This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
A new preprint design is coming!
We have improved readability, clarity, accessibility, and opportunities for engagement.
Research Article
Tom J.J. Schirris
Radboud University Medical Center
Corresponding Author
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Acute kidney injury accounts for 20% of all hospitalized adults, and 14 to 26% is drug-induced, emphasizing the importance of proper nephrotoxicity assessment. The ‘gold standard’ MTT assay is widely used to measure cell viability, but depends on cellular metabolic activity. Consequently, MTT may not be most optimal to assess cytotoxicity, as nephrotoxicity often involves mitochondrial dysfunction. We compared MTT with a direct cell death assay based on a compromised plasma membrane permeability. Mature conditionally immortalized proximal tubule epithelial cells were dose- (0.1-1,000 µM) and time- (0.5, 1, 2, 4, 8 and 24 hours) dependently exposed to a selection of prototypic nephrotoxicants. Dose-dependent reductions in cellular metabolic activity were stronger compared to declines in fluorescence-based cell death, most prominently for cisplatin (1.6 ± 2.0% and 68 ± 4% (mean ± SEM), respectively) and chloroacetaldehyde (2.13 ± 0.05% and 61.0 ± 0.8%). Similar, but more pronounced time-dependent effects were observed, particularly for sanguinarine. We show that assessing cellular metabolic activity by MTT provides a composite readout of cellular metabolic activity and cell death. A nuclear staining approach is preferable when assessing nephrotoxicity of metabolically active compounds. We recommend both assays during drug development to discriminate between metabolically active versus non-active compounds.
Keywords
drug-induced acute kidney injury, cellular metabolic activity, cell viability, metabolic toxicity
Figure 1
Figure 2
Figure 3
Competing interest reported. F.G.M.R. is co-inventor on patent EP2010/066792 ‘Novel conditionally immortalized human proximal tubule cell line expressing functional influx and efflux transporters’ assigned to Radboud University Medical Center and has conflict of interest through commercialization of ciPTEC models via Cell4Pharma. J.A.M.S. is the founding CEO of Khondrion BV, a Radboud University Medical Center spin-out company founded by J.A.M.S. Other authors do not have any conflict of interest.
Loading...
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
View the published article at Frontiers in Toxicology.
Version 1
Posted 12 Apr, 2021
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Learn more about our company.
A new preprint design is coming!
We have improved readability, clarity, accessibility, and opportunities for engagement.
This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research Article
Tom J.J. Schirris
Radboud University Medical Center
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
View the published article at Frontiers in Toxicology.
Version 1
Posted 12 Apr, 2021
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
View the published article at Frontiers in Toxicology.
Acute kidney injury accounts for 20% of all hospitalized adults, and 14 to 26% is drug-induced, emphasizing the importance of proper nephrotoxicity assessment. The ‘gold standard’ MTT assay is widely used to measure cell viability, but depends on cellular metabolic activity. Consequently, MTT may not be most optimal to assess cytotoxicity, as nephrotoxicity often involves mitochondrial dysfunction. We compared MTT with a direct cell death assay based on a compromised plasma membrane permeability. Mature conditionally immortalized proximal tubule epithelial cells were dose- (0.1-1,000 µM) and time- (0.5, 1, 2, 4, 8 and 24 hours) dependently exposed to a selection of prototypic nephrotoxicants. Dose-dependent reductions in cellular metabolic activity were stronger compared to declines in fluorescence-based cell death, most prominently for cisplatin (1.6 ± 2.0% and 68 ± 4% (mean ± SEM), respectively) and chloroacetaldehyde (2.13 ± 0.05% and 61.0 ± 0.8%). Similar, but more pronounced time-dependent effects were observed, particularly for sanguinarine. We show that assessing cellular metabolic activity by MTT provides a composite readout of cellular metabolic activity and cell death. A nuclear staining approach is preferable when assessing nephrotoxicity of metabolically active compounds. We recommend both assays during drug development to discriminate between metabolically active versus non-active compounds.
Figure 1
Figure 2
Figure 3
Competing interest reported. F.G.M.R. is co-inventor on patent EP2010/066792 ‘Novel conditionally immortalized human proximal tubule cell line expressing functional influx and efflux transporters’ assigned to Radboud University Medical Center and has conflict of interest through commercialization of ciPTEC models via Cell4Pharma. J.A.M.S. is the founding CEO of Khondrion BV, a Radboud University Medical Center spin-out company founded by J.A.M.S. Other authors do not have any conflict of interest.
Loading...