The abnormal phenotypes of cartilage and bone in calcium-sensing receptor deficient mice are dependent on the actions of calcium, phosphorus, and PTH.

Patients with neonatal severe hyperparathyroidism (NSHPT) are homozygous for the calcium-sensing receptor (CaR) mutation and have very high circulating PTH, abundant parathyroid hyperplasia, and severe life-threatening hypercalcemia. Mice with homozygous deletion of CaR mimic the syndrome of NSHPT. To determine effects of CaR deficiency on skeletal development and interactions between CaR and 1,25(OH)(2)D(3) or PTH on calcium and skeletal homeostasis, we compared the skeletal phenotypes of homozygous CaR-deficient (CaR(-/-)) mice to those of double homozygous CaR- and 1α(OH)ase-deficient [CaR(-/-)1α(OH)ase(-/-)] mice or those of double homozygous CaR- and PTH-deficient [CaR(-/-)PTH(-/-)] mice at 2 weeks of age. Compared to wild-type littermates, CaR(-/-) mice had hypercalcemia, hypophosphatemia, hyperparathyroidism, and severe skeletal growth retardation. Chondrocyte proliferation and PTHrP expression in growth plates were reduced significantly, whereas trabecular volume, osteoblast number, osteocalcin-positive areas, expression of the ALP, type I collagen, osteocalcin genes, and serum ALP levels were increased significantly. Deletion of 1α(OH)ase in CaR(-/-) mice resulted in a longer lifespan, normocalcemia, lower serum phosphorus, greater elevation in PTH, slight improvement in skeletal growth with increased chondrocyte proliferation and PTHrP expression, and further increases in indices of osteoblastic bone formation. Deletion of PTH in CaR(-/-) mice resulted in rescue of early lethality, normocalcemia, increased serum phosphorus, undetectable serum PTH, normalization in skeletal growth with normal chondrocyte proliferation and enhanced PTHrP expression, and dramatic decreases in indices of osteoblastic bone formation. Our results indicate that reductions in hypercalcemia play a critical role in preventing the early lethality of CaR(-/-) mice and that defects in endochondral bone formation in CaR(-/-) mice result from effects of the marked elevation in serum calcium concentration and the decreases in serum phosphorus concentration and skeletal PTHrP levels, whereas the increased osteoblastic bone formation results from direct effects of PTH.

Parathyroid hormone contributes to regulating milk calcium content and modulates neonatal bone formation cooperatively with calcium.

To determine whether PTH and calcium (Ca) interact in neonatal bone formation, female lactating mice either heterozygous (PTH(+/-)) or homozygous (PTH(-/-)) for targeted deletion of the pth gene were fed either a normal (1% Ca, 0.6% phosphate) or high-Ca diet (2% Ca and 0.4% phosphate). Dietary effects on milk Ca content and Ca-regulating hormones were determined in dams, and the effects of milk content were assessed on bone turnover in 3-wk-old pups. On the normal diet, milk Ca and 1,25-dihydroxyvitamin D(3) levels were lower, but milk PTH-related protein levels were higher in the PTH(-/-) dams compared with the PTH(+/-) dams. On the high-Ca diet, milk Ca levels were higher, but milk 1,25-dihydroxyvitamin D(3) and PTH-related protein levels were lower in both PTH(+/-) and PTH(-/-) dams. In pups fed by PTH(-/-) dams compared with pups fed by PTH(+/-) dams on normal diets, bone mineral density, trabecular bone volume relative to tissue volume, and the number of osteoblasts were reduced in both PTH(+/-) (32.5 +/- 1.2 vs. 39.6 +/- 1.5 mg/cm(2), P < 0.05; 23.3 +/- 1.6 vs. 29.2 +/- 2.8%, P < 0.01; and 94.2 +/- 8.2 vs. 123.5 +/- 3.5/mm(2), P < 0.01, respectively) and PTH(-/-) (20.4 +/- 0.9 vs. 27.0 +/- 1.2 mg/mm(2), P < 0.05; 16.8 +/- 1.9 vs. 19.3 +/- 2.1%, P < 0.05; and 48.6 +/- 7.9 vs. 90.5 +/- 8.6/mm(2), P < 0.01, respectively) pups but were lower in the PTH(-/-) pups compared with the PTH(+/-) pups. In contrast, in pups fed by either PTH(+/-) or PTH(-/-) dams on the high-Ca diet, bone mineral density, bone volume/tissue volume, and osteoblast numbers were significantly higher, in both PTH(+/-) (50.5 +/- 1.7 vs. 58.7 +/- 2.0 mg/mm(2), P < 0.05; 37.9 +/- 5.2 vs. 46.1 +/- 5.1, P < 0.05; and 120.5 +/- 9.2 vs. 159.3 +/- 14.7/mm(2), P < 0.01, respectively) and PTH(-/-) (33.0 +/- 1.2 vs. 47.5 +/- 2.2 mg/mm(2), P < 0.001; 23.8 +/- 3.1 vs. 35.9 +/- 2.0, P < 0.05; and 78.7 +/- 10.1 vs. 99.8 +/- 13.6/mm(2), P < 0.05, respectively), and were highest in the PTH(+/-) pups fed by the PTH(+/-) dams on the high-Ca diet. These results indicate that PTH can modulate Ca content of milk, and that PTH and Ca can each exert cooperative roles on osteoblastic bone formation in the neonate.