Renal claudin-14 expression is not required for regulating Mg²⁺ balance in mice

Abstract

The kidneys play a crucial role in maintaining calcium (Ca2+) and magnesium (Mg2+) homeostasis by regulating these minerals' reabsorption. In the thick ascending limb of Henle's loop (TAL), Ca2+ and Mg2+ are reabsorbed through the tight junctions by a shared paracellular pathway formed by claudin-16 and claudin-19. Hypercalcemia activates the Ca2+-sensing receptor (CaSR) in the TAL, causing upregulation of the pore-blocking claudin-14 (CLDN14) that reduces Ca2+ and Mg2+ reabsorption from this segment. Additionally, a high Mg2+ diet is known to increase both urinary Mg2+ and Ca2+ excretion. Since Mg2+ may also activate the CaSR, we aimed to investigate whether CaSR-dependent increases in CLDN14 expression also regulate urinary Mg2+ excretion in response to hypermagnesemia. Here we show that a Mg2+-enriched diet increased urinary Mg2+ and Ca2+ excretion in mice, however this occurred without detectable changes in renal CLDN14 expression. The administration of a high Mg2+ diet to Cldn14-/- mice did not cause more pronounced hypermagnesemia nor significantly alter urinary Mg2+ excretion. Finally, in vitro evaluation of CaSR-driven Cldn14 promoter activity in response to increasing Mg2+ concentrations revealed that Cldn14 expression only increases at supraphysiological extracellular Mg2+ levels. Together, these results suggest that CLDN14 is not involved in regulating extracellular Mg2+ balance following high dietary Mg2+ intake.

Differential parathyroid and kidney Ca2+-sensing receptor activation in autosomal dominant hypocalcemia 1

“…Furthermore, we have identified a Cldn14 promoter element that is directly regulated by the CaSR when the receptor is activated. 31 , 32 , 33 Collectively, these findings are in line with Cldn14 being a reasonable indicator of CaSR activity in the kidney. One should also consider that CaSR-dependent signaling in the TAL may differ from that reported in the parathyroid 39 so other surrogate markers for CaSR activity such as intracellular Ca 2+ release, would not necessarily be a better indicator.…”

“…To test the effect of increasing CaSR expression in the presence of a consistent extracellular Ca 2+ concentration sufficient to activate the CaSR (1.5 mM Ca 2+ ), a CaSR-responsive Cldn14 promoter reporter construct was transfected with increasing amounts of a myc-tagged CaSR into HEK293 cells. 33 Luminescence (and thus CaSR activity) significantly increased proportionately with increasing amounts of the CaSR construct transfected ( Figure 7 c). Western blotting confirmed increased CaSR expression after transfection with increased amount of construct ( Figures 7 d and S3).…”

“…Cloning of the Cldn14 promoter, which contains a CaSR-responsive element has been described previously. 31 , 32 , 33 HEK293 cells were purchased from the ATCC (Rockville, MD, USA; RRID:CVCL_0045) and cultured in DMEM with 10% FBS and 5% penicillin, streptomycin and glutamine as previously. 34 Cells were transfected via the calcium-phosphate method with the reporter construct, the pRL-TK renilla luciferase vector as an internal control and varying concentration of a vector encoding myc-tagged CaSR (Origene Inc.).…”

Gentamicin Inhibits Ca2+ Channel TRPV5 and Induces Calciuresis Independent of the Calcium-Sensing Receptor–Claudin-14 Pathway

“…Cloning of the Cldn14 promoter, which contains a CaSR responsive element, has been described 38 , 39 . HEK293 cells were purchased from the American Type Culture Collection (Rockville, MD) and cultured in DMEM with 10% FBS and 5% penicillin, streptomycin, and glutamine as described previously 40 .…”

Molecular mechanisms underlying paracellular calcium and magnesium reabsorption in the proximal tubule and thick ascending limb

“…Overall, it appears that CaSR activation via hypercalcemia leads to upregulation of CLDN14, which could contribute to reduced P Ca and P Mg and, thus, increased urinary excretion of divalent cations. Clearly, calcium is the main regulator of this response, and although magnesium can activate the CaSR, it does not lead to increased CLDN14 expression at concentrations observed under normal or hypermagnesemic conditions 148 …”