Rôle du microenvironnement dans le maintien et la résistance des cellules souches leucémiques de la Leucémie Myéloïde Chronique. voie BMP et contraintes mécaniques

One of the main causes of treatment failure in cancers is the development of drug resistance by cancer cells. The persistence of cancer stem cells (CSCs) might explain cancer relapses as they could allow reactivation of cancer cells proliferation following therapy, leading to disease persistence and ultimately to patients’ death. Clinically, it is crucial to develop therapeutic strategies able to target resistant CSCs in order to cure the patients. CSCs are controlled by a variety of biochemical and biomechanical signals from the leukemic niche. My project aims to determine the involvement of the tumor microenvironment (BMP signaling pathway and mechanical stress) in the maintenance and resistance of Leukemic Stem Cells (LSCs) in Chronic Myelogenous Leukemia (CML). For this, we combined functional and molecular assays to the analysis of tumor microenvironment on more than 200 CML patients’ samples. We demonstrated that alterations of intracellular BMP signaling pathway in CP-CML primary samples corrupt and amplify the response to exogenous BMP2 and BMP4, which are abnormally abundant in the tumor microenvironment. These results, recently published in Blood led us to evaluate the role of the BMP pathway in LSC maintenance under TKI treatment. Atomic force microscopy allowed us to demonstrate that BCR-ABL expression alone is sufficient to increases the rigidity of immature CML cells compared to healthy ones. Finally, using a unique cell confining system, we were able to demonstrate that mechanical stress controls the proliferation of immature leukemic cells by regulating the expression of mechano-sensitive genes such as Twist-1. These results could explain how LSCs can benefit from a mechanical stress exerted by their microenvironment to acquire a proliferative advantage over normal cells. Ultimately, we hope that this transdisciplinary approach will help to identify key molecules in the transduction of mechanical signals potentially involved in maintenance and resistance of CSCs and thus offer new targets to counter these effects.Une des principales causes d’échec dans le traitement des cancers est le développement de résistances aux drogues par les cellules tumorales. Les cellules souches cancéreuses (CSC) sont suspectées d’être responsables de ces rechutes, conduisant à la récurrence de la maladie et bien souvent au décès des patients. En clinique, il est donc nécessaire de développer des stratégies thérapeutiques capables de cibler ces CSC résistantes et aboutir à la guérison des patients. Les CSC sont régulées par un ensemble de signaux aussi bien biologiques que physiques au sein de la niche tumorale. Mon projet a pour objectif de déterminer l’implication du microenvironnement tumoral (voie de signalisation BMP et contraintes mécaniques) dans le maintien et la résistance des cellules souches leucémiques (CSLs) de la leucémie myéloïde chronique (LMC). Pour cela, nous avons combiné tests fonctionnels et moléculaires ainsi que l’analyse de la niche tumorale sur plus de 200 échantillons de patients atteints de LMC. Nous avons ainsi démontré que l’altération de la voie BMP intrinsèque aux cellules immatures de la LMC corrompt et amplifie la réponse à BMP2 et BMP4, présents en quantités anormalement abondantes au sein de la niche tumorale. Ces résultats récemment publiés dans Blood nous ont amenés à évaluer le rôle de la voie BMP dans le maintien des CSLs sous traitement par les ITK. La microscopie à force atomique nous a permis de démontrer que l’expression de BCR-ABL est suffisante pour induire une augmentation de la rigidité des cellules immatures de LMC par rapport à des cellules saines. Enfin, l’utilisation d’un système de confinement cellulaire nous a permis de démontrer que le stress mécanique contrôle la prolifération des cellules leucémiques immatures en régulant l’expression de gènes mécano-sensibles comme Twist-1. Ces résultats pourraient expliquer comment des CSLs tirent profit des contraintes mécaniques issues de leur microenvironnement afin d’acquérir un avantage prolifératif par rapport aux cellules saines. Ultimement, nous espérons que cette approche transdisciplinaire permettra d’identifier les molécules clés de la transduction de signaux mécaniques potentiellement impliqués dans le maintien et la résistance des CSC et ainsi proposer de nouvelles cibles pour contrer ces effets

Incidence of Apoptosis in the Lymphoid Organs of Normal or Malaria Infected Mice is Decreased in CD18 and Urokinase - Receptor (UPAR, CD87) Deficient Mice

Incidence of apoptosis was investigated in the spleen and lymph nodes of +/+, CD18 -/- and urokinase receptor (uPAR, CD87) -/- mice, untreated or Plasmodium Berghei Anka (PbA) infected. In non infected mice, incidence of apoptosis was lower in the lymph nodes of CD18 -/- and uPAR -/- than in +/+ mice, as seen by FACS analysis to count the number of hypodiploid and Annexin-V binding cells. Infection of mice with PbA resulted in a marked increase in the size of spleen and lymph nodes 7–8 days after infection, which was slightly higher in uPAR -/- and CD18 -/- than in +/+ mice. PbA infection increased about 7 fold the incidence of apoptosis in the lymphoid organs of +/+, especially in the white pulp and germinal centers of the spleen and lymph nodes, while in contrast it was unchanged in PbA infected CD18 -/- or uPAR -/- mice. Serum IgG levels, and number of circulating leukocytes were significantly higher in both uPAR and CD18 -/- than in +/+ mice. These results indicate that the CD18 and uPAR surface molecules, which are known to be associated in the cell membrane, have an important influence upon the incidence of cell survival in both normal or stimulated lymphoid organs

Cavitation Erosion Prediction on Francis Turbines Part 3 : Methodologies of Prediction

In the frame of a joint research program between EDF, Hydro Québec and IMHEF, different methods are investigated to predict cavitation erosion on Francis turbines from model. They are based on measurement of pitting, pressure fluctuations and acceleration. The measurement techniques have been detailed in Part I and Part 2. The present article describes essentially the theoretical and practical aspects of the methods and discusses the results obtained until now from the mode! and prototype tests. The first analysis shows that the methods proposed are suitable to measure cavitation aggressiveness on mode! and on prototype, and that the level on the mode! is several orders of magnitude smaller than on the prototype. To adjust transposition laws, a more complete set of data is needed

Direct transfection of clonal organoids in matrigel microbeads : a promising approach toward organoid-based genetic screens

Organoid cultures in 3D matrices are relevant models to mimic the complex in vivo environment that supports cell physiological and pathological behaviors. For instance, 3D epithelial organoids recapitulate numerous features of glandular tissues including the development of fully differentiated acini that maintain apico-basal polarity with hollow lumen. Effective genetic engineering in organoids would bring new insights in organogenesis and carcinogenesis. However, direct 3D transfection on already formed organoids remains challenging. One limitation is that organoids are embedded in extracellular matrix and grow into compact structures that hinder transfection using traditional techniques. To address this issue, we developed an innovative approach for transgene expression in 3D organoids by combining single-cell encapsulation in Matrigel microbeads using a microfluidic device and electroporation. We demonstrate that direct electroporation of encapsulated organoids reaches up to 80% of transfection efficiency. Using this technique and a morphological read-out that recapitulate the different stages of tumor development, we further validate the role of p63 and PTEN as key genes in acinar development in breast and prostate tissues. We believe that the combination of controlled organoid generation and efficient 3D transfection developed here opens new perspectives for flow-based high-throughput genetic screening and functional genomic applications

Highly integrated multi-material fibers for soft robotics

Soft robots are envisioned as the next generation of safe biomedical devices in minimally invasive procedures. Yet, the difficulty of processing soft materials currently limits the size, aspect-ratio, manufacturing throughput, as well as, the design complexity and hence capabilities of soft robots. Multi-material thermal drawing is introduced as a material and processing platform to create soft robotic fibers imparted with multiple actuations and sensing modalities. Several thermoplastic and elastomeric material options for the fibers are presented, which all exhibit the rheological processing attributes for thermal drawing but varying mechanical properties, resulting in adaptable actuation performance. Moreover, numerous different fiber designs with intricate internal architectures, outer diameters of 700 µm, aspect ratios of 103, and a fabrication at a scale of 10s of meters of length are demonstrated. A modular tendon-driven mechanism enables 3-dimensional (3D) motion, and embedded optical guides, electrical wires, and microfluidic channels give rise to multifunctionality. The fibers can perceive and autonomously adapt to their environments, as well as, probe electrical properties, and deliver fluids and mechanical tools to spatially distributed targets

Cavitation Erosion Prediction on Francis Turbines Part 2 : Model Tests and Flow Analysis

Different measurement techniques have been used to detect cavitation on a Francis turbine model. The results are compared to those obtained on the prototype and presented in the first of this series of articles. The runner mode! used for that study is built on the basis of a geometrical recovery of one of most eroded blade of the prototype. The results of the different measurements are presented and commented by comparison with prototype measurements. This comparison leads to a proposal of the physics which should be involved in transposition laws for the prediction of prototype erosion from cavitation mode! tests. The consequences of such scaling laws, as well as their application to the prototype and mode! results, are part of the third facet of this work

Restauration morpho-dynamique et redynamisation de la section court-circuitée du Rhin en aval du barrage de Kembs (projet INTERREG / EDF)

National audienceThe Upper Rhine River has been heavily impacted by channelization for flood protection and navigation, and then by damming for hydropower generation. In normal non flooding conditions, most of the flows are diverted in a canalized section whereas the regulated “old Rhine” bypassed reach runs a minimum flow. Between Huningue and Neuf-Brisach, engineering works induced simplification and stabilization of the channel pattern from a formerly braiding sector to a single incised channel, hydrological modifications, bottom armouring due to bedload decrease, and thus ecological alterations. Two complementary and interdisciplinary projects have been initiated to restore alluvial morphodynamics: i) the international “INTERREG IV - Redynamisation of the old Rhine” project (2009-2012) coordinated by the Alsace region, France; ii) the left bank “controlled erosion” project launched by Electricité de France (EDF) within Kembs hydroelectric station relicensing process since 2003-2004. The purpose of these projects is to evaluate the feasibility of an important hydro-morphological and ecological restoration plan on a 45 km long reach, through both field testing of bank erosion techniques at favourable locations, and artificial sediments input from right bank excavations. This will help define possible long term prospective scenarios, in order to restore sustainable sediment transport, morphodynamics variability and associated ecological functions. The study will involve historical analysis, hydro-morphological / hydraulic physical and numerical modelling, physical and ecological monitoring, and sociological aspectsLe Rhin alsacien-allemand a enregistré de profondes modifications morphologiques et hydrologiques à la suite de sa correction et de sa régularisation pour la protection contre les crues et la navigation, puis après la construction de barrages hydro-électriques. Les aménagements réalisés entre Huningue et Neuf-Brisach ont engendré une simplification et une stabilisation du style fluvial. Un fleuve en tresses a cédé la place à un chenal unique incisé. Le fond de chenal est devenu pavé à cause d’une diminution des apports de charge de fond et des altérations écologiques ont été observées (simplification des habitats aquatiques et riverains). Deux projets complémentaires et interdisciplinaires ont été engagés afin de restaurer une dynamique des formes alluviales : i) le projet international INTERREG IV – Redynamisation du Vieux Rhin (2009-2012) sous l’impulsion de la région Alsace ; ii) le projet d’érosion maitrisée des berges de la rive gauche conduit par Electricité de France (EDF) dans le cadre du renouvellement de la concession de l’aménagement de Kembs. L’objectif des deux projets est de définir un plan de restauration hydro-morphologique et écologique conduisant à la redynamisation d’un tronçon de 45 km. L’étude repose sur une analyse historique, l’exploitation de modèles à la fois physiques et numériques, et les suivis morphologiques in situ d’une recharge artificielle en sédiments et d’érosions de berge contrôlées. Ces études de faisabilité sont complétées par des analyses écologique et sociologique pour apprécier l’impact socio-environnemental de ces projets

Role of the bone morphogenic protein pathway in developmental haemopoiesis and leukaemogenesis

Myeloid leukaemias share the common characteristics of being stem cell-derived clonal diseases, characterised by excessive proliferation of one or more myeloid lineage. Chronic myeloid leukaemia (CML) arises from a genetic alteration in a normal haemopoietic stem cell (HSC) giving rise to a leukaemic stem cell (LSC) within the bone marrow (BM) ‘niche’. CML is characterised by the presence of the oncogenic tyrosine kinase fusion protein breakpoint cluster region-abelson murine leukaemia viral oncogene homolog 1 (BCR-ABL), which is responsible for driving the disease through activation of downstream signal transduction pathways. Recent evidence from our group and others indicates that important regulatory networks involved in establishing primitive and definitive haemopoiesis during development are reactivated in myeloid leukaemia, giving rise to an LSC population with altered self-renewal and differentiation properties. In this review, we explore the role the bone morphogenic protein (BMP) signalling plays in stem cell pluripotency, developmental haemopoiesis, HSC maintenance and the implication of altered BMP signalling on LSC persistence in the BM niche. Overall, we emphasise how the BMP and Wnt pathways converge to alter the Cdx–Hox axis and the implications of this in the pathogenesis of myeloid malignancies