Chen Y C, Chen S J, Chang H T, Huang J K, Wang J L, Tseng L L, Chang H J, Su W, Law Y P, Chen W C, Jan C R
Department of Orthopaedic Surgery, Chang-Gung Memorial General Hospital, Kaohsiung, Taiwan.
Toxicol Lett. 2001 Jul 6;122(3):245-53. doi: 10.1016/s0378-4274(01)00370-8.
The effect of the estrogen diethylstilbestrol (DES) on cytosolic free Ca(2+) levels (Ca(2+)) in MG63 human osteoblasts was explored by using fura-2 as a Ca(2+) indicator. DES at concentrations between 5--20 microM induced an immediate increase in Ca(2+) in a concentration-dependent manner with an EC(50) of 10 microM. Removing extracellular Ca(2+) reduced the Ca(2+) signal by 70%. Pretreatment with 50 microM La(3+) or 10 microM of nifedipine, verapamil and diltiazem did not change 20 microM DES-induced Ca(2+) increases. Addition of 3 mM Ca(2+) increased Ca(2+) in cells pretreated with 20 microM DES in Ca(2+)-free medium. Pretreatment with 1 microM thapsigargin (an endoplasmic reticulum Ca(2+) pump inhibitor) to deplete the endoplasmic reticulum Ca(2+) store partly inhibited 20 microM DES-induced Ca(2+) release, but addition of carbonylcyanide m-chlorophenylhydrazone (CCCP; a mitochondrial uncoupler) and thapsigargin together abolished DES-induced Ca(2+) release. Conversely, pretreatment with 20 microM DES abrogated CCCP- and thapsigargin-induced Ca(2+) release. Inhibition of phospholipase C activity with 2 microM U73122 did not alter 20 microM DES-induced Ca2+ release. Another estrogen 17beta-estradiol also increased Ca(2+) in a concentration-dependent manner with an EC50 of 7 microM. Together, the data indicate that in human osteoblasts, DES increased Ca(2+) via causing Ca(2+) release from both mitochondria and the endoplasmic reticulum in a phospholipase C-independent manner, and by causing Ca(2+) influx.
