Characterization of vitamin D-deficient klotho-/- mice: Do increased levels of serum 1,25(OH)2D3 cause disturbed calcium and phosphate homeostasis in klotho-/- mice?

摘要

BackgroundKlotho-/- mice display disturbed Ca2+ and vitamin D homeostasis. Renal cytochrome p450 27b1 (Cyp27b1), the enzyme that catalyzes the hydrolysis to 1,25-dihydroxyvitamin D3 (1,25(OH) 2D3), is increased in klotho-/- mice, and a 1,25(OH)2D3-deficient diet partially normalized Ca 2+ homeostasis in these klotho-/- mice. The aim of the present study was to further delineate the interplay between 1,25(OH) 2D3 and klotho and their relative contribution to the Ca2+ homeostasis of klotho-/- mice.MethodsDouble- klotho-/-/Cyp27b1-/- mice were generated and mice aged 8-12 weeks were housed in metabolic cages to collect 24-h urine. Blood samples were taken and the animals were sacrificed, and the kidney and duodenum tissues were sampled for RNA extraction. The bone was fixed in 10 v/v formalin and analysed by microcomputed tomography (μCT) scans.ResultsKlotho -/-/Cyp27b1-/- mice, like Cyp27b1-/- mice, displayed significantly decreased serum total calcium concentrations compared with wild-type mice (1.44 ± 0.03 and 2.25 ± 0.02 mM) along with normal urinary total calcium excretion. Hyperphosphataemia of klotho -/- mice normalized to wild-type levels in klotho-/-/ Cyp27b1-/- mice. The mRNA levels of duodenal transient receptor potential vanilloid subtype 6 (TRPV6) and calcium-binding protein-D 9K, and renal calbindin-D28K and NCX1 were significantly reduced in the double knockouts compared with wild-type or klotho-/- mice. Elevated TRPV5 protein levels in klotho-/- mice normalized to wild type in klotho-/-/Cyp27b1-/- mice, but were decreased in Cyp27b1-/- mice. μCT scans showed that klotho -/-/Cyp27b1-/- mice, as Cyp27b1-/- mice, display significant bone hypomineralization and severely decreased bone mass. Klotho-/- mice show a reduced bone mass and increased trabecular numbers.ConclusionsKlotho-/-/Cyp27b1-/- mice resemble Cyp27b1-/- mice. Since 1,25(OH)2D3 is absent in these mice, our results imply that Ca2+ homeostasis in klotho -/- mice is affected by their excessive 1,25(OH)2D 3 levels.