Abstract: FR-PO555
Modeling Kidney Proximal Tubular Crystallopathy In Vitro
Session Information
- Fluid, Electrolyte, and Acid-Base Disorders: Basic
October 25, 2024 | Location: Exhibit Hall, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Fluid, Electrolytes, and Acid-Base Disorders
- 1101 Fluid, Electrolyte, and Acid-Base Disorders: Basic
Authors
- Masereeuw, Rosalinde, Universiteit Utrecht, Utrecht, Utrecht, Netherlands
- Vonk, Mara, Universiteit Utrecht, Utrecht, Utrecht, Netherlands
- Klein, Chaja, Universiteit Utrecht, Utrecht, Utrecht, Netherlands
- Janssen, Manoe J., Universiteit Utrecht, Utrecht, Utrecht, Netherlands
- Barnes, Devon, Universiteit Utrecht, Utrecht, Utrecht, Netherlands
Background
Kidney crystallopathy can be characterized by the presence of crystals within renal tubules, leading to obstruction, cytotoxic effects, and inflammatory responses. Here, a novel proximal tubule cell model was developed to investigate the mechanisms underlying toxicant-induced crystal formation and renal inflammation.
Methods
Conditionally immortalized proximal tubule epithelial cells (ciPTECs) were used to investigate the role of uric acid, a known crystal-forming metabolite, and the influence of pH on crystal formation. A series of in vitro assays were used to assess cytotoxicity, oxidative stress, mitochondrial function, inflammation as well as the expression of NLRP3 inflammasome related markers on the protein level.
Results
Exposing ciPTECs to uric acid decreased cell viability, which was more pronounced under reduced pH conditions with a significant reduction starting at 200 μg/ml at pH 5.5 vs 800 μg/ml at pH 7.4. Various shapes of uric acid crystals were formed (Figure 1). Reactive oxygen species production was increased at 200 μg/ml at all pH levels tested (viz. 5.5, 6.0 and 7.4). Furthermore, uric acid induced the expression of the inflammasome-related markers caspase-1, ASC, TNFα at the mRNA level, and IL-1β at the protein level. In line with cell viability, these effects were most pronounced with higher uric acid concentrations and at lower pH levels.
Conclusion
A novel model was established simulating kidney crystallopathy and its impact on inflammasome components, caspase activity, cytokine production, and the NF-κβ pathway. This model will be used to study crystallopathy in more detail to provide an adverse outcome pathway for crystal-forming nephrotoxicants.
This project received funding from the European Union’s Horizon 2020 research and innovation programme (GA No 963845, ONTOX).
Figure 1. Uric acid crystals formed after 72h incubation with A) 800 μg/ml at pH 5.5 and B) 600 μg/ml at pH 5. 10x magnification.
Funding
- Government Support – Non-U.S.