Mitogenic action of calcium-sensing receptor on rat calvarial osteoblasts.
The parathyroid calcium-sensing receptor (CaR) plays a nonredundant role in systemic calcium homeostasis. In bone, Ca(2+)(o), a major extracellular factor in the bone microenvironment during bone remodeling, could potentially serve as an extracellular first messenger, acting via the CaR, that stimulates the proliferation of preosteoblasts and their differentiation to osteoblasts (OBs). Primary digests of rat calvarial OBs express the CaR as assessed by RT-PCR, Northern, and Western blot analysis, and immunocolocalization of the CaR with the OB marker cbfa-1. Real-time PCR revealed a significant increase in CaR mRNA in 5- and 7-d cultures compared with 3-d cultures post harvesting. High Ca(2+)(o) did not affect the expression of CaR mRNA during this time but up-regulated cyclin D (D1, D2, and D3) genes, which are involved in transition from the G1 to the S phase of the cell cycle, as well as the early oncogenes, c-fos and early growth response-1; high Ca(2+)(o) did not, however, alter IGF-I expression, a mitogenic factor for OBs. The high Ca(2+)(o)-dependent increase in the proliferation of OBs was attenuated after transduction with a dominant-negative CaR (R185Q), confirming that the effect of high Ca(2+)(o) is CaR mediated. Stimulation of proliferation by the CaR involves the Jun-terminal kinase (JNK) pathway, as high Ca(2+)(o) stimulated the phosphorylation of JNK in a CaR-mediated manner, and the JNK inhibitor SP600125 abolished CaR-induced proliferation. Our data, therefore, show that the parathyroid/kidney CaR expressed in rat calvarial OBs exerts a mitogenic effect that involves activation of the JNK pathway and up-regulation of several mitogenic genes.
Chattopadhyay
Naibedya
N
Division of Endocrinology, Diabetes and Hypertension, Beth Israel Seaconess Medical Center, Harvard Institutes of Medicine, Boston, Massachusetts 02115, USA. Naibedya@rics.bwh.harvard.edu.
Yano
Shozo
S
Tfelt-Hansen
Jacob
J
Rooney
Paul
P
Kanuparthi
Deepthi
D
Bandyopadhyay
Sanghamitra
S
Ren
Xianghui
X
Terwilliger
Ernest
E
Brown
Edward M
EM
D000818:Animals; D000918:Antibody Specificity; D015152:Blotting, Northern; D002118:Calcium / Q000378:metabolism; D002455:Cell Division / Q000502:physiology; D056741:Cyclin D; D016213:Cyclins / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics; D051766:Early Growth Response Protein 1; D015870:Gene Expression / Q000502:physiology; D017874:Immediate-Early Proteins / Q000235:genetics; D007334:Insulin-Like Growth Factor I / Q000235:genetics; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology* / Q000502:physiology*; D010766:Phosphorylation; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D044169:Receptors, Calcium-Sensing / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012886:Skull / Q000166:cytology; D014157:Transcription Factors / Q000235:genetics; D015854:Up-Regulation
D056741:Cyclin D; D016213:Cyclins; D004268:DNA-Binding Proteins; D051766:Early Growth Response Protein 1; C486110:Egr1 protein, rat; D017874:Immediate-Early Proteins; D016760:Proto-Oncogene Proteins c-fos; D012333:RNA, Messenger; D044169:Receptors, Calcium-Sensing; D014157:Transcription Factors; D007334:Insulin-Like Growth Factor I; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D002118:Calcium