Kidney-specific claudin-2 deficiency leads to medullary nephrocalcinosis in mice
Christine V. Behm, Duuamene Nyimanu, Ony Araujo Galdino, Sadhana Kanoo, Young Chul Kim, Natalia Lopez, Helen Goodluck, Peter S. Rowe, Andrew P. Evan, André J. Sommer, Matthew N. Barr, Tracy Punshon, Volker Vallon, Brian P. Jackson, James C. Williams Jr., Alan S.L. Yu
Christine V. Behm, Duuamene Nyimanu, Ony Araujo Galdino, Sadhana Kanoo, Young Chul Kim, Natalia Lopez, Helen Goodluck, Peter S. Rowe, Andrew P. Evan, André J. Sommer, Matthew N. Barr, Tracy Punshon, Volker Vallon, Brian P. Jackson, James C. Williams Jr., Alan S.L. Yu
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Research Article Metabolism Nephrology

Kidney-specific claudin-2 deficiency leads to medullary nephrocalcinosis in mice

Abstract

Deposits of hydroxyapatite called Randall’s plaques are found in the renal papilla of calcium oxalate kidney stone formers and likely serve as the nidus for stone formation, but their pathogenesis is unknown. Claudin-2 is a paracellular ion channel that mediates calcium reabsorption in the renal proximal tubule. To investigate the role of renal claudin-2, we generated kidney tubule–specific claudin-2 conditional KO mice (KS-Cldn2 KO). KS-Cldn2 KO mice exhibited transient hypercalciuria in early life. Normalization of urine calcium was accompanied by a compensatory increase in expression and function of renal tubule calcium transporters, including in the thick ascending limb. Despite normocalciuria, KS-Cldn2 KO mice developed papillary hydroxyapatite deposits, beginning at 6 months of age, that resembled Randall’s plaques and tubule plugs. Bulk chemical tissue analysis and laser ablation–inductively coupled plasma mass spectrometry revealed a gradient of intrarenal calcium concentration along the corticomedullary axis in normal mice that was accentuated in KS-Cldn2 KO mice. Our findings provide evidence for the “vas washdown” hypothesis for Randall’s plaque formation and identify the corticomedullary calcium gradient as a potential target for therapies to prevent kidney stone disease.

Authors

Christine V. Behm, Duuamene Nyimanu, Ony Araujo Galdino, Sadhana Kanoo, Young Chul Kim, Natalia Lopez, Helen Goodluck, Peter S. Rowe, Andrew P. Evan, André J. Sommer, Matthew N. Barr, Tracy Punshon, Volker Vallon, Brian P. Jackson, James C. Williams Jr., Alan S.L. Yu

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Figure 1

General phenotype of constitutive, kidney-specific Cldn2-KO mice.

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General phenotype of constitutive, kidney-specific Cldn2-KO mice. (A) We...
(A) Western blot for claudin-2 in kidney and colon tissue from KO mice (Cre+) and control littermates (Cre–). (B) Cldn2 mRNA expression by quantitative PCR in kidneys, relative to ezrin, plotted as 2–ΔCt values. Differences are significant for genotype (P < 0.001), age (P = 0.03 for 10 vs. 4 weeks), and sex (P = 0.04) by 3-way ANOVA. (C) Immunofluorescence staining of kidney cortex and medulla with antibodies to claudin-2 (green) and ZO-1 (red). Orange/yellow fluorescence indicates colocalization of claudin-2 with ZO-1. Claudin-2 is detectable at the tight junctions (arrowheads) and basolateral membrane (arrows) of PTs in the cortex, and in thin descending limbs in the medulla of Cre– control but not Cre+ KO mice. Scale bars: 50 μm. (D) GFR determined from FITC-sinistrin clearance. n = 10–13 per group. (E) Determination of fractional excretion of lithium. Following pretreatment with bumetanide to block thick ascending limb sodium and lithium transport, male mice were given an i.v. bolus of LiCl, and plasma creatinine concentration, plasma lithium (mean of samples 3 minutes and 60 minutes after injection), and lithium excretion in a 1-hour urine collection were determined. n = 5 per group. (F) Systolic BP determined by tail cuff measurement. n = 10–13 per group.