High doses of bisphosphonates reduce osteoblast-like cell proliferation by arresting the cell cycle and inducing apoptosis

Abstract

Objectives: The study objective was to evaluate the effect on osteoblast growth of high concentrations of three nitrogen-containing bisphosphonates (pamidronate, alendronate, and ibandronate) and one non-nitrogen-containing bisphosphonate (clodronate), using the MG-63 cell line as an osteoblast model, in order to determine the role of osteoblasts in bisphosphonate-related osteonecrosis of the jaw (BRONJ).

Materials and methods: Osteoblasts were incubated in culture medium with different doses of pamidronate, alendronate, ibandronate or clodronate. The proliferative capacity of the osteoblasts was determined by spectrophotometry (MTT-based) at 24 h of culture. Flow cytometry was used to determine the percentage of cells in each cell cycle phase (G0/G1, G2/M, and S) and to discriminate apoptotic cell death from necrotic cell death in the cell cycle at 24 h of treatment.

Results: All the bisphosphonates assayed produced a significant and dose-dependent reduction in MG-63 proliferation at the high doses assayed (10(-4) and 5 × 10(-5) M) in comparison with controls (p <0.001). Cell cycle study revealed that all assayed bisphosphonates significantly arrested the cell cycle in phase G0/G1 at doses of 10(-4) and 5 × 10(-5) M, increasing the percentage of cells in this phase (p <0.05). Apoptosis/necrosis studies showed significant changes compared with control cells, with an increased percentage of cells in apoptosis after treatment with 10(-4) or 5 × 10(-5) M of pamidronate, alendronate, ibandronate, or clodronate (p <0.05).

Conclusions: High doses of nitrogen-containing or non-nitrogen-containing bisphosphonates can reduce the proliferation of MG-63 osteoblast-like cells by arresting the cell cycle and inducing apoptosis/necrosis.

Keywords: Apoptosis; BRONJ; Bisphosphonates; Cell cycle; Osteoblast; Proliferation.