title: Evidence for multiple bone resorption-stimulating factors produced by normal human keratinocytes in culture.
abstract: Conditioned medium from cultured normal human foreskin keratinocytes enhanced the release of calcium from neonatal mouse calvaria in organ culture. Unfractionated keratinocyte-conditioned medium (KCM) stimulated bone resorption in a dose-dependent manner, but it did not increase the concentration of prostaglandin E2 (PGE2) in the bone culture medium until a maximal dose of KCM for resorption was used. Furthermore, inhibitors of PGE2 synthesis, indomethacin, ibuprofen, and piroxicam, did not inhibit KCM-induced calcium release. High concentrations of KCM increased cAMP production by calvaria in the presence of isobutylmethylxanthine, but the increase was small compared with that produced by a dose of bovine PTH that caused a similar level of bone resorption. The bone resorption-stimulating activity of KCM was not lost after incubation at 56 C for 60 min, but it was lost after heating at 100 C for 10 min. Fractionation of KCM by gel filtration chromatography revealed two distinct peaks of bone resorption-stimulating activity. One peak, KCMI, caused a significant increase in bone resorption at 2 micrograms protein/ml. KCMI did not increase medium PGE2, and inhibition of PGE2 synthesis in bone had no effect on KCMI-induced bone resorption. KCMI failed to increase cAMP production by human osteosarcoma SaOS-2 cells. Another peak, KCMII, caused a dose-dependent increase in bone resorption, and a significant increase in medium calcium was noted at a 20-fold lower concentration (0.1 microgram protein/ml) than with KCMI. In contrast to KCMI, the increase in bone resorption stimulated by KCMII was accompanied by a parallel increase in the production of PGE2, and inhibition of PGE2 synthesis completely inhibited the bone resorption-stimulating activity of KCMII. KCMII also caused an increase in cAMP production by SaOS-2 cells. We conclude that KCM contains at least two distinct bone resorption-stimulating factors, one of which acts via a PG-mediated mechanism and the other by a PG-independent pathway.
mesh_terms: D000818:Animals; D001681:Biological Assay; D001862:Bone Resorption / Q000187:drug effects*; D001842:Bone and Bones / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D002850:Chromatography, Gel; D003470:Culture Media / Q000032:analysis; D000242:Cyclic AMP / Q000096:biosynthesis / Q000494:pharmacology; D015232:Dinoprostone; D004817:Epidermis / Q000378:metabolism*; D006801:Humans; D007213:Indomethacin / Q000494:pharmacology; D007633:Keratins / Q000378:metabolism*; D051379:Mice; D009924:Organ Culture Techniques; D012516:Osteosarcoma / Q000378:metabolism; D010281:Parathyroid Hormone / Q000494:pharmacology; D011458:Prostaglandins E / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D003470:Culture Media; D010281:Parathyroid Hormone; D011458:Prostaglandins E; D007633:Keratins; D000242:Cyclic AMP; D015232:Dinoprostone; D007213:Indomethacin