Glucocorticoids induce apoptosis in primary rat osteoblasts and in mice in vivo: Prevention by estrogen

Date of Completion

January 1998


Biology, Molecular|Biology, Cell|Health Sciences, Medicine and Surgery




The ability of glucocorticoids to induce apoptosis, and estrogen to prevent glucocorticoid-induced apoptosis was studied both in vitro and in vivo. In primary rat osteoblasts treated with glucocorticoids for 72 h, a dose-dependent increase in the number of apoptotic cells was seen by acridine orange/ethidium bromide staining with the maximal response of 31 + 2% with 100 nM corticosterone. Glucocorticoid-induced DNA fragmentation was confirmed by DNA laddering. Western blots demonstrated a decrease in Bcl-2 and an increase in Bax protein levels as early as 24 h. The Bcl-2/Bax ratio decreased from 1 in control cells to 0.18 in osteoblasts treated with 1000 nM corticosterone for 72 h. There was a reduction in the levels of retinoblastoma protein and proliferating nuclear antigen in osteoblasts treated with glucocorticoids. In addition, there was a dose-dependent increase in p53 protein level following glucocorticoid treatment. Simultaneous administration of 0.01 nM 17$\beta$-estradiol and 100 nM corticosterone decreased apoptotic osteoblasts by 60%. The reduction in Bcl-2/Bax by corticosterone was abolished by treating osteoblasts simultaneously with estrogen. In seven day old mice, administration of 0.3 mg/kg dexamethasone and 1 mg/kg dexamethasone resulted in apoptosis of osteoblasts as demonstrated by DNA laddering and in situ staining of the calvaria. Injection of 5 mg/kg 17$\beta$-estradiol with 1 mg/kg dexamethasone blocked dexamethasone-induced apoptosis at 72 h as determined by TUNEL and DNA laddering. In osteoblasts, glucocorticoid-induced apoptosis may be prevented, at least in part, by estrogen. ^