Assessing the Nephrotoxicity of Benzalkonium Chlorides in vitro and in vivo

Abstract

Benzalkonium chlorides (BACs) are widely used antimicrobial compounds found across a variety of settings, including consumer products, agriculture, the food processing industry, and clinical settings. Interest in BACs has intensified in recent years, driven by increased regulatory scrutiny by the U.S. Federal Food and Drug Administration (FDA) regarding their safety. The surge in disinfectant use during COVID-19 resulted in elevated concentrations of BACs in human serum, raising concerns about their potential health impacts. BACs exhibit broad tissue distribution, but the kidney has been identified as a primary site of accumulation, particularly for long-chain BACs. The kidney is especially vulnerable to environmental toxicants due to its high perfusion rate and its physiological role in filtering and concentrating xenobiotics for excretion. In addition, renal tissue contains lower levels of xenobiotic-metabolizing enzymes than the liver, limiting its capacity to detoxify xenobiotics effectively. Therefore, we hypothesize that the low BAC-metabolizing capacity contributes to the extent of BAC buildup in the kidney and consequently, to BAC-induced kidney injury. In this dissertation, I first evaluated the metabolic capacity for BACs in 2D-cultured human proximal tubule epithelial cells (PTECs). I subsequently investigated nephrotoxicity in a human-relevant model using the novel organ-on-a-chip microphysiological system (MPS). I further examined altered biochemical pathways in PTECs cultured in the MPS using RNA sequencing under low-dose, chronic exposure to BACs in the kidney MPS to better reflect real-world exposure. Next, I assessed the spatial accumulation of BACs in the kidneys of mice exposed to BACs using matrix-assisted laser desorption ionization imaging mass spectrometry (MALDI-IMS), enabling the identification of kidney regions that are most susceptible to BAC accumulation and toxicity. I also characterized the global lipidome changes in mouse kidneys, identifying potential lipid biomarkers associated with acute kidney injury (AKI). Finally, I performed a comprehensive transcriptomic analysis of BAC-exposed mice kidneys and 2D-cultured human PTECs, aiming to identify biochemical pathways that are consistent with those identified in MPS. I conclude the dissertation by discussing the overall findings and implications of this work in human health, providing insight into future research directions that would further elucidate the mechanisms of BAC toxicity in the kidney. Collectively, this work provides a comprehensive, multifaceted evaluation of BAC-induced kidney toxicity, integrating in vitro, in vivo, and systems-level approaches. These findings emphasize the potential human health risks associated with chronic, low-level exposure to BACs and highlight the need for further investigation into environmental toxicant accumulation and its long-term consequences on renal health.