Targeted ablation of the PTH/PTHrP receptor in osteocytes impairs bone structure and homeostatic calcemic responses

Abstract

Parathyroid hormone (PTH) is a major physiologic regulator of calcium, phosphorous and skeletal homeostasis. Cells of the osteoblastic lineage are key targets of PTH action in bone, and recent evidence suggests that osteocytes might be important in the anabolic effects of PTH. To understand the role of PTH signaling through the PTH/PTHrP receptors (PPR) in osteocytes and to determine the role(s) of these cells in mediating the effects of the hormone, we have generated mice in which PPR expression is specifically ablated in osteocytes. Transgenic mice in which the 10Kb-Dmp1 promoter drives a tamoxifen-inducible Cre –recombinase were mated with animals in which exon1 of PPR is flanked by Lox-P sites. In these animals, osteocyte-selective PPR knockout (Ocy-PPRcKO mice) could be induced by administration of tamoxifen. Histological analysis revealed a reduction in trabecular bone and mild osteopenia in Ocy-PPRcKO mice. Reduction of trabeculae number and thickness was also detected by μCT analysis whereas BV/TV% was unchanged. These findings were associated with an increase in Sost and sclerostin expression. When Ocy-PPRcKO mice were subjected to a low calcium diet, to induce secondary hyperparathyroidism, their blood calcium levels were significantly lower than littermate controls. Moreover, PTH was unable to suppress Sost and sclerostin expression in the Ocy-PPRcKO animals, suggesting an important role of PTH signaling in osteocytes for proper bone remodeling and calcium homeostasis.

Loss of XLαs (extra-large αs) imprinting results in early postnatal hypoglycemia and lethality in a mouse model of pseudohypoparathyroidism Ib

“…4D), suggesting that XLαs might still play a role in mediating PTH actions in bone. Consistent with this interpretation, XLαs protein expression has been detected in adult mouse osteocytes (51), which are importantly involved in the PTHdependent regulation of bone remodeling and calcium homeostasis (52)(53)(54).…”

The Wnt Inhibitor Sclerostin Is Up-regulated by Mechanical Unloading in Osteocytes in Vitro

“…Protein concentrations were quantified (Bio-Rad Protein Assay, Bio-Rad), and 10 g was separated on a 4 -20% Tris-glycine denaturing gel (Life Technologies) and transferred to a PVDF membrane using the Trans-Blot Turbo (Bio-Rad) system according to the manufacturer's recommendations. The membrane was blocked with 3% BSA and 5% nonfat milk in Tris-buffered saline containing 0.05% Tween 20 (TBST) for 1 h and then incubated with goat polyclonal mouse sclerostin antibody (1:200; R&D Systems, Minneapolis, MN) overnight at 4°C (30). After washing, secondary antibody (1:5000) was incubated for 1 h at room temperature and then developed using enhanced chemiluminescence (Thermo Scientific) (30).…”

“…Indeed, there is an inverse correlation between PTH and sclerostin in male hypoparathyroid subjects (27), and PTH infusion in healthy men induces a decline in circulating sclerostin (28). Both in vivo and in vitro, PTH decreases sclerostin expression via activation of the PTH receptor expressed on osteocytes (29), and mice lacking the PTH receptor specifically in osteocytes have elevated expression of sclerostin (30). Thus, in vivo studies cannot determine whether suppression of PTH or other changes in cytokines, such as prostaglandin E 2 (PGE 2 ), are driving the increases in serum sclerostin following unloading.…”

El receptor 2 de VEGF (VEGFR2) y el receptor 1 de la PTH (PTH1R) actúan como mediadores de la respuesta anti-apoptótica al estímulo mecánico en las células osteocíticas MLO-Y4

“…Recent studies in genetically engineered mice indicate that the action of PTH requires a functional PTH1R in osteocitos 22 . From a pharmacological perspective, intermittent administration of PTH in mice attenuates rapidly osteoblast apoptosis in the vertebrae; this effect appears to be only a consequence of direct hormone action on osteoblasts, but also indirectly through its inhibitory effect on the expression of Sost/sclerostin in osteocytes 20,36,37 .…”

“…Mice heterozygous deletion of the gene for PTHrelated protein (PTHrP), local counterpart bone, osteoblasts show osteopenia associated with decreased survival of osteoblasts and osteocytes 21 . Furthermore, mice with conditional suppression of PTH1R osteocytes specifically exhibit altered homeostasis of calcium and osteopenia 22 . In contrast, mice with constitutive overexpression of this receptor in the osteocytes show increased periosteal bone formation, associated with activation of the Wnt pathway and decreased osteoblastic apoptosis 23 .…”