Investigations of geometric cues on cell physiology in vitro

Cells in the human body show drastic differences in shape and morphology depending on the tissue that they are embedded in, and pathological tissue is often accompanied by changes in cell architecture. However, the question of whether changes in cell architecture play a regulatory role in physiology and disease is still mostly unexplored. As the cell architecture is influenced by the cell micro environment, such as the surrounding ECM, it might play a central role as a regulator and mediator of physical, or even chemical, environmental cues in vivo. As it is not possible to study the effect of cell architecture in vivo, various in vitro bioengineering systems based on microfabrication were developed in this thesis. The use of cell micropatterns allowed precise engineering of cell geometry, which was found to result in specific cell cytoskeletal arrangements and changes in cell contractility. It was found that those changes in contractility mediate cell geometry-dependent lineage commitment of mesenchymal stem cells. Development of several new analytical techniques and subsequent application to cell micropattern systems resulted in many new and unexpected cell geometry-dependent effects. For the first time, a link between lipid raft formation and cellcell geometry was observed. Additionally, plasma membrane curvature as a new cell characteristic is described and reported to be cell geometry-dependent in this thesis. Furthermore, a new mechanism involving lipid raft dependent Akt signalling was found to mediate cell geometry-dependent stem cell lineage commitment independent of PI3K activity in mesenchymal stem cells. Considering the involvement of Akt signalling in a wide variety of cell physiological events, and in many challenging diseases such as type-2 diabetes and cancer, the evidence of a link between cell geometry and Akt signalling could have broader implications and warrants further investigation in other cell systems. Moreover, depending on the soluble factor environment, cell geometry was observed to have different, seemingly antagonistic effects, on cell differentiation. In the absence of soluble differentiation cues, β-catenin was found to mediate cell differentiation in a most likely Wnt independent mechanism. Furthermore, by developing a novel system based on decellularized ECM micropatterns, it was reported that cell architecture also changes ECM structure by cell shape-dependent secretion of ECM proteins, suggesting a bi-directional regulatory mechanism between cell shape and ECM. It was also found that downstream effects caused by other mechanical cues in the cell microenvironment, such as matrix elasticity and topography, are indirect effects mediated by changes in cell geometry. Therefore, cell architecture seems to have an important role in mediating cell physiological effects of ECM and might be a main component that transduces effects of mechanobiological cues into specific downstream behaviour. The work in this thesis suggests a possible key role of cell geometry within the cell microenvironment, and is highly relevant for improved targeted therapies, especially in cancer, the design of tissue engineering scaffolds and in vitro drug screening systems.Open Acces

Triggerable tough hydrogels for gastric resident dosage forms

Systems capable of residing for prolonged periods of time in the gastric cavity have transformed our ability to diagnose and treat patients. Gastric resident systems for drug delivery, ideally need to be: ingestible, be able to change shape or swell to ensure prolonged gastric residence, have the mechanical integrity to withstand the forces associated with gastrointestinal motility, be triggerable to address any side effects, and be drug loadable and release drug over a prolonged period of time. Materials that have been primarily utilized for these applications have been largely restricted to thermoplastics and thermosets. Here we describe a novel set of materials, triggerable tough hydrogels, meeting all these requirement, supported by evaluation in a large animal model and ultimately demonstrate the potential of triggerable tough hydrogels to serve as prolonged gastric resident drug depots. Triggerable tough hydrogels may be applied in myriad of applications, including bariatric interventions, drug delivery, and tissue engineering.Bill & Melinda Gates Foundation (Grant OPP1096734)Bill & Melinda Gates Foundation (Grant OPP1139927)National Institutes of Health (U.S.) (Grant EB000244

High resolution Raman spectroscopy mapping of stem cell micropatterns

We report on the use of high resolution Raman spectroscopy mapping combined with a micro-engineered stem cell platform.</p

Sparse feature selection methods identify unexpected global cellular response to strontium-containing materials

Despite the increasing sophistication of biomaterials design and functional characterization studies, little is known regarding cells’ global response to biomaterials. Here, we combined nontargeted holistic biological and physical science techniques to evaluate how simple strontium ion incorporation within the well-described biomaterial 45S5 bioactive glass (BG) influences the global response of human mesenchymal stem cells. Our objective analyses of whole gene-expression profiles, confirmed by standard molecular biology techniques, revealed that strontium-substituted BG up-regulated the isoprenoid pathway, suggesting an influence on both sterol metabolite synthesis and protein prenylation processes. This up-regulation was accompanied by increases in cellular and membrane cholesterol and lipid raft contents as determined by Raman spectroscopy mapping and total internal reflection fluorescence microscopy analyses and by an increase in cellular content of phosphorylated myosin II light chain. Our unexpected findings of this strong metabolic pathway regulation as a response to biomaterial composition highlight the benefits of discovery-driven nonreductionist approaches to gain a deeper understanding of global cell–material interactions and suggest alternative research routes for evaluating biomaterials to improve their design

Cell-geometry-dependent changes in plasma membrane order direct stem cell signalling and fate

Cell size and shape affect cellular processes such as cell survival, growth and differentiation1,2,3,4, thus establishing cell geometry as a fundamental regulator of cell physiology. The contributions of the cytoskeleton, specifically actomyosin tension, to these effects have been described, but the exact biophysical mechanisms that translate changes in cell geometry to changes in cell behaviour remain mostly unresolved. Using a variety of innovative materials techniques, we demonstrate that the nanostructure and lipid assembly within the cell plasma membrane are regulated by cell geometry in a ligand-independent manner. These biophysical changes trigger signalling events involving the serine/threonine kinase Akt/protein kinase B (PKB) that direct cell-geometry-dependent mesenchymal stem cell differentiation. Our study defines a central regulatory role by plasma membrane ordered lipid raft microdomains in modulating stem cell differentiation with potential translational applications