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2024/06/20

Creation and application of a new high-throughput individual zebrafish nephrotoxicity screening system

Friday, July 5, 2024, 15:24-15:36
The 51st Annual Meeting of the Japanese Society of Toxicology, Fukuoka International Congress Center, Fukuoka, Japan
We will report the following at the 51st Annual Meeting of the Japanese Society of Toxicology, Fukuoka International Conference Hall 7, Fukuoka, Japan.

Zebrafish have been extensively used for nephrotoxicity screening worldwide. This is due to its 84% homology to the human pathological genome, which suggests similarity to humans in toxicity mechanisms, and its whole-body transparency, which allows direct analysis of renal morphology and renal function. In addition, zebrafish is the most suitable vertebrate for in vivo nephrotoxicity high-throughput screening due to its high fecundity and rapid organogenesis. Initially, zebrafish nephrotoxicity screening used morphological observations and blood tests such as creatinine and urea nitrogen to assess nephrotoxicity. However, these methods were not sensitive enough for early diagnosis. Therefore, we decided to apply intravenous fluorescent labeled dextran of one molecular weight for nephrotoxic proteinuria quantification. In this case, several challenges remained due to the fact that it is not an endogenous protein and has only one molecular weight. Therefore, the determination of nephrotoxic proteinuria by transgenic zebrafish of endogenous plasma proteins (1/2vdbp-mCherry:50kDa, NL-D3:35.5kDa) was reported. However, in these cases, the molecular weight was still one of a kind, and the inability to discriminate between glomerular and tubular functions in nephrotoxicity remained an issue. Therefore, in 2013 (ACS Chem Neurosci. 4:1183-93), we independently developed a novel fluorescent dye ZMB741 that rapidly binds to all molecular weight plasma proteins in vivo when added to zebrafish breeding water, and developed a non-invasive We have established a protocol for in vivo quantification of nephrotoxic proteinuria in individual zebrafish. As a result, we report the creation of an in vivo high-throughput nephrotoxicity screening system based on a completely new principle that can discriminate between glomerular and urinary repopulation disorders of nephrotoxicity.

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