ESPN 51th Annual Meeting

ESPN 2018


 
Cysteamine improves proximal tubular reabsorption of low molecular weight compounds in cystinotic zebrafish, but has no effect on the defective megalin expression
MOHAMED ELMONEM 1 SANTE P. BERLINGERIO 1 JOSEPH MORGAN 2 LADAN KHODAPARAST 1 LALEH KHODAPARAST 1 MARTIN LOWE 2 LAMBERTUS VAN DEN HEUVEL 1 ELENA LEVTCHENKO 1

1- KU LEUVEN
2- UNIVERSITY OF MANCHESTER
 
Introduction:

Nephropathic cystinosis is an autosomal recessive lysosomal storage disorder characterized by cystine accumulation and early renal damage particularly at the level of the proximal tubules. We have developed and validated a mutant zebrafish model for the disease that shows significant reduction in the proximal tubular reabsorption capacity, especially for LMW compounds. Moreover, we have shown that in the zebrafish the expression of the multi-ligand receptor megalin is defective, as it is mainly trapped in subapical and cytoplasmic vacuoles instead of being evenly distributed at the brush border of the tubular lumen, where it should perform its function. We aimed to investigate the effect of cysteamine on this process in the zebrafish model.

Material and methods:

At three days post fertilization (3 dpf), we injected wild-type and cystinosis zebrafish larvae (treated or not with 100 µM of cysteamine), with fluorescently labelled LMW dextran (4.4 kDa). This was followed by cryosectioning to evaluate the fluorescence intensity in the proximal tubules. We also evaluated the protein expression of megalin, early endosomal antigen-1 (EEA1) and Rab11 at the level of the proximal tubules in each condition at 5 dpf using immunofluorometric staining of frozen sections. 

Results:

At 100 µM, cysteamine significantly increased the amount of fluorescent dextran reabsorbed in the proximal tubules of cystinosis zebrafish larvae, compared to the untreated cystinosis larvae. However, the defective pattern of megalin expression was not improved after cysteamine treatment. Furthermore, the protein expression of EEA1 (a tethering protein of early endosomes) and the small GTPase Rab11 associated with recycling endosome trafficking to the plasma membrane was significantly decreased in the proximal tubules of the cystinosis zebrafish compared to the wild-type. Cysteamine treatment did not restore the protein expression of either. 

Conclusions:

 Cysteamine therapy partially improves proximal tubular endocytosis in cystinotic zebrafish larvae but not through the megalin-receptor-mediated endocytic machinery.