Chapter 2 34 of cysteamine (0.1 mM) confirming the importance of cystine depletion in the correction of cystinotic phenotype. Another key finding of our study is that the ctns‒/‒ zebrafish model exhibits the early renal phenotype. Resorption of the 4-kDa dextran, which readily passes the GFB, was virtually absent in the ctns‒/‒ larvae, indicating perturbed proximal tubular reabsorption. The 70-kDa tracer, which does not readily pass the GFB, was lost more rapidly in the ctns‒/‒ larvae than in the wt. Moreover, the loss of glomerular permeability was probably underestimated in the tubular reabsorption analysis, as the 4-kDa experiment has shown that tubular reabsorption capacity was reduced. This phenotype closely mimics the human disease which usually manifests with both tubular and glomerular impairment during infancy and childhood, at a much earlier age than in its mouse counterpart. Although the functional glomerular defect in the ctns-/- larvae with the HMW dextran is evident at this early stage, the data on partial podocyte effacement and minimal loss of inulin clearance were not that impressive. Thus, these data should be further evaluated at later time points which may be more representative for the podocyte damage. On the other hand, unlike in mammals, podocytes can regenerate in the adult zebrafish47 which can mitigate renal disease progression. Interestingly, the presence of the renal phenotype in the Ctns‒/‒ mouse depended on its genetic background. While the first cystinosis mice generated on a FVB/N background did not develop any signs of renal pathology44,Ctns‒/‒ mice on a pure C57BL/6 background showed defective proximal tubular reabsorption starting from 2 months of age corresponding to late adolescence in humans; however, evidence of decreased GFR was only detectable in adult mice at 10 months of age10 and no podocyte damage was present11. This points to compensatory mechanisms in the FVB/N mice which are not present in C57BL/6 mouse, Danio rerio or in humans. A concise comparison of cystinosis in humans, mice and zebrafish is presented in Table 1. In line with functional abnormalities, the morphological changes in the ctns‒/‒ zebrafish strikingly resembled those described in human cystinotic kidneys. Similar to humans39, pronephric podocytes showed partial foot process effacement and narrowed slit diaphragmatic spaces. Larval cystinotic PTECs showed lysosomal abundance and enlargement similar to human andmice cells12,15. We further elucidated the key mechanism behind the defective proximal tubular reabsorption in the mutant zebrafish larval model, which could be explained by the altered expression of the multi-ligand receptor, megalin at the proximal tubular brush border of ctns‒/‒ zebrafish larvae. The mis-trafficking of intracytoplasmic megalin evidenced by the sub-apical punctate
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