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Since the emergence of spheroid/organoid technology, a significant amount of effort has gone toward improving culture techniques to better recapitulate the native organ and preserve cell phenotype. While hydrogels of various composi-tions offer interesting opportunities to model human physiology, there are still un-optimized factors to consider in continuing to move the field forward. The goals of this PhD dissertation are (1) to engineer a defined microenvironment that can support the malignant state, function, and phenotype of liver cancer cells (2) to justify the developed liver model’s application for in vitro drug screening that is predictive of patient responses. The focus is on key microenvironmental parame-ters affecting the physiological function of cells, mainly oxygenation and extracel-lular matrix ligands. To this end, monodispersed and cell-sized chitosan-based microparticles conjugated with fluorine molecules were generated using a custom microfluidic device. Next, they were co-cultured with commonly used human hepatocellular carcinoma cell line (HepG2) and hepatic stellate cells (HSCs) to enhance oxygen levels in aggregate culture. In the next step, the surface of mi-croparticles was decorated with different ligands derived from the native liver ex-tracellular matrix to understand how different proteins modulate cellular behavior. Finally, two tumor models including immortalized and patient-derived liver sphe-roids cultured with microparticles were applied for toxicity assessments in vitro and the mechanism underlying cell apoptosis was studied. Together, this disserta-tion advances efforts to create more physiologically relevant organ models and allows a well-defined condition and phenotypic cell signaling to improve the rele-vance of spheroid and organoid models. |
