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We present a theoretical framework based on an extension of Dynamical Density Functional Theory (DDFT) to describe the structure and dynamics of cells in living tissues and tumours. DDFT is a microscopic statistical mechanical theory for the time evolution of the density distribution of interacting many-particle systems. The theory accounts for cell pair-interactions, different cell types, phenotypes and cell birth and death processes (including cell division), in order to provide a biophysically consistent description of processes bridging across the scales, including the description of the tissue structure down to the level of the individual cells. Analysis of the model is presented for a single species and a two-species cases, the latter describing competition between a cancerous and healthy cells. In suitable parameter regimes, model results are consistent with biological observations. Of particular note, divergent tumour growth behaviour, mirroring metastatic and benign growth characteristics, are shown to be dependent on the cell pair-interaction parameters.

Alors que le rôle des cellules du système immunitaires innées sur la progression tumorale est l'objet d'une investigation croissante, le rôle des neutrophiles sur la sensibilité à la thérapie n'a pas été précédemment décrit. Jusqu’au présent, nous avons effectué des cocultures de neutrophiles et des différentes lignées cellulaires de lymphome non hodgkinien (LNH) en présence de divers agents cytotoxiques ou des thérapies ciblées. Afin d’évaluer l'effet du traitement sur la prolifération cellulaire et la mort des cellules, des marquages CFSE et DAPI ont été effectués respectivement, en utilisant la cytométrie en flux. Les neutrophiles ainsi que les cellules HL60 différenciées avec des propriétés de neutrophiles, ont atténué la sensibilité de cellules de lymphome à des agents anticancéreux in vitro, à la fois dans les modèles 2D et 3D. L'effet protecteur des neutrophiles a été testée in vivo en injectant des cellules de LNH et des neutrophiles chez des souris SCID/CB17 traités avec vincristine. La coinjection de neutrophiles réduit la sensibilité des cellules LNH à la chimiothérapie. Cet effet protecteur a été validé en utilisant des cellules primaires, purifiée à partir de patients atteints de leucémie lymphoïde chronique, exposés à des agents cytotoxiques ou des agents ciblés en présence de neutrophiles autologues. La protection par les neutrophiles est contact dépendante. Elle est médiée par l'interaction de CD11b et ICAM1, exprimé par les neutrophiles et les lymphocytes B, respectivement, et par la molécule d'adhésion CD44. Elle est également dépendante de Mcl1 et est partiellement abrogée par un composé anti-Mcl1 / While the role of innate immune cells on tumor progression is the object of increasing scrutiny, the role of neutrophils on sensitivity to therapy has not been previously described. To this end, we performed cocultures of freshly purified human neutrophils and different non- Hodgkin lymphoma (NHL) cell lines in the presence of various cytotoxic and targeted agents. CFSE and DAPI assays were performed to assess the therapeutic effect on cell proliferation and cell death, respectively, using flow cytometry. Neutrophils and differentiated HL60 cells with neutrophil-like properties attenuated the sensitivity of lymphoma cells to anti-cancer agents both in 2D and 3D models in vitro. The protective effect of neutrophils was tested in vivo using SCID/CB17 mice inoculated with NHL cells together with neutrophils, and treated with vincristine. The co-inoculation of neutrophils reduced the sensitivity of NHL cells to chemotherapy. Similar findings were made on primary cells, purified from patients diagnosed with chronic lymphocytic leukemia, exposed to cytotoxic agents or recently approved targeted agents (ibrutinib and idelalisib) in the presence of autologous neutrophils. Neutrophil-induced protection was dependent on cell-cell contact mediated by the interaction of CD11b and ICAM-1, expressed by neutrophils and B cells respectively, and by the adhesion molecule CD44. This protective effect was Mcl-1-dependent and was partially abrogated by an anti- Mcl-1 compound

<p>The gonad is a unique primordial organ that retains the ability to adopt one of two morphological fates through much of mammalian embryonic development. Previous work in our lab found that dimorphic vascular remodeling was one of the earliest steps during sex-specific morphogenesis. In particular, vessels in XY gonads display highly ordered behavior that coincides with testis cord formation. It was unknown how the vasculature may influence testis cord morphogenesis and, if so, how this was mechanistically related to sex determination. The work in this thesis addresses a single over-arching hypothesis: Male-specific vascular remodeling is required for testis morphogenesis and orchestrates differentiation of the XY gonad. </p><p>To address this question we have modified and developed techniques that allow us to isolate aspects of vascular behavior, gene expression, and endothelial influence on surrounding cells. In particular, the application of live imaging was instrumental to understanding the behavior of various gonadal cell-types in relation to remodeling vessels. It is difficult to grasp the complexity of an organ without understanding the dynamics of its constituents. A critical aim of my work was to identify specific inhibitors of the vasculature that do not affect the early stages of sex determination. Combining inhibitors, live imaging, cell sorting, qRT-PCR, mouse models, and whole organ culture has led to a far richer understanding of how the vasculature behaves and the cell-types that mediate its influence on organ morphogenesis. The beauty of our system is that we do not have to settle for a snapshot of the fate of cells in vivo, but can document their journeys and their acquaintances along the way. </p><p>Vascular migration is required for testis cord morphogenesis. Specific inhibitors revealed that in the absence of vessels, testis cords do not form. The work below shows that vessels establish a feedback loop with mesenchymal cells that results in both endothelial migration and subsequent mesenchymal proliferation. Interstitial control of testis morphogenesis is a new model within the field. The mechanisms regulating this process include Vegf mediated vascular remodeling, Pdgf induced proliferation, and Wnt repression of coordinated endothelial-mesenchymal dynamics. Our work also suggests that vascular patterning underlies testis patterning and, again, is mediated by signals within the interstitial space not within testis cords themselves. </p><p>A final aspect of my work has been focused on how vessels continue to influence morphology of the testis and the fate of surrounding cells. Jennifer Brennan, a graduate student in our lab, previously showed that loss of Pdgfr&#945; antagonizes cord formation and development of male-specific lineages. The mechanisms and cell-types related to this defect were not clear. I began to reanalyze Pdgfr&#945; mutants after finding remarkable similarity to gonads after vascular inhibition. This work is providing data suggesting that vessels are not simply responsible for testis morphology but also for the fate of specialized cells within the testis. On the whole, this thesis describes specific roles for endothelial cells during gonad development and mechanisms by which they are regulated.</p> / Dissertation

Microcystine sind die wohl bekanntesten cyanobakteriellen Toxine. Sie werden im Wesentlichen durch die im Süßwasser weltweit verbreitete, koloniebildende Gattung Microcystis synthetisiert. Die biologische Funktion dieser Peptide ist jedoch ungeklärt. In dieser Studie wurde die Fragestellung erstmals über einen globalen Ansatz auf molekularer Ebene analysiert. Die proteomischen Analysen zwischen M. aeruginosa PCC 7806/ Wildtyp und einigen Microcystin-freien Mutanten deuten auf eine physiologische Rolle der Microcystine. Microcystine beeinflussen die Abundanz zahlreicher Proteine. Prominentester Vertreter ist RubisCO – Schlüsselenzym des Calvin Zyklus. RubisCO und andere im 2D selektierte Proteine konnten außerdem als mögliche zelluläre Bindepartner des Microcystins identifiziert werden. Möglicherweise bindet MC an bestimmte Cysteinreste dieser Proteine. Mit dem Knockout der mcy-Gene geht außerdem eine Überexpression eines Morphotyp-spezifischen Proteins einher, das MrpC genannt wurde. Dieses Protein vermittelt möglicherweise Zell-Zell-Interaktionen in Microcystis. / Microcystins are the most common cyanobacterial toxins found in freshwater lakes and reservoirs throughout the world. They are frequently produced by the unicellular, colonial cyanobacterium Microcystis; however, the role of the peptide for the producing organismen is poorly understood. In this study we describe the first global approach to investigate this topic on a molecular level. Proteomic studies with M. aeruginosa PCC 7806 wild-type and several microcystin-deficient mutants indicated a physiological function for microcystin. Microcystin was shown to influence the abundance of several proteins which have an intra- or extracellular function. A prominent candidate is RubisCO, the key enzyme of the calvin cycle. RubisCO and other proteins, initially selected by 2D analysis, are putative cellular binding partners of microcystin. A potentially interaction mechanismen is the kovalent binding of microcystin to cysteine residues of the protein. Moreover, several knockouts of microcystin biosynthesis genes result in an overexpression of a putative morpho-type specific factor, named MrpC. This protein possibly mediates cell-cell interactions in Microcystis.

La motilité cellulaire est fondamentale dans de nombreux processus physiologiques, qu’ils soient normaux ou pathologiques. Cependant, bien que ces derniers impliquent la plupart du temps de nombreuses cellules se mouvant en même temps, les effets des interactions entre cellules sur leur dynamique, à la fois individuelle et collective, restent assez mal connu. Dans cette thèse, j’ai utilisé Dictyostelium discoideum à l’état végétatif pour étudier cette régulation collective de la motilité. Je me suis principalement appuyé sur une analyse minutieuse de nombreuses trajectoires cellulaires dans des situations variées pour (i) caractériser un facteur sécrété qui régule négativement la motilité cellulaire (nature chimique, voie de signalisation, dynamique de sécrétion et de réponse) et (ii) analyser et modéliser quantitativement la dynamique d’étalement de colonie cellulaires de forme, dimension et densité contrôlées. Je décris enfin un phénomène d’agrégation dynamique observé lorsque les cellules sont placées à haute densité dans un milieu nutritif / Cell motility is fundamental in many physiological, either normal or pathological, phenomena. Yet, although these most often involve several cells moving at the same time, how the interactions between cells affect both individual and collective dynamics remains a poorly understood question. In this thesis, I used vegetative Dictyostelium discoideum cells as a model to study this collective regulation of the motility. I relied mainly on the thorough analysis of numerous cell trajectories in various situations to (i) characterise a secreted factor used to down-regulate the cells’ motility (biochemical nature, response pathway, secretion and response dynamics) and (ii) quantitatively analyse and model the dynamics of spreading cell colonies of controlled initial shape, size and density. Last, I describe a dynamic aggregation phenomenon that occurs when the cells are seeded at high density in a nutrient-rich medium

The mortality rate is high for cancer patients and treatments are only efficient in a fraction of patients. To be able to cure more patients, new treatments need to be invented. Immunotherapy activates the immune system to fight against cancer and one treatment targets immune checkpoints. If more targets are found, more patients can be treated successfully. In this project, interactions between immune and cancer cells that drive tumour growth were investigated in an attempt to find new potential targets. This was achieved by creating a machine learning model that finds genes expressed in cells involved in tumour-driving interactions. Single-cell RNA sequencing and spatial transcriptomic data from breast cancer patients were utilised as well as single-cell RNA sequencing data from healthy patients. The tumour rate was based on the cumulative expression of G2/M genes. The G2/M related genes were excluded from the analysis since these were assumed to be cell cycle genes. The machine learning model was based on a supervised variational autoencoder architecture. By using this kind of architecture, it was possible to compress the input into a low dimensional space of genes, called a latent space, which was able to explain the tumour rate. Optuna hyperparameter optimizer framework was utilised to find the best combination of hyperparameters for the model. The model had a R2 score of 0.93, which indicated that the latent space was able to explain the growth rate 93% accurately. The latent space consisted of 20 variables. To find out which genes that were in this latent space, the correlation between each latent variable and each gene was calculated. The genes that were positively correlated or negatively correlated were assumed to be in the latent space and therefore involved in explaining tumour growth. Furthermore, the correlation between each latent variable and the growth rate was calculated. The up- and downregulated genes in each latent variable were kept and used for finding out the pathways for the different latent variables. Five of these latent variables were involved in immune responses and therefore these were further investigated. The genes in these five latent variables were mapped to cell types. One of these latent variables had upregulated immune response for positively correlated growth, indicating that immune cells were involved in promoting cancer progression. Another latent variable had downregulated immune response for negatively correlated growth. This indicated that if these genes would be upregulated instead, the tumour would be thriving. The genes found in these latent variables were analysed further. CD80, CSF1, CSF1R, IL26, IL7, IL34 and the protein NF-kappa-B were interesting finds and are known immune-modulators. These could possibly be used as markers for pro-tumour immunity. Furthermore, CSF1, CSF1R, IL26, IL34 and the protein NF-kappa-B could potentially be targeted in immunotherapy.