[Show abstract][Hide abstract] ABSTRACT: Prostate cancer metastasizes frequently to bone. Elevated extracellular calcium concentrations ([Ca(2+)](o)) stimulate parathyroid hormone-related protein (PTHrP) secretion from normal and malignant cells, potentially acting via the [Ca(2+)](o)-sensing receptor (CaR). Because prostate cancers produce PTHrP, if high [Ca(2+)](o) stimulates PTHrP secretion via the CaR, this could initiate a mechanism whereby osteolysis caused by bony metastases of prostate cancer promotes further bone resorption. We investigated whether the prostate cancer cell lines LnCaP and PC-3 express the CaR and whether polycationic CaR agonists stimulate PTHrP release. Both PC-3 and LnCaP prostate cancer cell lines expressed bona fide CaR transcripts by Northern analysis and RT-PCR and CaR protein by immunocytochemistry and Western analysis. The polycationic CaR agonists [Ca(2+)](o), neomycin, and spermine each concentration dependently stimulated PTHrP secretion from PC-3 cells, as measured by immunoradiometric assay, with maximal, 3.2-, 3.6-, and 4.2-fold increases, respectively. In addition, adenovirus-mediated infection of PC-3 cells with a dominant negative CaR construct attenuated high [Ca(2+)](o)-evoked PTHrP secretion, further supporting the CaR's mediatory role in this process. Finally, pretreating PC-3 cells with transforming growth factor (TGF)-beta(1) augmented both basal and high [Ca(2+)](o)-stimulated PTHrP secretion. Thus, in PTHrP-secreting prostate cancers metastatic to bone, the CaR could initiate a vicious cycle, whereby PTHrP-induced bone resorption releases [Ca(2+)](o) and TGF-beta stored within bone, further increasing PTHrP release and osteolysis.
Full-text · Article · Jan 2002 · AJP Endocrinology and Metabolism
[Show abstract][Hide abstract] ABSTRACT: Recently, substantial evidence has accumulated that the G-protein-coupled, extracellular calcium (Ca2+o)-sensing receptor (CaR) is expressed in bone marrow-derived cells, including osteoblasts, stromal cells, monocytes–macrophages, and osteoclast precursor cells. Our previous studies have shown that the mouse osteoblastic MC3T3-E1 cell line also expresses the CaR and exhibits mitogenic responses when exposed to various CaR agonists. In this study, in order to understand the signaling pathway(s) mediating this response, we studied the effects of CaR agonists on the phosphorylation of p42/44 mitogen-activated protein kinase (MAPK) (Erk1/2), p38 MAPK, and c-Jun N-terminal kinase (JNK) in MC3T3-E1 cells. Raising the level of Ca2+o (4.5 mM) or addition of the polycationic CaR agonists, gadolinium (Gd3+) (25 μM), neomycin (300 μM) or spermine (1 mM), each stimulated phosphorylation of both p42/44 and p38 MAPKs, but not JNK, as assessed using phospho-specific antibodies to the respective MAPKs. Furthermore, phosphorylation of p42/44 and p38 MAPK were markedly inhibited by their selective and potent inhibitors, PD98059 (50 μM) and SB203580 (10 μM), respectively. Finally, the two inhibitors suppressed [3H]thymidine incorporation into DNA in MC3T3-E1 cells at a normal level of Ca2+o (1.8 mM) as well as when stimulated by high (4.5 mM) Ca2+o, Gd3+, or neomycin. Thus, in mouse osteoblastic MC3T3-E1 cells, both the p42/44 and p38 MAPK cascades play pivotal roles in CaR-stimulated mitogenic responses.
No preview · Article · Jan 2001 · Biochemical and Biophysical Research Communications
[Show abstract][Hide abstract] ABSTRACT: Metastasis of breast cancer to bone occurs with advanced disease and produces substantial morbidity. Secretion of PTH-related peptide (PTHrP) from breast cancer cells is thought to play a key role in osteolytic metastases and is increased by transforming growth factor-beta (TGFbeta), which is released from resorbed bone. Elevated extracellular calcium (Ca2+(o)) also stimulates PTHrP secretion from various normal and malignant cells, an action that could potentially be mediated by the Ca2+(o)-sensing receptor (CaR) originally cloned from the parathyroid gland. Indeed, we previously showed that both normal breast ductal epithelial cells and primary breast cancers express the CaR. In this study we investigated whether the MCF-7 and MDA-MB-231 human breast cancer cell lines express the CaR and whether CaR agonists modulate PTHrP secretion. Northern blot analysis and RT-PCR revealed bona fide CaR transcripts, and immunocytochemistry and Western analysis with a specific anti-CaR antiserum demonstrated CaR protein expression in both breast cancer cell lines. Furthermore, elevated Ca2+(o) and the polycationic CaR agonists, neomycin and spermine, stimulated PTHrP secretion dose dependently, with maximal, 2.1- to 2.3-fold stimulation. In addition, pretreatment of MDA-MB-231 cells overnight with TGFbeta1 (0.2, 1, or 5 ng/ml) augmented both basal and high Ca2+-stimulated PTHrP secretion. Thus, in PTHrP-secreting breast cancers metastatic to bone, the CaR could potentially participate in a vicious cycle in which PTHrP-induced bone resorption raises the levels of Ca2+(o) and TGFbeta within the bony microenvironment, which then act in concert to evoke further PTHrP release and worsening osteolysis.
[Show abstract][Hide abstract] ABSTRACT: Humoral hypercalcemia of malignancy (HHM) occurs when secretion of parathyroid hormone-related peptide (PTHrP) by cancer cells causes hypercalcemia in the absence of skeletal metastases. High extracellular calcium (Ca2+o) increases secretion of PTH-like bioactivity by rat H-500 leydig cells, a transplantable model of HHM, an action potentially mediated by the Ca2+o-sensing receptor (CaR). In this study we investigated whether H-500 cells express the CaR and, if so, whether CaR agonists modulate PTHrP secretion. Northern blot analysis and RT-PCR revealed bona fide CaR transcript(s), and immunocytochemistry and Western analysis with a specific anti-CaR antiserum demonstrated CaR protein expression in H-500 cells. Furthermore, high Ca2+o and neomycin stimulated PTHrP secretion dose-dependently with maximal 2.7- and 3.3-fold increases at 5 mM Ca2+o and 300 μM neomycin, respectively. Thus in HHM caused by H-500 cells, the CaR could participate in a vicious cycle whereby PTHrP-induced increases in Ca2+o further stimulate PTHrP release and exacerbate hypercalcemia.
No preview · Article · Apr 2000 · Biochemical and Biophysical Research Communications