Cldn2-Trpv6 double knockout mice have hypocalcaemia and reduced bone mineral density

Central to calcium homeostasis and bone health is adequate intestinal calcium absorption. This occurs via either a transcellular or a paracellular pathway. In cases of low dietary calcium availability, a transcellular pathway that relies at least in part on the apical calcium channel Trpv6 predominates. However, when calcium is abundant in the diet, paracellular calcium absorption occurs through tight junction proteins, including claudin-2 (Cldn2). This tight junction protein also plays a role in calcium reabsorption from the renal proximal tubule. Despite this, Cldn2 knockout mice only display mildly altered calcium homeostasis characterized by hypercalciuria without evidence of altered bone mineral density (BMD), plasma calcium or calciotropic hormone levels. Similarly, mice expressing non-functional mutant Trpv6 (TRPV6 D541A/D541A ) channels have reduced intestinal calcium absorption without altered urine or plasma calcium levels, and only when fed a low calcium diet. We hypothesized that the lack of a more pronounced phenotype in these single functional knockout mice was due to the compensation of one pathway in the absence of the other. To test this hypothesis, we crossed claudin-2 knockout mice with mice expressing the mutant Trpv6, generating a functional double knockout (dko, Trpv6 mut/mut /Cldn2 -/- mice). Metabolic cage studies revealed hypocalcaemia (Male WT: 1.07 ± 0.02 mM, Male DKO: 0.97 ± 0.02 mM, p-value=0.0065, Female WT: 1.07 ± 0.02 mM, Female dko: 0.96 ± 0.03 mM , p-value= 0.0050) and hypercalciuria (Male WT Ca 2+ /Creatinine: 0.51 ± 0.05, Male dko:1.93 ± 0.18, p-value < 0.0001 Female WT Ca 2+ /Creatinine: 1.10 ± 0.13, Female dko: 3.56 ± 0.28 , p-value < 0.0001) in the dko mice. In contrast to the single claudin-2 KO mice and Trpv6 mutant animals the dko mice had elevated plasma calcitriol and parathyroid hormone. Moreover, BMD was reduced in the dko animals (Cortical BMD WT: 1.36 ± 0.02 g/cm 3 vs dko: 1.29 ± 0.02g/cm 3 ). RT-qPCR of intestinal segments and whole kidney revealed increased calbindin-D9k expression in the proximal colon (but not duodenum) and calbindin-D28k in the kidneys, consistent with compensatory increased transcellular absorption from these intestinal segments. Together these results are consistent with the paracellular pathway compensating for the loss of transcellular calcium absorption from the intestine. Funding: Alberta Innovates Graduate Studentship, NSERC, CIHR This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.