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Generation of extracellular matrixprotein-based microcapsules for investigating single cells


Pashapour,  Sadaf
Cellular Biophysics, Max Planck Institute for Medical Research, Max Planck Society;

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Pashapour, S. (2022). Generation of extracellular matrixprotein-based microcapsules for investigating single cells. PhD Thesis, Ruprecht-Karls-Universität Heidelberg, Heidelberg.

Cite as: https://hdl.handle.net/21.11116/0000-0009-775F-8
Interactions between cells and the extracellular matrix (ECM) activate multiple signaling pathways that initiate, drive and regulate nearly all motions of cells in their native environment. Cell-ECM interactions are also emphasizing the importance of research that aims to better understand such interactions. Consequently, engineering 3D ECM systems for controlled manipulation of cells in vitro has become an important strategy, particularly in medical applications. These systems will contribute to understanding the mechanisms underlying the ability of cells to perform different tasks as a response to environmental information. In this PhD thesis, I have established two novel droplet-based microfluidic approaches for the controlled assembly of; (1) cell-laden ECM-based protein microcapsules; and (2) ECM-coated crescent hydrogel-based microparticles. Towards the production of ECM-based microcapsules, water-in-oil emulsion droplets consisting of negatively or positively charged block-copolymer surfactants are used as a template for the charge-mediated formation of an either pure laminin-, laminin/collagen mixed- or fibronectin-based continuous layer on the inner droplet periphery. Following the protein layer formation, different microfluidic technologies are implemented to encapsulate cells and under the appropriate ionic conditions for the controlled polymerization of the protein layer. Sequential release of the assembled cell-laden ECM based microcapsules from the surfactant-stabilized droplets into a physiological environment allows for analysis of cell-ECM interactions on the single-cell level. The second technology, invented within the scope of this thesis, is the application of ECM-coated crescent PEGDA microparticles for the analysis of cell behavior on curved substrates. By making use of an aqueous two-phase microfluidic system, it was possible to establish PEGDA crescent microparticles with a layer of ECM proteins coating the bucket. Towards this end, ECM proteins are dissolved with dextran molecules and are encapsulated with PEGDA into water-in-oil droplets. Due to a phase separation between PEGDA and dextran the characteristic crescent shape is established. Upon polymerization of PEGDA it becomes feasible to release the particles and wash away the dextran phase, which generates a remaining protein layer on the inner bucket. By tailoring the biochemical properties of both systems, we are able to produce a wide variety of ECM-based microcapsules that are tunable in terms of protein composition and cellular encapsulation. Ultimately, this technology will be used for investigating cell-ECM interactions in various environments and on a variety of substrate geometries.