Thiol oxidation by 2,2′-dithiodipyridine induced calcium mobilization in MG63 human osteosarcoma cells

2,2'-dithiodipyridine (2,2'-DTDP), a reactive disulphide that mobilizes Ca2+ in muscle, induced an increase in cytoplasmic free Ca2+ concentrations ([Ca2+](i)) in MG63 human osteosarcoma. cells loaded with the Ca2+-sensitive dye fura-2. 2,2'-DTDP acted in a concentration-independent manner with an EC50 of 50 [M. The Ca2+ signal comprised an initial spike and a prolonged increase. Removing extracellular Ca2+ did not alter the Ca2+ signal, suggesting that the Ca2+ signal was due to store Ca2+ release. In Ca2+-free medium, the 2,2-DTDP-induced [Ca2+](i) increase was not changed by depleting store Ca2+ with 50 muM breffeldin A (a Golgi apparatus permeabilizer), 2 muM carbonylcyanide m-chlorophenylhydrazone (CCCP, a mitochondrial uncoupler), I [muM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor) or 5 muM ryanodine. Conversely, 2,2'-DTDP pretreatment abolished CCCP and thapsigargin-induced [Ca2+](i) increases. 2,2'-DTDP-induced Ca2+ signals in Ca2+-containing medium were not affected by modulation of protein kinase C activity or suppression of phospholipase C activity. However, 2,2'-DTDPinduced Ca2+ release was inhibited by a thiol-selective reducing reagent, dithiothreitol (5-25 muM) in a concentration-dependent manner. Collectively, this study shows that 2,2'-DTDP induced [Ca2+](i) increases in human osteosarcoma. cells via releasing store Ca2+ from multiple stores in a manner independent of protein kinase C or phospholipase C activity. The 2,2'-DTDP-induced store Ca2+ release appeared to be dependent on oxidation of membranes. (C) 2002 Elsevier Science Inc. All rights reserved.