Extracellular signal-regulated kinases and calcium channels are involved in the proliferative effect of bisphosphonates on osteoblastic cells in vitro

Bisphosphonates (BPs) are analogues of pyrophosphate, which are widely used for the treatment of different pathologies associated with imbalances in b one turnover. Recent evidence suggested that cells of the osteoblastic line age might be targets of the action of BPs. The objective of this work was t o determine whether BPs induce proliferation of osteoblasts and whether thi s action involves activation of the extracellular signal-regulated kinases (ERKs). We have shown that three different BPs (olpadronate, pamidronate, a nd etidronate) induce proliferation in calvaria-derived osteoblasts and ROS 17/2.8 as measured by cell count and by [H-3]thymidine uptake. Osteoblast proliferation induced by all BPs diminished to control levels in the presen ce of U0126, a specific inhibitor of the upstream kinase MEK 1 responsible for ERK phosphorylation. Consistent with this, BPs induced ERK activation a s assessed by in-gel kinase assays. Phosphorylation of ERK1/2 was induced b y the BPs olpadronate and pamidronate within 30 s, followed by rapid dephos phorylation, whereas etidronate induced phosphorylation of ERKs only after 90 s of incubation and returned to basal levels within 15-30 minutes. In ad dition, both BP-induced cell proliferation and ERK phosphorylation were red uced to basal levels in the presence of nifedipine, an L-type voltage-sensi tive calcium channel (VSCC) inhibitor. These results show that BP-induced p roliferation of osteoblastic cells is mediated by activation of ERKs and su ggest that this effect requires influx of Ca2+ from the extracellular space through calcium channels.