Recent evidence suggests that the bone anabolic effects of the active hormone 1a, 25(OH) 2 D 3 ] are mediated via a direct action on cells of the osteoblastic lineage.1) In a number of studies, however, the doses required for anabolic effects were associated with toxicity due to hypercalcaemia and hypercalciuria.2,3) Previous studies have reported the syntheses of a number of analogs of 1a,25(OH) 2 D 3 , and these analogs can mimic the effects of the parent compound without causing hypercalcaemia.4) As such, these analogs may prove to be useful as potential therapeutic agents for the treatment for osteoporosis 5,6) and other metabolic bone diseases associated with bone loss. 7)The mechanisms of action of 1a,25(OH) 2 D 3 are far from clear although its effects on osteoblastic function have been well-documented. 4,8) Briefly, 1a,25(OH) 2 D 3 has been shown to affect alkaline phosphatase (ALP) activity, 9-13) the production, 14,15) and gene expression 16,17) of osteocalcin (OC) and insulin-like growth factor-1, 18,19) cell proliferation, 9,20) collagen synthesis 9,21,22) and expression, 16,17) parathyroid hormone (PTH)-stimulated adenylate cyclase activity, 8,23) as well as the regulation of its own receptor levels.14,24-26) The effects of 1a,25(OH) 2 D 3 have been reported to be dependent upon the stage of osteoblast maturation. 8,10,12,13) On the other hand, while the actions of several analogs have been well documented on calcium mobilization in mice or chicks 4,27) and on cell differentiation and proliferation in leukemic, 28,29) prostate, 30) and breast cancer 31) cells and human retinoblastoma tumors, 32) little is known of its effects on bone cells including the bone-forming osteoblasts. We recently showed that differences exist in the actions of 1a,25(OH) 2 D 3 and the analog 1a,25(OH) 2 -16-ene-23-yne-D 3 in the osteosarcoma SaOS-2 cells. 33) In the present report, our objective was to investigate the actions of another analog, 1a,25(OH) 2 -16-ene-D 3 , on osteoblast function. In addition, we wanted to compare its effects with those of the parent compound and their modulation by 17b -estradiol (E 2 ) in our two human cell models representing two stages of differentiation, the more differentiated cells grown in the presence of dexamethasone (DEX), the normal human osteoblasts HOBϩDEX and osteosarcoma SaOSϩDEX cells, and the corresponding less differentiated cells grown in the absence of DEX, the HOB-DEX and SaOS-DEX cells.