New Mexico Geological Society Annual Spring Meeting — Abstracts

Gadolinium-based contrast agents (GBCAs) enhance magnetic resonance imaging (MRI) examinations in clinical care. Long-term tissue retention of elemental gadolinium, nephrogenic systemic fibrosis, and acute kidney injury are directly linked to GBCA exposures. In addition to being the primary organ for GBCA excretion, the kidneys also serve as long-term reservoirs for these agents, increasing the risk of nephrotoxicity. Therefore, it is crucial to define the cellular targets of gadolinium-based contrast agents. This study aimed to determine the role of cytoplasmic organelles in the pathogenesis of GBCA-induced renal injury using an in vitro model of renal proximal tubular epithelial cells.

Mouse renal proximal tubular epithelial cells (MRPTEpiCs) were seeded at a density of 10 × 10³ cells/cm² and allowed to reach ~70% confluence prior to GBCA exposure. Cells were exposed to 2mM concentrations of commercially available GBCAs from the two general classes, linear (Omniscan) or macrocyclic (Dotarem), for 1 – 48 hrs. Protease inhibitor cocktail (P1860-1mL, Sigma) or Cathepsin B inhibitor, CA-074Me (S7420, Selleck Chem) were incubated simultaneously with the indicated contrast agents. Untreated cells at each time point served as controls. Inductively coupled plasma mass spectroscopy (ICP-MS) was used to measure intracellular gadolinium levels. Transmission electron and confocal microscopy, fluorescence-based assays, and western blotting were used to monitor and quantify lysosomal and mitochondrial morphology and function.

The cellular uptake of GBCAs has not been thoroughly investigated. ICP-MS bulk analysis of MRPTEpiCs exposed to either linear or macrocyclic, demonstrates that these cells accumulate gadolinium in a time-dependent manner, indicating a cellular uptake process. Exposures to linear and macrocyclic GBCAs have induced lysosomal damage, characterized by lysosomal accumulation and enlargement, followed by lysosomal membrane permeabilization and the release of lysosomal contents into the cytosol. Disturbance of mitochondrial dynamics and loss of cell viability occur downstream of lysosome damage.

In this study, we investigate the interplay between critical cytosolic organelles in GBCA-induced renal tubular cell injury. Functional organelles, such as mitochondria and lysosomes, are crucial for maintaining cellular homeostasis. We demonstrate that the accumulation of these contrast agents in the endolysosomal compartment causes lysosomal damage and the release of lysosomal contents into the cytosol. This protease leakage initiates contrast agent-induced mitochondrial damage, impairing cellular function. Here, we reveal the unexpected lysosomotropic behavior of gadolinium-based contrast agents and how interorganelle dyshomeostasis contributes to GBCA-mediated renal injury.