Submitted on July 14, 2005
Accepted on April 4, 2006
CGRP inhibits osteoprotegerin production in human osteoblast-like cells via cAMP/PKA dependent pathway
Isabella Villa1, Emanuela Mrak1, Alessandro Rubinacci1, Flavio Ravasi1, and Francesca Guidobono1*
1 Bone Metabolic Unit, Scientific Institute H San Raffaele, Milano, Italy; Dept. of Pharmacology, Chempherapy and Medical Toxicology, University of Milan, Milano, Italy
* To whom correspondence should be addressed. E-mail: francesca.guidobono{at}unimi.it.
The OPG/RANKL/RANK system was evaluated as a potential target of CGRP anabolic activity on bone. Primary cultures of human osteoblast-like cells (hOB) express calcitonin receptor-like receptor (CLR) and receptor activity modifying protein 1 (RAMP1) and, as CGRP stimulates cAMP (one of the modulators of osteoprotegerin, OPG, production in osteoblasts), it was investigated whether it affects OPG secretion and expression in hOB. CGRP treatment of hOB (10-11M-10-7M) dose-dependently inhibited OPG secretion with an EC50 of 1.08x10-10M, and also decreased its expression. This action was blocked by the antagonist CGRP8-37. Forskolin, a stimulator of cAMP production, and dibutyryl cAMP also reduced the production of OPG. CGRP (10-8M) enhanced protein kinase A (PKA) activity in hOB, and hOB exposure to the PKA inhibitor, H89 (2x10-6M), abolished the inhibitory effect of CGRP on OPG secretion. Conditioned media from CGRP treated hOB increased the number of multinucleated tartrate resistant acid phosphatase positive cells (TRAP +) and the secretion of cathepsin K in human peripheral blood mononuclear cells (PBMCs) compared to the conditioned media of untreated hOB.
These results show that the cAMP/PKA pathway is involved in the CGRP inhibition of OPG mRNA and protein secretion in hOB and that this effect favours osteoclastogenesis. CGRP could thus modulate the balance between osteoblast and osteoclast activity, participating in the fine tuning of all of the bone remodeling phases necessary for the subsequent anabolic effect.
