[en] Megalin/LRP2 is a major receptor supporting apical endocytosis in kidney proximal tubular cells. We have previously reported that kidney-specific perinatal ablation of the megalin gene in cystinotic mice, a model of nephropathic cystinosis, essentially blocks renal cystine accumulation and partially preserves kidney tissue integrity. Here, we examined whether inhibition of the megalin pathway in adult cystinotic mice by dietary supplementation (5x-fold vs control regular diet) with the dibasic amino-acids (dAAs), lysine or arginine, both of which are used to treat patients with other rare metabolic disorders, could also decrease renal cystine accumulation and protect cystinotic kidneys. Using surface plasmon resonance, we first showed that both dAAs compete for protein ligand binding to immobilized megalin in a concentration-dependent manner, with identical inhibition curves by L- and D-stereoisomers. In cystinotic mice, 2-month diets with 5x-L-lysine and 5x-L-arginine were overall well tolerated, while 5x-D-lysine induced strong polyuria but no weight loss. All diets induced a marked increase of dAA urinary excretion, most prominent under 5x-D-lysine, without sign of kidney insufficiency. Renal cystine accumulation was slowed down approx. twofold by L-dAAs, and totally suppressed by D-lysine. We conclude that prolonged dietary manipulation of the megalin pathway in kidneys is feasible, tolerable and can be effective in vivo.
Disciplines :
Urology & nephrology
Author, co-author :
Rega, L R; Nephrology Research Unit, Translational Pediatrics and Clinical Genetics Research Area, Bambino Gesù Children's Hospital, IRCCS, Rome, Italy
Janssens, V; Cell Biology Unit, de Duve Institute and Louvain University Medical School, Brussels, Belgium
Graversen, J H; Department of Molecular Medicine, University of Southern Denmark, Odense, Denmark
Moestrup, S K; Department of Molecular Medicine, University of Southern Denmark, Odense, Denmark
Cairoli, S; Division of Metabolic Diseases and Drug Biology, Bambino Gesù Children's Hospital, IRCCS, Rome, Italy
Goffredo, B M; Division of Metabolic Diseases and Drug Biology, Bambino Gesù Children's Hospital, IRCCS, Rome, Italy
Nevo, N; Laboratoire des Maladies Rénales Héréditaires, Inserm UMR 1163, Institut Imagine, Université Paris Cité, Paris, France
Courtoy, G E; Imaging Platform (2IP), Institut de Recherche Expérimentale et Clinique, Louvain University Medical School, Brussels, Belgium
Jouret, François ; Centre Hospitalier Universitaire de Liège - CHU > > Service de néphrologie
Antignac, C; Laboratoire des Maladies Rénales Héréditaires, Inserm UMR 1163, Institut Imagine, Université Paris Cité, Paris, France
Emma, F; Nephrology Research Unit, Translational Pediatrics and Clinical Genetics Research Area, Bambino Gesù Children's Hospital, IRCCS, Rome, Italy
Pierreux, C E; Cell Biology Unit, de Duve Institute and Louvain University Medical School, Brussels, Belgium. christophe.pierreux@uclouvain.be
Courtoy, P J; Cell Biology Unit, de Duve Institute and Louvain University Medical School, Brussels, Belgium. pierre.courtoy@uclouvain.be
Nielsen, R., Christensen, E. I. & Birn, H. Megalin and cubilin in proximal tubule protein reabsorption: From experimental models to human disease. Kidney Int. 89(1), 58–67 (2016). DOI: 10.1016/j.kint.2015.11.007
Perez Bay, A. E. et al. The fast-recycling receptor Megalin defines the apical recycling pathway of epithelial cells. Nat. Commun. 7, 11550 (2016). DOI: 10.1038/ncomms11550
Shipman, K. E. et al. An adaptable physiological model of endocytic megalin trafficking in opossum kidney cells and mouse kidney proximal tubule. Function 3(6), zqac046 (2022). DOI: 10.1093/function/zqac046
Grieco, G. et al. Vps34/PI3KC3 deletion in kidney proximal tubules impairs apical trafficking and blocks autophagic flux, causing a Fanconi-like syndrome and renal insufficiency. Sci. Rep. 8(1), 14133 (2018). DOI: 10.1038/s41598-018-32389-z
Rinschen, M. M. et al. VPS34-dependent control of apical membrane function of proximal tubule cells and nutrient recovery by the kidney. Sci. Signal. 15(762), eabo7940 (2022). DOI: 10.1126/scisignal.abo7940
Christensen, E. I., Wagner, C. A. & Kaissling, B. Uriniferous tubule: Structural and functional organization. Compr. Physiol. 2(2), 805–861 (2012). DOI: 10.1002/cphy.c100073
Nielsen, R. et al. Endocytosis provides a major alternative pathway for lysosomal biogenesis in kidney proximal tubular cells. Proc. Natl. Acad. Sci. USA 104(13), 5407–5412 (2007). DOI: 10.1073/pnas.0700330104
Johanns, M. et al. Cellular uptake of proMMP-2:TIMP-2 complexes by the endocytic receptor megalin/LRP-2. Sci. Rep. 7(1), 4328 (2017). DOI: 10.1038/s41598-017-04648-y
Moestrup, S. K. et al. Evidence that epithelial glycoprotein 330/megalin mediates uptake of polybasic drugs. J. Clin. Investig. 96(3), 1404–1413 (1995). DOI: 10.1172/JCI118176
Mogensen, C. E. & Sølling, K. Studies on renal tubular protein reabsorption: Partial and near complete inhibition by certain amino acids. Scand. J. Clin. Lab. Investig. 37(6), 477–486 (1977). DOI: 10.3109/00365517709101835
Ottosen, P. D. et al. Inhibition of protein reabsorption in the renal proximal tubule by basic amino acids. Ren. Physiol. 8(2), 90–99 (1985).
Andersen, C. B. & Moestrup, S. K. How calcium makes endocytic receptors attractive. Trends Biochem. Sci. 39(2), 82–90 (2014). DOI: 10.1016/j.tibs.2013.12.003
Thelle, K. et al. Characterization of proteinuria and tubular protein uptake in a new model of oral l-lysine administration in rats. Kidney Int. 69(8), 1333–1340 (2006). DOI: 10.1038/sj.ki.5000272
Jamar, F. et al. 86Y-DOTA0)-D-Phe1-Tyr3-octreotide (SMT487)–a phase 1 clinical study: Pharmacokinetics, biodistribution and renal protective effect of different regimens of amino acid co-infusion. Eur. J. Nucl. Med. Mol. Imaging 30(4), 510–518 (2003). DOI: 10.1007/s00259-003-1117-1
Barone, R. et al. Endocytosis of the somatostatin analogue, octreotide, by the proximal tubule-derived opossum kidney (OK) cell line. Kidney Int. 67(3), 969–976 (2005). DOI: 10.1111/j.1523-1755.2005.00160.x
Molema, F. et al. Evaluation of dietary treatment and amino acid supplementation in organic acidurias and urea-cycle disorders: On the basis of information from a European multicenter registry. J. Inherit. Metab. Dis. 42(6), 1162–1175 (2019). DOI: 10.1002/jimd.12066
Elpeleg, N. & Korman, S. H. Sustained oral lysine supplementation in ornithine delta-aminotransferase deficiency. J. Inherit. Metab. Dis. 24(3), 423–424 (2001). DOI: 10.1023/A:1010545811361
Nevo, N. et al. Renal phenotype of the cystinosis mouse model is dependent upon genetic background. Nephrol. Dial Transplant. 25(4), 1059–1066 (2010). DOI: 10.1093/ndt/gfp553
Cheung, P. Y. et al. In vitro and in vivo models to study nephropathic cystinosis. Cells 11(1), 6 (2021). DOI: 10.3390/cells11010006
Hollywood, J. A. et al. Cystinosin-deficient rats recapitulate the phenotype of nephropathic cystinosis. Am. J. Physiol. Renal Physiol. 323(2), F156-f170 (2022). DOI: 10.1152/ajprenal.00277.2021
Krohn, P. et al. Multisystem involvement, defective lysosomes and impaired autophagy in a novel rat model of nephropathic cystinosis. Hum. Mol. Genet. 31(13), 2262–2278 (2022). DOI: 10.1093/hmg/ddac033
Janssens, V. et al. Protection of cystinotic mice by kidney-specific megalin ablation supports an endocytosis-based mechanism for nephropathic cystinosis progression. J. Am. Soc. Nephrol. 30(11), 2177–2190 (2019). DOI: 10.1681/ASN.2019040371
Cherqui, S. et al. The targeting of cystinosin to the lysosomal membrane requires a tyrosine-based signal and a novel sorting motif. J. Biol. Chem. 276(16), 13314–13321 (2001). DOI: 10.1074/jbc.M010562200
Kalatzis, V. et al. Cystinosin, the protein defective in cystinosis, is a H(+)-driven lysosomal cystine transporter. EMBO J. 20(21), 5940–5949 (2001). DOI: 10.1093/emboj/20.21.5940
Gahl, W. A. & Thoene, J. G. Cystinosis: A disorder of lysosomal membrane transport. In The Online Metabolic and Molecular Bases of Inherited Disease (eds Valle, D. L. et al.) (McGraw-Hill Education, 2019).
Cherqui, S. & Courtoy, P. J. The renal Fanconi syndrome in cystinosis: Pathogenic insights and therapeutic perspectives. Nat. Rev. Nephrol. 13(2), 115–131 (2017). DOI: 10.1038/nrneph.2016.182
Adelmann, C. H. et al. MFSD12 mediates the import of cysteine into melanosomes and lysosomes. Nature 588(7839), 699–704 (2020). DOI: 10.1038/s41586-020-2937-x
Thoene, J. G. & Lemons, R. M. Cystine accumulation in cystinotic fibroblasts from free and protein-linked cystine but not cysteine. Biochem. J. 208(3), 823–830 (1982). DOI: 10.1042/bj2080823
Lloyd, J. B. Disulphide reduction in lysosomes. The role of cysteine. Biochem. J. 237(1), 271–272 (1986). DOI: 10.1042/bj2370271
Jamalpoor, A. et al. molecular mechanisms and treatment options of nephropathic cystinosis. Trends Mol. Med. 27(7), 673–686 (2021). DOI: 10.1016/j.molmed.2021.04.004
Gaide Chevronnay, H. P. et al. Time course of pathogenic and adaptation mechanisms in cystinotic mouse kidneys. J. Am. Soc. Nephrol. 25(6), 1256–1269 (2014). DOI: 10.1681/ASN.2013060598
Raggi, C. et al. Dedifferentiation and aberrations of the endolysosomal compartment characterize the early stage of nephropathic cystinosis. Hum. Mol. Genet. 23(9), 2266–2278 (2014). DOI: 10.1093/hmg/ddt617
Ivanova, E. A. et al. Endo-lysosomal dysfunction in human proximal tubular epithelial cells deficient for lysosomal cystine transporter cystinosin. PLoS ONE 10(3), e0120998 (2015). DOI: 10.1371/journal.pone.0120998
Fotiadis, D., Kanai, Y. & Palacín, M. The SLC3 and SLC7 families of amino acid transporters. Mol. Aspects Med. 34(2–3), 139–158 (2013). DOI: 10.1016/j.mam.2012.10.007
Rinschen, M. M. et al. Accelerated lysine metabolism conveys kidney protection in salt-sensitive hypertension. Nat. Commun. 13(1), 4099 (2022). DOI: 10.1038/s41467-022-31670-0
Mizoguchi, K. et al. Human cystinuria-related transporter: Localization and functional characterization. Kidney Int. 59(5), 1821–1833 (2001). DOI: 10.1046/j.1523-1755.2001.0590051821.x
Boger, R. H. & Bode-Boger, S. M. The clinical pharmacology of l-arginine. Annu. Rev. Pharmacol. Toxicol. 41, 79–99 (2001). DOI: 10.1146/annurev.pharmtox.41.1.79
Prencipe, G. et al. Inflammasome activation by cystine crystals: Implications for the pathogenesis of cystinosis. J. Am. Soc. Nephrol. 25(6), 1163–1169 (2014). DOI: 10.1681/ASN.2013060653
Jouret, F. et al. Cystic fibrosis is associated with a defect in apical receptor-mediated endocytosis in mouse and human kidney. J. Am. Soc. Nephrol. 18(3), 707–718 (2007). DOI: 10.1681/ASN.2006030269
Schuh, C. D. et al. Combined structural and functional imaging of the kidney reveals major axial differences in proximal tubule endocytosis. J. Am. Soc. Nephrol. 29(11), 2696–2712 (2018). DOI: 10.1681/ASN.2018050522
Grahammer, F. et al. mTOR regulates endocytosis and nutrient transport in proximal tubular cells. J. Am. Soc. Nephrol. 28(1), 230–241 (2017). DOI: 10.1681/ASN.2015111224
Thoene, J. G. & Lemons, R. Modulation of the intracellular cystine content of cystinotic fibroblasts by extracellular albumin. Pediatr. Res. 14(6), 785–787 (1980). DOI: 10.1203/00006450-198006000-00001
Johnson, J. L. et al. Upregulation of the Rab27a-dependent trafficking and secretory mechanisms improves lysosomal transport, alleviates endoplasmic reticulum stress, and reduces lysosome overload in cystinosis. Mol. Cell Biol. 33(15), 2950–2962 (2013). DOI: 10.1128/MCB.00417-13
Andrzejewska, Z. et al. Cystinosin is a component of the vacuolar H+-ATPase-ragulator-rag complex controlling mammalian target of rapamycin complex 1 signaling. J. Am. Soc. Nephrol. 27(6), 1678–1688 (2016). DOI: 10.1681/ASN.2014090937
Ivanova, E. A. et al. Altered mTOR signalling in nephropathic cystinosis. J. Inherit. Metab. Dis. 39(3), 457–464 (2016). DOI: 10.1007/s10545-016-9919-z
Hollywood, J. A. et al. Use of human induced pluripotent stem cells and kidney organoids to develop a cysteamine/mTOR inhibition combination therapy for cystinosis. J. Am. Soc. Nephrol. 31(5), 962–982 (2020). DOI: 10.1681/ASN.2019070712
Berquez, M. et al. Lysosomal cystine export regulates mTORC1 signaling to guide kidney epithelial cell fate specialization. bioRxiv 2022.08.28.505580 (2022).
Christensen, H. N. Role of amino acid transport and countertransport in nutrition and metabolism. Physiol. Rev. 70(1), 43–77 (1990). DOI: 10.1152/physrev.1990.70.1.43
Moestrup, S. K. et al. Epithelial glycoprotein-330 mediates endocytosis of plasminogen activator-plasminogen activator inhibitor type-1 complexes. J. Biol. Chem. 268(22), 16564–16570 (1993). DOI: 10.1016/S0021-9258(19)85456-X
Moestrup, S. K. et al. Megalin-mediated endocytosis of transcobalamin-vitamin-B12 complexes suggests a role of the receptor in vitamin-B12 homeostasis. Proc. Natl. Acad. Sci. USA 93(16), 8612–8617 (1996). DOI: 10.1073/pnas.93.16.8612
Hansen, M. & Nexø, E. Cobalamin binding proteins in human seminal plasma. Scand. J. Clin. Lab. Investig. 52(7), 647–652 (1992). DOI: 10.3109/00365519209115508
McNeal, C. J. et al. Safety and effectiveness of arginine in adults. J. Nutr. 146(12), 2587S-2593S (2016). DOI: 10.3945/jn.116.234740
Courtoy, G. E. et al. Digital image analysis of picrosirius red staining: A robust method for multi-organ fibrosis quantification and characterization. Biomolecules 10(11), 1585 (2020). DOI: 10.3390/biom10111585
Vogel, B. et al. Determination of collagen content within picrosirius red stained paraffin-embedded tissue sections using fluorescence microscopy. MethodsX 2, 124–134 (2015). DOI: 10.1016/j.mex.2015.02.007
