<p>Cellular structural mechanics play a key role in homeostasis by transducing mechanical signals to regulate gene expression and by providing adaptive structural stability for the cell. The alteration of nuclear mechanics in various laminopathies and in natural aging can damage these key functions. Arterial endothelial cells appear to be especially vulnerable due to the importance of shear force mechanotransduction to structure and gene regulation as is made evident by the prominent role of atherosclerosis in Hutchinson-Gilford progeria syndrome (HGPS) and in natural aging. Computational models of cellular mechanics may provide a useful tool for exploring the structural hypothesis of laminopathy at the intracellular level. This thesis explores this topic by introducing the biological background of cellular mechanics and lamin proteins in arterial endothelial cells, investigating disease states related to aberrant lamin proteins, and exploring computational models of the cell structure. It then presents a finite element model designed specifically for investigation of nuclear shear forces in arterial endothelial cells. Model results demonstrate that changes in nuclear material properties consistent with those observed in progerin-expressing cells may result in substantial increases in stress concentrations on the nuclear membrane. This supports the hypothesis that progerin disrupts homeostatic regulation of gene expression in response to hemodynamic shear by altering the mechanical properties of the nucleus.</p>

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<p dir="ltr">The actin cytoskeleton is a dynamic structure that regulates various important cellular processes, such as cell protrusion, migration, transport, and cell shape changes. Cells employ different actin architectures best suited for each of these functions. We have employed an agent-based model to illuminate how the actin cytoskeleton plays such functions in animal and plant cells, via dynamic interactions between molecular players.</p><p dir="ltr">Lamellipodia found in animal cells are two-dimensional actin protrusion formed on the leading edge of cells, playing an important role in sensing surrounding mechanical environments via focal adhesions. Various molecular players, architecture, and dynamics of the lamellipodia have been investigated extensively during recent decades. Nevertheless, it still remains elusive how each component in the lamellipodia mechanically interacts with each other to attain a stable, dynamic steady state characterized by a retrograde flow emerging in the branched actin network. Using the agent-based model, we investigated how the balance between different subcellular processes is achieved for the dynamic steady state. We simulated a branched network found in the lamellipodia, consisting of actin filament (F-actin), myosin motor, Arp2/3 complex, and actin crosslinking protein. We found the importance of a balance between F-actin assembly at the leading edge of cells and F-actin disassembly at the rear end of the lamellipodia. We also found that F-actin severing is crucial to allow for the proper disassembly of an actin bundle formed via network contraction induced by motor activity. In addition, it was found that various dynamic steady states can exist.</p><p dir="ltr">The actin cytoskeleton in plant cells plays a crucial role in intracellular transport and cytoplasmic streaming, and its structure is very different from the actin cytoskeleton in animal cells. The plant actin cytoskeleton is known to show distinct dynamic behaviors with homeostasis. We used the agent-based model to simulate the plant actin cytoskeleton with the consideration of the key governing mechanisms, including F-actin polymerization/depolymerization, different types of F-actin nucleation events, severing, and capping. We succeeded in reproducing experimental observations in terms of F-actin density, length, nucleation frequency, and rates of severing, polymerization, and depolymerization. We found that the removal of nucleators results in lower F-actin density in the network, which supports recent experimental findings.</p>

L'ingénierie tissulaire, qui consiste à remplacer un tissu lésé par un biosubstitut constitué de cellules réparatrices ensemencées dans une matrice de support biodégradable, possède un potentiel prometteur pour la réparation du Ligament Croisé Antérieur (LCA). Or, aucune solution opérationnelle n'a encore été proposée à ce jour, notamment au vu du nombre de domaines scientifiques impliqués. Dans ce travail, nous avons dressé un cahier des charges pour la définition de cette matrice en nous appuyant sur l'état de l'art. Une matrice de support tressée multicouche constituée de fibres de P(LL85/CL15) a été imaginée, puis les outils nécessaires à sa fabrication à l'échelle du laboratoire ont été mis en place. Nous avons ensuite développé des outils numériques spécifiques permettant la modélisation de sa géométrie et de son comportement biomécanique multi-échelles, qui ont été mis à profit afin d?optimiser les caractéristiques de la matrice compte tenu du cahier des charges établi. De plus, des caractérisations biologiques ont montré que la matrice était compatible avec la culture de cellules souches, et était susceptible d?accueillir la formation d'un néo-tissu. Par ailleurs, nous avons mis en place un bioréacteur spécifique permettant d'imposer à la matrice de support des cycles de traction-torsion sous environnement contrôlé. L'utilisation des informations locales issues de la modélisation biomécanique, afin d'interpréter ou d'optimiser les résultats de culture cellulaire sous sollicitations cycliques, constitue une perspective majeure du présent travail. Notre investigation permet en outre de penser qu'un nouveau biosubstitut pour le LCA pourrait prochainement être proposé / Tissue engineering, which consists in replacing an injured tissue with a biodegradable scaffold seeded with cells, has the potential to overcome the limitations associated with current reconstructions strategies of the Anterior Cruciate Ligament (ACL). However, no relevant solution has been proposed yet, especially due to the variety of scientific fields involved in this approach. In the current study, the key requirements for the design of a new scaffold have been listed from the current state of art. A scaffold based on P(LL85/CL15) fibers arranged into a multilayer braided structure has been proposed, and the tools needed to process this scaffold have been developed. Dedicated numerical tools have been proposed in order to predict the morphological and multiscale biomechanical behavior of the scaffold. These simulation tools have enabled to optimize the scaffold geometry in order to match the selected key requirements for ACL tissue engineering. Moreover, preliminary biological assessments have shown that the scaffold was suited for the culture of stem cells and for tissue formation. In addition, a dedicated bioreactor has been developed in order to prescribe tension-torsion cycles within a controlled environment. The use of local information issued from the biomechanical simulations open large perspectives as far as the optimization of culture conditions and the understanding of mechanisms that govern the formation of a ligamentous tissue are concerned. As a conclusion, the present study is likely to enable a new solution for ACL tissue engineering to emerge in the next years

Mesenchymal stem cells (MSCs) may benefit vascular cell-based therapies as smooth muscle or endothelial cell substitutes or through paracrine actions to repair, replace, or regenerate vascular tissue. Previous studies have demonstrated that MSCs can adopt traits of smooth muscle cells (SMCs) or endothelial cells (ECs), as well as secrete specific factors that tune signaling and material properties in the local environment. Few studies have investigated the cell signaling response of MSCs to mechanical forces present in the vasculature: specifically, shear stress due to blood flow and cyclic strain due to pulsatile blood flow. Thus, the central objective of this dissertation was to determine the signaling responses of MSCs to vascular-relevant applied physical forces, in comparison with that of differentiated vascular cells. Vascular-relevant mechanosignaling of MSCs was assessed through two comparisons: (1) MSC and SMC responses to applied cyclic strain and (2) MSC and EC responses to applied fluid shear stress. MSCs and SMCs were seeded on fibronectin-coated silicone and subjected in vitro to cyclic strain (10%, 1 Hz) or parallel static culture using a custom-built equibiaxial cyclic strain device. Gene expression analysis of 84 signal transduction molecules demonstrated both cell types respond with significant (p<0.05, n=3) fold-changes (|FC|≥ 1.5) within 24 hours of applied equibiaxial strain. Most strain-responsive genes identified were significantly strain-responsive in only one cell type. A signaling trio of Interleukin 8, Vascular cell adhesion molecule 1, and Heme oxygenase 1 was significantly altered in both MSCs and SMCs, suggesting cyclic strain regulates immune and inflammatory functions in both cell types. The response to shear stress of MSCs and ECs was compared using cells seeded on type I collagen or fibronectin and exposed to steady laminar shear stress (5 or 15 dyn/sq-cm) using a parallel plate shear chamber system. Gene expression was compared in MSCs and ECs for a panel of immune and inflammation-related markers. Expression of Cox-2 and Hmox-1 increased significantly (p<0.05, n≥3; |FC|≥1:5) in both cell types. Reduced shear stress-responses of Mcp-1, Pecam-1, and VE-Cad in MSCs relative to ECs suggests that MSCs promote less inflammation and immune activation in response to shear stress than ECs. Mechanosensitivity profiles for MSCs and differentiated vascular cells were broadened using whole genome microarrays. These high-throughput studies confirmed that (1) signaling profiles between sample groups vary significantly more (p<0.05, n=3) with cell type than applied force condition and (2) a subset of conserved mechanosensitive genes alter expression levels significantly and in the same direction fold-change in multiple cell types. Bioinformatics analysis of these conserved mechanoresponsive genes highlighted oxidative stress, cell cycle, and DNA replication as functions regulated by vascular-relevant mechanical cues. These studies demonstrate that MSCs partially reproduce differentiated vascular cell mechanosignaling, while simultaneously altering expression of genes not typically force-responsive in vascular cells. This work defines a role for conserved mechanosignals, based on genes whose expression in response to applied force alters significantly (p<0.05, n≥3) and by at least 1.5-fold change in multiple cell types and/or force types. Comparisons completed for this dissertation motivate future studies to track the functional impact of specific similar or unique MSC mechanoresponses. This work contributes to design of MSC-based vascular therapies and an understanding of stem and differentiated cell mechanobiology.

Dans le domaine des biomatériaux, cette thèse s’intéresse à l’influence de l’élasticité du substrat sur la division et la plasticité de la chromatine de cellules épithéliales. La létalité des cellules est corrélée aux faibles rigidités des substrats. Cependant, quelques cellules tumorales SW480, incluant celles portant des anomalies de ségrégation des chromosomes, progressent en mitose. Ces anomalies seraient à l’origine de réarrangements chromosomiques, sources de nombreuses mutations. Les substrats mous conduisent à la formation d’hétérochromatine tandis que les substrats très mous induisent la nécrose des cellules PtK2. Sur ces substrats, l’euchromatine est maintenue après inhibition de HDAC, permettant aux cellules de résister à la nécrose, indépendamment de la compétence transcriptionnelle du noyau. Ces cellules s’étalent à nouveau après transfert sur un substrat rigide. Ces résultats suggèrent 1) une voie de signalisation entrante initiée par le substrat conduisant à la nécrose via la formation d’hétérochromatine 2) une voie de signalisation sortante initiée par l’euchromatine permettant la survie cellulaire. / In the biomaterials field, this PhD work is about influence of substrate elasticity on cell division and chromatin plasticity of epithelial cells. Soft substrates cause massive death.However, some SW480 tumor cells, including those bearing chromosomal segregation abnormalities progress in mitosis. These abnormalities could result in more chromosomal rearrangements, increasing mutations. Soft substrates lead to heterochromatin remodelling and very soft substrates promote necrosis of PtK2 cells. On these substrates, euchromatin could be maintained after HDAC inhibition independently of the nuclear transcriptional competence.These cells spread again after tranfer on stiff substrates. These results suggest i) outside-insignalling cascade initiated at the soft substrate surface leading to heterochromatin remodelling and ultimately necrosis, ii) inside-out signaling cascade initiated from euchromatin allowing cell to overcome necrosis on soft substrate.

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<p>The subcutaneous injection has emerged as a common approach for self-administration of biotherapeutics due to the patient comfort and cost-effectiveness. However, the available knowledge about transport and absorption of these agents after subcutaneous injection is limited. Here we aim to find drug distribution in the tissue and lymphatic uptake after subcutaneous (SC) injection. In the first part of the study, a mathematical framework to study the subcutaneous drug delivery from injection to lymphatic uptake is presented. A three-dimensional poroelastic model is exploited to find the biomechanical response of the tissue by taking into account tissue deformation during the injection. The results show that including tissue deformability noticeably changes tissue poromechanical response due to the significant dependence of interstitial pressure on tissue deformation. Moreover, the importance of the amount of lymph fluid at the injection site and injection rate on the drug uptake to lymphatic capillaries is highlighted. Finally, the variability of lymphatic uptake due to uncertainty in parameters, including tissue poromechanical and lymphatic absorption parameters, is evaluated. It is found that interstitial pressure due to injection is the major contributing factor in short-term lymphatic absorption, while the amount of lymph fluid at the site of injection determines the long-term absorption of the drug. Finally, it is shown that the lymphatic uptake results are consistent with experimental data available in the literature.</p><p>In the second part, drug transport and distribution in different tissue layers are studied. A single-layer model of the tissue as a base study was first explored. During injection, the difference between the permeability of the solvent and solute results in a higher drug concentration proportional to the inverse of the permeability ratio. Then the effects of layered tissue properties with primary layers, including epidermis, dermis, subcutaneous, and muscle layers, on tissue biomechanical response to injection and drug transport are studied. The drug distributes mainly in the SQ layer due to its lower elastic moduli. Finally, the effect of secondary tissue elements like the deep fascia layer and the network of septa fibers inside the SQ tissue is investigated. The Voronoi algorithm is exploited to create random geometry of the septa network. It is shown how drug molecules accumulate around these tissue components as observed in experimental SC injection. Next, the effect of injection rate on drug concentration is studied. Higher injection rates slightly increase the drug concentration around septa fibers. Finally, it is demonstrated that the concentration-dependent viscosity increases the concentration of biotherapeutics in the direction of septa fibers.</p><p>In the third part of this thesis, a poro-hyperelastic model of the tissue is exploited to find the biomechanical response of the tissue together with a transport model based on an advection-diffusion equation in large-deformation poro-hyperelastic Media. The process of mAbs transport to the lymphatic system is explored. This process has two major parts. First, the initial phase, where mAbs are dispersed in the tissue as a result of momentum exerted by injection. This stage last for only a few minutes after the injection. Then there is the second stage, which can take tens of hours, and as a result, monoclonal antibodies (mAbs) molecules are transported from the subcutaneous layer towards initial lymphatics in the dermis to enter the lymphatic system. In third chapter, both stages are studied. The process of plume formation, interstitial pressure, and velocity development is explored. Then the effect of the injection device, injection site, and sensitivity of long-term lymphatic uptake due to variability in permeability, diffusivity, viscosity, and binding of mAbs are investigated. Then the results are used to find an equivalent lymphatic uptake coefficient that is widely used in pharmacokinetic (PK) models to study the absorption of mAbs. We show that the injection rate is the least, and the injection site is the most important parameter in the uptake of mAbs. Injection depth and mAbs dose also significantly alter lymphatic absorption. Finally, the computational model is validated against experimental studies available in the literature.</p>

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