Regulation of osteoprotegerin secretion from primary cultures of human bone marrow stromal cells

Osteoprotegerin (OPG) is a soluble receptor for receptor activator of NF ka ppaB-ligand, a factor required for osteoclastogenesis. OPG secreted from bo ne marrow stromal cells is believed to inhibit osteoclast differentiation a nd several agents known to influence bone resorption have been demonstrated to regulate mRNA levels of OPG. In this report we have investigated the se cretion of OPG protein from primary cultures of human bone marrow stromal c ells. An ELISA was developed for measuring the concentration of OPG in cult ure medium. OPG; secretion was decreased by 50% when the human bone marrow stromal cells were treated with 1 muM of prostaglandin E-2, possibly throug h activation of the protein kinase A-pathway since stimulation of protein k inase A by forskolin also inhibited OPG; secretion. Treatment with phorbol 12,13 di butyrate, an activator of the protein kinase C-pathway, potently s timulated the secretion of OPG from human bone marrow stromal cells. The ce lls were also stimulated with inflammatory mediators and glucocorticoids. T reatment with interleukin-1 alpha (IL-1 alpha) and tumor necrosis factor-al pha (TNF-alpha) stimulated OPG secretion to 500% and 400% of control wherea s dexamethasone decreased OPG production by 40%, In conclusion, an ELISA me asuring OPG in cell culture media was developed. Using this ELISA, the amou nt of OPG secreted from human bone marrow stromal cells was clearly detecta ble, and the secretion of OPG-protein was potently regulated by prostagland in E-2, forskolin, phorbol 12,13 di butyrate, IL-1 alpha, TNF-alpha, and de xamethasone. (C) 2001 Academic Press.