Regulation of intracellular Ca(2+) by reactive oxygen species in osteoblasts treated with antimycin A

This study evaluated the effects of antimycin A (AMA), an inhibitor of electron transport in mitochondria, on the release of intracellular calcium ion ([Ca(2+) ](i) ), ROS and bone resorbing factors in osteoblastic MC3T3-E1 cells. Pretreatment of osteoblasts with trolox, a ROS scavenger, and cyclosporin A, a potent inhibitor of calcium release from mitochondria, prevented the AMA-induced increases in [Ca(2+) ](i) . However, [Ca(2+) ](i) increase by AMA was unaffected by dantrolene, which blocks the ryanodine receptor channel of the endoplasmic reticulum. BAPTA/AM (an intracellular Ca(2+) chelator), dantrolene and cyclosporine A did not reverse the effect of AMA on ROS release. We also investigated whether intracellular calcium release inhibitor and antioxidant protect against AMA-induced bone resorbing cytokine release. Trolox prevented the release of receptor activator of nuclear factor-κB ligand (RANKL), IL-6, and TNF-α induced by AMA. Moreover, the increased IL-6 and TNF-α release by AMA was markedly reduced by BAPTA/AM and cyclosporin A. However, BAPTA/AM did not reverse the effect of AMA on osteoprotegerin and RANKL. Taken together, these results demonstrate that mitochondrial ROS generation and Ca(2+) influx by AMA is required for osteoblast death and bone resorbing cytokine release.