Simulation of solute transport in a heterogeneous vadose zone describing the hydraulic properties using a multistep stochastic approach

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/95612/1/wrcr10619.pd

Impact of ligand presentation on biophysics of T-lymphocyte adhesion

Les lymphocytes T jouent un rôle central dans la réponse immunitaire adaptative. par l'intermédiaire d'un récepteur distinctif - le RCT (Récepteur des Cellules T), qui est responsable de la reconnaissance spécifique de peptides étrangers dans le corps. Les interactions moléculaires entre le RCT et son ligand, présenté par des cellules spécialisées (CPA cellules présentatrices d'antigène) sont suivies par un processus d'adhésion et d'étalement cellulaire caractérisé par d'importantes réorganisations à l'échelle supramoléculaire et cellulaire. Le rôle des propriétés physiques des cellules à ces échelles est encore mal compris. Dans cette thèse, nous nous sommes intéressés aux événements suivant cette reconnaissance en mimant la CPA par des substrats artificiels contrôlés exploitant plusieurs techniques de microscopie avancé. Nous avons réalisé une étude systématique de la dynamique d'étalement ainsi que de la morphologie (en terme d'aire d'adhésion et d'organisation de l'actine) des lymphocytes T humains de la lignée Jurkat sur des substrats présentant des anti-CD3 (un ligand spécifique du RCT) mobiles ou non. L'étalement des LT est fortement réduit sur les ligands mobiles par rapport aux ligands fixes. De plus le blocage de la myosine-II ou l'inclusion d'ICAM (le ligand des intégrines LFA-1) sur les substrats permettent de compenser ces différences. Nous proposons un modèle, inspiré de ceux sur la migration ou la croissance du cône neuronal , qui lie la dynamique du front d'étalement des lymphocytes T à la friction générée au niveau de la surface en tirant ou en fixant les ligands.T- cells play a central role in cell mediated adaptive immune response. They are a class of lymphocytes that carry distinctive receptors - the TCRs (T cell receptor), which are responsible for specific recognition of foreign peptides in the body. Molecular interactions between the TCR complex and its ligand is followed by cell adhesion and spreading accompanied by dramatic changes at molecular and cellular scale. In this thesis we focus on the events that immediately follow recognition. I report a systematic study of the spreading dynamics and morphology (in terms of adhesion area and actin organization) of T lymphocytes on substrates with mobile or immobilized anti-CD3 (a ligand targeting the TCR complex). The spreading behaviour is seen to be dramatically different on the two types of substrates. Interestingly, on blocking myosin-II or by inclusion of ICAM (ligand of LFA1 integrins) on the substrate, these differences tend to disappear. We propose a model, partially inspired by earlier models of neuronal growth cones and filopodia, that links the dynamics of the leading edge of the spreading T cell to the friction generated at the surface by dragging or pinning of the ligands. I also report studies on T cells interacting with substrates decorated with nano-scale islands of anti-CD3 in a sea of repulsive PEG or a supported lipid bilayer. Dramatic differences in adhesion, organization of actin, and receptor distribution are observed on different geometries. I report advances in multi channel RICM towards the reconstruction of not only the topography of the proximal membrane of the T cell with high level of confidence but also the distal surface of lamellipodia

Multichannel Image Regularisation Using Anisotropic Geodesic Filtering

This paper extends a recent image-dependent regularisation approach introduced in [11, 12] aiming at edge-preserving smoothing. For that purpose, geodesic distances equipped with a Riemannian metric need to be estimated in local neighbourhoods. By deriving an appropriate metric from the gradient structure tensor, the associated geodesic paths are constrained to follow salient features in images. Following, we design a generalised anisotropic geodesic filter, incorporating not only a measure of the edge strenght, like in the original method, but also further directional information about the image structures. The proposed filter is particularly efficient at smoothing heterogeneous areas while preserving relevant structures in multichannel imagesJRC.DG.G.2-Global security and crisis managemen

Nano-clustering of ligands on synthetic APCs influences T-cell membrane and actin organization

International audienc

Nanometric Protein-Patch Arrays on Glass and Polydimethylsiloxane for Cell Adhesion Studies

International audienc

Biphasic mechanosensitivity of T cell receptor-mediated spreading of lymphocytes

International audienc

Nano-clustering of ligands on surrogate antigen presenting cells modulates T cell membrane adhesion and organization

International audienceWe investigate the adhesion and molecular organization of the plasma membrane of T lymphocytes interacting with a surrogate antigen presenting cell comprising glass supported ordered arrays of antibody (alpha-CD3) nano-dots dispersed in a non-adhesive matrix of polyethylene glycol (PEG). The local membrane adhesion and topography, as well as the distribution of the T cell receptors (TCRs) and the kinase ZAP-70, are influenced by dot-geometry, whereas the cell spreading area is determined by the overall average density of the ligands rather than specific characteristics of the dots. TCR clusters are recruited preferentially to the nano-dots and the TCR cluster size distribution has a weak dot-size dependence. On the patterns, the clusters are larger, more numerous, and more enriched in TCRs, as compared to the homogeneously distributed ligands at comparable concentrations. These observations support the idea that non-ligated TCRs residing in the non-adhered parts of the proximal membrane are able to diffuse and enrich the existing clusters at the ligand dots. However, long distance transport is impaired and cluster centralization in the form of a central supramolecular cluster (cSMAC) is not observed. Time-lapse imaging of early cell-surface contacts indicates that the ZAP-70 microclusters are directly recruited to the site of the antibody dots and this process is concomitant with membrane adhesion. These results together point to a complex interplay of adhesion, molecular organization and activation in response to spatially modulated stimulation

Targeting KRAS mutations with HLA class II-restricted TCRs for the treatment of solid tumors

T-cell receptor (TCR) redirected T cells are considered as the next generation of care for the treatment of numerous solid tumors. KRAS mutations are driver neoantigens that are expressed in over 25% of all cancers and are thus regarded as ideal targets for Adoptive Cell Therapy (ACT). We have isolated four KRAS-specific TCRs from a long-term surviving pancreatic cancer patient vaccinated with a mix of mutated KRAS peptides. The sequence of these TCRs could be identified and expressed in primary cells. We demonstrated stable expression of all TCRs as well as target-specific functionality when expressing T cells were co-incubated with target cells presenting KRAS peptides. In addition, these TCRs were all partially co-receptor independent since they were functional in both CD4 and CD8 T cells, thus indicating high affinity. Interestingly, we observed that certain TCRs were able to recognize several KRAS mutations in complex with their cognate Human leukocyte antigen (HLA), suggesting that, here, the point mutations were less important for the HLA binding and TCR recognition, whereas others were single-mutation restricted. Finally, we demonstrated that these peptides were indeed processed and presented, since HLA-matched antigen presenting cells exogenously loaded with KRAS proteins were recognized by TCR-transduced T cells. Taken together, our data demonstrate that KRAS mutations are immunogenic for CD4 T cells and are interesting targets for TCR-based cancer immunotherapy