Regulation of osteocalcin secretion by human primary bone cells and by the human osteosarcoma cell line MG-63

We present evidence that the regulation of osteocalcin secretion by PTH and PGE₂ in normal human bone cells can be produced in the human osteoblast-like cell line MG-63. Both cell cultures showed time-and dose-dependent stimulation of osteocalcin secretion in response to 1,25(OH)₂D₃. Bovine parathyroid hormone (PTH) amino acid fragment 1-34 (40 nM) and prostaglandin E₂ (PGE₂, 5 nM) significantly inhibited 1,25(OH)₂D₃-induced osteocalcin secretion by these cells. The inhibition reached 20 and 36%, respectively. In contrast, PTH 3-34 had no effect on osteocalcin secretion. Both cell cultures produced cAMP in response to PTH. Dexamethasone (Dex) (100 nM) potentiated PTH-induced (40 nM) cAMP synthesis in subconfluent MG-63 cells (1.5-fold increase, P < 0.05). This treatment with Dex resulted in a greater inhibition of 1,25(OH)₂D₃-induced osteocalcin secretion (-30%, P < 0.005) by PTH in MG-63 cells as compared to cells exposed to PTH and 1,25(OH)₂D₃ alone. Pretreatment of subconfluent MG-63 cells with Dex (100 nM) for 48 h also increased 1,25(OH)₂D₃-induced osteocalcin secretion by 40% (P < 0.025). In contrast, treatments of confluent MG-63 cells with Dex inhibited osteocalcin secretion regardless of the 1,25(OH)₂D₃ doses used. Forskolin (10^-7-10^-5 M) and dibutyryl cAMP (10^-6-10^-3 M) both reproduced the effects observed with PTH and PGE₂ in the two cell cultures. Forskolin's action was time-dependent: addition of forskolin (10^-6 M) 12 h after 1,25(OH)₂D₃ (50 nM) resulted in a progressively weaker inhibition of osteocalcin secretion. Increasing the extracellular calcium concentration of the incubation media resulted in a dose-dependent increase in osteocalcin secretion (P < 0.01). These results indicate that PTH and PGE₂ inhibit osteocalcin secretion by a mechanism involving cAMP production. In contrast, an increase in extracellular calcium stimulated osteocalcin release. Thus the human osteosarcoma cell line MG-63 is a useful osteoblast-like cell model to study the regulation of osteocalcin secretion. Furthermore, a factor (or factors) between hormone-receptor coupling and gene induction can regulate the expression of the osteocalcin gene or affect pre- or posttranslational mechanisms implicated in osteocalcin synthesis and secretion.