Skip to Content
MilliporeSigma
  • Transgenic zebrafish modeling low-molecular-weight proteinuria and lysosomal storage diseases.

Transgenic zebrafish modeling low-molecular-weight proteinuria and lysosomal storage diseases.

Kidney international (2020-02-18)
Zhiyong Chen, Alessandro Luciani, José María Mateos, Gery Barmettler, Rachel H Giles, Stephan C F Neuhauss, Olivier Devuyst
ABSTRACT

Epithelial cells lining the proximal tubule of the kidney reabsorb and metabolize most of the filtered low-molecular-weight proteins through receptor-mediated endocytosis and lysosomal processing. Congenital and acquired dysfunctions of the proximal tubule are consistently reflected by the inappropriate loss of solutes including low-molecular-weight proteins in the urine. The zebrafish pronephros shares individual functional segments with the human nephron, including lrp2a/megalin-dependent endocytic transport processes of the proximal tubule. Although the zebrafish has been used as a model organism for toxicological studies and drug discovery, there is no available assay that allows large-scale assessment of proximal tubule function in larval or adult stages. Here we establish a transgenic Tg(lfabp::½vdbp-mCherry) zebrafish line expressing in the liver the N-terminal region of vitamin D-binding protein coupled to the acid-insensitive, red monomeric fluorescent protein mCherry (½vdbp-mCherry). This low-molecular-weight protein construct is secreted into the bloodstream, filtered through the glomerulus, reabsorbed by receptor-mediated endocytosis and processed in the lysosomes of proximal tubule cells of the fish. Thus, our proof-of-concept studies using zebrafish larvae knockout for lrp2a and clcn7 or exposed to known nephrotoxins (gentamicin and cisplatin) demonstrate that this transgenic line is useful to monitor low-molecular-weight proteinuria and lysosomal processing. This represents a powerful new model organism for drug screening and studies of nephrotoxicity.

MATERIALS
Product Number
Brand
Product Description

Supelco
Ethyl 3-aminobenzoate methanesulfonate salt, analytical standard
Sigma-Aldrich
Ethyl 3-aminobenzoate methanesulfonate, 98%
HiTrap® Chelating High Performance, Cytiva 17-0408-01, pack of 5 × 1 mL
Sigma-Aldrich
Monoclonal Anti-β-Actin antibody produced in mouse, clone AC-15, ascites fluid
Sigma-Aldrich
Gentamicin sulfate salt, powder, BioReagent, suitable for cell culture
Sigma-Aldrich
Taurine, ≥99%
Cisplatin, European Pharmacopoeia (EP) Reference Standard
Sigma-Aldrich
RIPA Buffer
Sigma-Aldrich
Bafilomycin A1 from Streptomyces griseus, ≥90% (HPLC)