||Experimental models that faithfully reflect both normal physiological and pathological conditions represent an essential cornerstone of both the basic and applied sciences. However, the degrees to which these models represent actual in vivo conditions and environments greatly affect their ability to accurately produce clinically relevant responses to stimuli such as drug exposures. This dissertation introduces an in vitro kidney proximal tubule model that aims to retain tissue architecture, cell organization, cell-matrix and cell-cell interactions, and physiological cellular responses in long-term culture, and assess its ability to preserve differentially and functionally stable primary cells during exposure to drugs known to be nephrotoxic. These features are achieved by encapsulating suspended harvested viable proximal tubule fragments containing their epithelial cells in a three-dimensional hyaluronic acid (HA) hydrogel matrix. Proximal tubule fragment (i.e., "organoid") encapsulation, and not their isolated cells suspensions, in the hydrogel endows the model with the capability to retain native cellular organization, extracellular matrix, and intra- and extra-cellular interactions, while limiting non-native cell-polymer interfaces. Data collected for these cultures indicate that proximal tubule epithelial cells sustain cellular gluconeogenic potential, differential markers, and functional ligands. Moreover, these kidney cells in vitro were capable of responding to exogenous agents (i.e., drug introductions) with induction of in vivo-relevant gene and protein biomarkers as well as accurate cytokine, cytochrome, and iv metabolite expression patterns. The utility of the model for assessing potential nephrotoxic agents was further validated by comparison to the "gold standard" of current in vitro toxicology: immortalized cell monolayers grown on plastic. Results demonstrate significant improvements in cell phenotypic fidelity for 3D organoid cultures over cells supported via standard in vitro culture techniques that produce most clinically measured biomarkers. The obtained results indicate that preservation of the cellular microenvironment is key to not only sustaining functional proximal tubule cells in vitro but also in preserving their natural responses to the same nephrotoxic indicators as expressed in animal toxicity models. The proposed 3D in vitro culture model bridges the gap between overly simplistic cellular and costly in vivo assessment, and opens possibilities for direct, reliable and predictive comparisons between in vitro and clinical data.