Vip Inhibits Osteoclast-like Cell Formation in Mouse Bone Marrow Culture

Objective: There are evidences those indicate neuropeptide, including vasoactive intestinal peptide (VIP), regulation of bone metabolism. In the present study, the effect of VIP on osteoclast formation in mouse bone marrow cultures was examined. Methods: Femur and tibia from 5 to 9 weeks old CsA male mice were aseptically excised and bone marrow tissues were flushed out and cultured. After 7-day-culture-period, 1) cells were fixed and stained to count tartrate acid phosphatase positive multinuclear cells (TRAP+MNC), 2) cells were incubated with 125I-calcitonin (CT) to evaluate the number of calcitonin biding site and 3)RNA was extracted from the culture and processed to RT-PCR. Results: VIP reduced, in a concentration-dependent manner, osteoclast formation, the number of binding sites for radiolabelled calcitonin and the amounts of resorption pits in cultures stimulated by PTH or D3. Osteoclast formation stimulated by D3 was associated with stimulations of mRNA for calcitonin receptor, cathepsin K and TRAP. The stimulatory effect by D3 on calcitonin receptor and cathepsin K mRNA, but not that on TRAP mRNA, was down regulated by VIP. The inhibition of TRAP+MNC formation by VIP was mimicked by forskolin. cAMP concentration of VIP or forskolin treated culture is higher that of D3 stimulated culture The presence of VIP during last two days of a 7-day-culture-period was sufficient to inhibit osteoclast formation. Not only VIP, but also PACAP-38 and secretin, but not glucagon, inhibited osteoclast formation and the number of calcitonin binding sites in D3 stimulated cultures. RT-PCR revealed the expression of VIP-1 and VIP-2 receptor subtypes with the VIP-2 receptor mRNA down regulated by D3. These findings indicate that VIP, via VIP-1 and/or VIP-2 receptors, inhibits osteoclastogenesis by cAMP-dependent mechanism, due to an inhibitory effect at the late stage of osteoclastogenesis. Conclusion: The present study supports the idea of a neuro-osteogenic interaction in bone metabolism.