (pO2) of the niche is below 5% as compared to 21% O2 in the air and 12-15% in the arterial blood. As developed in our recent review, this physiological hypoxia protects stem cells from oxidative stress and maintains their multipotential state. Our hypothesis is that MSC cultured in hypoxia should be closer to their physiological condition and therefore more "multipotent". MSC from human BM were cultured et 21% and at 5% pO2. Their morphology, their ability to differentiate into osteocytes and adipocytes, and their transcriptome were compared at different passages. We observed a decrease of proliferation rate in early times in hypoxia, characterized by inhibition of the expression of genes involved in cell replication and cell cycle, and an increase in later passages. Whatever the passage, the genes encoding adhesion molecules and extracellular matrix are stimulated by hypoxia. At later times, the ability of MSC differentiation is stimulated by hypoxia, the cells look to be more immature and show decreased synthesis of mitochondria. Indeed, hypoxia stimulates the synthesis of plasticity genes according to "Gene Ontology" (GO) terms, and of several genes involved in neuronal- and epithelial-cell development. In conclusion, the culture of MSC from BM in hypoxia seems to be more physiological and may be useful for regenerative medicine applications
Ferrari, Caroline. « Études cinétiques de procédés d'expansion de cellules souches mésenchymateuses cultivées sur microporteurs en systèmes agités ». Thesis, Université de Lorraine, 2012. http://www.theses.fr/2012LORR0117/document.
Texte intégralThe extensive use of mesenchymal stem cells (MSC) in tissue engineering increases the necessity to improve the expansion performance. This work aimed at studying an efficient expansion process for porcine MSC in agitated mode. First, a culture medium was adapted to the multipotent porcine MSC. Then, various expansion modes and agitation conditions were evaluated with the cells fixed on microcarriers. Cultures on the Cytodex 1 microcarrier enabled to reach a specific growth rate of 0.54 d-1, which was higher than the one observed in static T-flasks (0.31 d-1), with the same culture conditions. In parallel, an innovative counting method was proposed for the automatic enumeration of cells cultivated on Cytodex 1, without passing by a trypsination step. Finally, the operating conditions of the expansion process were studied. Compared to a culture of MSC on non-agitated Cytodex 1 microcarriers, cell aggregation occurred and an apparent decrease in the cell concentration was observed at an agitation rate of 25 and 75 rpm. Moreover, the addition of microcarriers during a 300 h culture, performed in an agitated culture at 25 rpm and in a volume of 200 mL enabled to prolong the cell proliferation without any aggregation, while maintaining the multipotency of the cells. A cell concentration of 3 x 105 cells/mL was obtained, instead of the 1.2 x 105 cells/mL in static flasks with the same culture conditions. An efficient expansion process for porcine MSC under agitated conditions has therefore been proposed
Ben, Azouna Nesrine. « Etude phénotypique et fonctionnelle des cellules souches mésenchymateuses et hématopoïétiques du sang placentaire en comparaison avec la moëlle osseuse ou le sang périphérique adulte ». Thesis, Tours, 2012. http://www.theses.fr/2012TOUR3321.
Texte intégralThe mesenchymal stem cells (MSC) are adult stem cells which are at the origin of the ostebiastic lignage cells (O), adipocytes (A) and chondrobiasts (C). The MSC are initially found in the bone marrow (BM) but it also exists there in other tissues as the umbilical cord blood (UCB).Endowed with a regenerative potential, MSC can used in diverse degenerative pathologies in a purpose of tissular repair. Besides, their immunosuppressive properties allowed to envisage their use in a purpose of immunomodulation as during the reactions of transplant against the host in allogenic hematopoietic stem cell transplants (HSC). The essential purpose of this work was to compare the characteristics of the MSC derived from the UCB in comparison to those stemming from the bone marrow (BM)
Loubière, Céline. « Characterization and impact of the hydrodynamics on the performance of umbilical-cord derived stem cells culture in stirred tank bioreactors ». Electronic Thesis or Diss., Université de Lorraine, 2018. http://www.theses.fr/2018LORR0220.
Texte intégralMesenchymal stem cells (MSC) are becoming increasingly involved in the regenerative medicine field, particularly to treat diseases that are not effectively curable with the current therapies. Two scientific barriers are nevertheless responsible for MSC use and commercialization limitations. On one side, large amounts of cells are needed to reach the high cell dose requirements. On the other side, cells being the final product themselves, directly injected into the patient, their quality have to be controlled (stem cell phenotype, differentiation capability). MSC cultivation on microcarriers in a stirred bioreactor seems to meet these challenges. However, a precise knowledge about the impact of the technologies and the hydrodynamics generated, on the physiological cell response, is necessary to improve the scale-up of MSC cultures in bioreactors. In this context, present work is dedicated to the study of the impact of the agitation mode (orbital or mechanical) on the cell attachment, expansion and detachment on various microcarrier types, in the case of MSC derived from the Wharton’s jelly (WJ-MSC) of umbilical cords. To quantify more precisely cell distribution and expansion on microcarriers, an automatic and in situ counting method was developed, which need no detachment step. This allowed the identification of commercial microcarriers suitable for WJ-MSC cultures, which were then compared to home-made microcarriers, synthesized by a partner laboratory, in terms of cell attachment and expansion, and detachment efficiency. In parallel to these works, the impact of the impeller design on the microcarrier suspension in stirred tank bioreactors was investigated. Based on a dimensional analysis and CFD simulations, it resulted in the establishment of two models relating the minimal agitation rate to ensure all particle suspension (Njs) with the impeller geometrical characteristics (design, size, off-bottom clearance) and the material properties of both the solid and the liquid phases. CFD models validation allowed then to develop a strategy to optimize the geometrical configuration of an impeller, dedicated to MSC cultures on microcarriers in a minibioreactor. Parameters characterizing the hydromechanical stress encountered by the solid phase were wisely chosen and integrated into CFD simulations. Based on a design of experiments, and the hydrodynamics data recovered from simulations, response surfaces were built and a multiobjective optimization was achieved in order to determine the geometry minimizing the particle stress, and also by adhered cells. WJ-MSC cultures in minibioreactors equipped with impellers displaying various geometries were finally validated, with a preliminary comparison of the impact of these geometries on the cell expansion
Guérin, Coralie. « Biomarqueurs cellulaires circulants de la dysfonction endothéliale : détection et potentiel vasculaire ». Thesis, Paris 5, 2014. http://www.theses.fr/2014PA05P608.
Texte intégralIn endothelial dysfunction, circulating endothelial compartment simultaneously plays the role of actor involved in the regeneration of injured tissue and reflects endothelium state. In peripheral arterial disease (PAD), one of the research areas is the development of a cellular therapy product capable of inducing the formation of neo-Vessels. Faced with the difficulty to obtain and amplify endothelial progenitor cells (EPC) in adults, one of the assumptions lets consider the use of other cell types with vasculogenic properties. In patients with cardiovascular disease, and PAD in particular, bone marrow mononuclear cells and EPC show reduced angiogenic properties. We have demonstrated the ability of isolated mesenchymal stem cells (MSCs) from PAD patients to induce reperfusion by recruitment of endothelial cells in situ, with the same efficiency as that of healthy donors MSCs. MSCs do not differentiate into endothelial cells but act by paracrine. The second hypothesis of obtaining an autologous angiogenic cell therapy product is to sort cells more immature than the CPE and to differentiate them secondarily into endothelial lineage as the pathological cell model of hemangioma stem cells CD133 + which lets consider the Very Small Embryonic like stem cells (VSEL), CD133 + multipotent stem cells as a potential candidate of postnatal vascular cell. We have derived and cultured in angiogenic conditions VSEL that acquired a mesenchymal phenotype but exhibited a secretory profile similar to that of EPC. VSEL promote post-Ischemic revascularization and acquire an endothelial phenotype in vitro and in vivo suggesting that VSEL may be responsible for the endothelial lineage. VSEL also appear as a biomarker of endothelial dysfunction mobilized from bone marrow (BM) to peripheral blood (PB) in patients with PAD. Cellular circulating biomarkers are not only non-Invasive markers of endothelium but can also provide useful information for the diagnosis, prognosis and therapeutic monitoring of patients with endothelial dysfunction associated pathologies. Changing the number of EPC and circulating endothelial cells (CEC) in the circulation has been reported in different pathological situations respectively associated with endothelial regeneration and alteration such as the increase of CEC in patients with pulmonary arterial hypertension (PAH). The reference technique for the enumeration of CEC in peripheral blood is magnetic immunoseparation (IMS). This non-Automated and time-Consuming method, based on the enumeration by fluorescence microscopy of CD146 + cells isolated. Although reproducible, this count is subject to many through quantification, difficult to implement and subject to interpretation. The development of an acoustic focusing cytometry (AFC) method for automated detection of CEC has proved reliable and robust results, in a cohort of patients with PAH treated or not, constituting a relevant alternative analysis to microscopy. All of this work opens new perspectives in the detection of cellular circulating biomarkers involved in endothelial dysfunction, suggesting VSEL as new vasculogenic actor
Loubière, Céline. « Characterization and impact of the hydrodynamics on the performance of umbilical-cord derived stem cells culture in stirred tank bioreactors ». Thesis, Université de Lorraine, 2018. http://www.theses.fr/2018LORR0220/document.
Texte intégralMesenchymal stem cells (MSC) are becoming increasingly involved in the regenerative medicine field, particularly to treat diseases that are not effectively curable with the current therapies. Two scientific barriers are nevertheless responsible for MSC use and commercialization limitations. On one side, large amounts of cells are needed to reach the high cell dose requirements. On the other side, cells being the final product themselves, directly injected into the patient, their quality have to be controlled (stem cell phenotype, differentiation capability). MSC cultivation on microcarriers in a stirred bioreactor seems to meet these challenges. However, a precise knowledge about the impact of the technologies and the hydrodynamics generated, on the physiological cell response, is necessary to improve the scale-up of MSC cultures in bioreactors. In this context, present work is dedicated to the study of the impact of the agitation mode (orbital or mechanical) on the cell attachment, expansion and detachment on various microcarrier types, in the case of MSC derived from the Wharton’s jelly (WJ-MSC) of umbilical cords. To quantify more precisely cell distribution and expansion on microcarriers, an automatic and in situ counting method was developed, which need no detachment step. This allowed the identification of commercial microcarriers suitable for WJ-MSC cultures, which were then compared to home-made microcarriers, synthesized by a partner laboratory, in terms of cell attachment and expansion, and detachment efficiency. In parallel to these works, the impact of the impeller design on the microcarrier suspension in stirred tank bioreactors was investigated. Based on a dimensional analysis and CFD simulations, it resulted in the establishment of two models relating the minimal agitation rate to ensure all particle suspension (Njs) with the impeller geometrical characteristics (design, size, off-bottom clearance) and the material properties of both the solid and the liquid phases. CFD models validation allowed then to develop a strategy to optimize the geometrical configuration of an impeller, dedicated to MSC cultures on microcarriers in a minibioreactor. Parameters characterizing the hydromechanical stress encountered by the solid phase were wisely chosen and integrated into CFD simulations. Based on a design of experiments, and the hydrodynamics data recovered from simulations, response surfaces were built and a multiobjective optimization was achieved in order to determine the geometry minimizing the particle stress, and also by adhered cells. WJ-MSC cultures in minibioreactors equipped with impellers displaying various geometries were finally validated, with a preliminary comparison of the impact of these geometries on the cell expansion