Nanoparticle toxicity assessment using an in vitro 3-D kidney organoid culture model.

Nanocarriers and nanoparticles remain an intense pharmaceutical and medical imaging technology interest. Their entry into clinical use is hampered by the lack of reliable in vitro models that accurately predict in vivo toxicity. This study evaluates a 3-D kidney organoid proximal tubule culture to assess in vitro toxicity of the hydroxylated generation-5 PAMAM dendrimer (G5-OH) compared to previously published preclinical in vivo rodent nephrotoxicity data. 3-D kidney proximal tubule cultures were created using isolated murine proximal tubule fractions suspended in a biomedical grade hyaluronic acid-based hydrogel. Toxicity in these cultures to neutral G5-OH dendrimer nanoparticles and gold nanoparticles in vitro was assessed using clinical biomarker generation. Neutral PAMAM nanoparticle dendrimers elicit in vivo-relevant kidney biomarkers and cell viability in a 3-D kidney organoid culture that closely reflect toxicity markers reported in vivo in rodent nephrotoxicity models exposed to this same nanoparticle.

Comparing predictive drug nephrotoxicity biomarkers in kidney 3-D primary organoid culture and immortalized cell lines.

The cellular microenvironment is recognized to play a key role in stabilizing cell differentiation states and phenotypes in culture. This study addresses the hypothesis that preservation of in vivo-like tissue architecture in vitro produces a cell culture more capable of responding to environmental stimuli with clinically relevant toxicity biomarkers. This was achieved using kidney proximal tubules in three-dimensional organoid hydrogel culture, with comparisons to conventional monolayer kidney cell cultures on plastic. Kidney proximal tubule cultures and two immortalized kidney cell line monolayer cultures exposed to known nephrotoxic drugs were evaluated for inflammatory cytokines, nephrotoxicity-associated genes, Kim-1 protein, cytochrome enzymes, and characteristic cellular enzyme shedding. Significant similarities are shown for these traditional biomarkers of kidney toxicity between in vivo and 3-D organoid endpoints of drug toxicity, and significantly, a consistent lack of clinically relevant endpoints produced by traditional 2-D kidney cell cultures. These findings impact both in vitro bioreactor-based kidney functional and regenerative medicine models, as well as high-throughput cell-based drug screening validations.

A 3-D organoid kidney culture model engineered for high-throughput nephrotoxicity assays.

Cell-cell and cell-matrix interactions control cell phenotypes and functions in vivo. Maintaining these interactions in vitro is essential to both produce and retain cultured cell fidelity to normal phenotype and function in the context of drug efficacy and toxicity screening. Two-dimensional (2-D) cultures on culture plastics rarely recapitulate any of these desired conditions. Three dimensional (3-D) culture systems provide a critical junction between traditional, yet often irrelevant, in vitro cell cultures and more accurate, yet costly, in vivo models. This study describes development of an organoid-derived 3-D culture of kidney proximal tubules (PTs) that maintains native cellular interactions in tissue context, regulating phenotypic stability of primary cells in vitro for up to 6 weeks. Furthermore, unlike immortalized cells on plastic, these 3-D organoid kidney cultures provide a more physiologically-relevant response to nephrotoxic agent exposure, with production of toxicity biomarkers found in vivo. This biomimetic primary kidney model has broad applicability to high-throughput drug and biomarker nephrotoxicity screening, as well as more mechanistic drug toxicology, pharmacology, and metabolism studies.