Estrogen prevents glucocorticoid-induced apoptosis in osteoblasts in vivo and in vitro

The ability of estrogen to prevent glucocorticoid-induced apoptosis in osteoblasts was studied both in vitro and in vivo. Glucocorticoid treatment for 72 h produced a dose-dependent increase in the number of apoptotic cells, determined by acridine orange/ethidium bromide staining, with a maximal response of 31 +/- 2% and 26 +/- 3% with 100 nM corticosterone in primary rat and mouse osteoblasts, respectively. Simultaneous administration of varying concentrations of 17 beta-estradiol and 100 nM corticosterone decreased apoptotic osteoblasts in a dose-dependent manner, with a maximal decrease of 70% with 0.01 nM 17 beta-estradiol. Terminal deoxynucleotidyltransferase-mediated deoxy-UTP-biotin nick end labeling also demonstrated glucocorticoid-induced DNA fragmentation that was inhibited by estrogen. Estrogen was shown to inhibit apoptosis induced by lipopolysaccharide treatment. As early as 6 h, Western blots demonstrated a dose-dependent decrease in the Bcl-2/Bax ratio, which reached a minimum of 0.18 in osteoblasts treated with 1000 nM corticosterone for 72 h. This reduction in Bcl-2/Bax was abolished by treating osteoblasts simultaneously with 17 beta-estradiol, but not with 17 alpha-estradiol. In 7-day-old mice, administration of varying concentrations of dexamethasone for 72 h resulted in a dose-dependent increase in the number of apoptotic osteoblasts as demonstrated by in situ terminal deoxynucleotidyltransferase-mediated deoxy-UTP-biotin nick end labeling staining of calvaria. A maximum of 22 +/- 1% apoptotic osteoblasts on the bone surface was found with 1 mg/kg BW dexamethasone compared with 2 +/- 1% in vehicle-treated mice. Injection of varying concentrations of 17 beta-estradiol (0.5-5 mg/kg BW), but not 17 alpha-estradiol, with I mg/kg dexamethasone produced a dose-dependent decrease in the number of apoptotic osteoblasts to 5 +/- 1% with 5 mg/kg 17 beta-estradiol. Thus, glucocorticoid-induced apoptosis of osteoblasts may be prevented at least in part by 17 beta-estradiol.