title: Changing RANKL/OPG mRNA expression in differentiating murine primary osteoblasts.
abstract: Osteoblast-osteoclast coordination is critical in the maintenance of skeletal integrity. The modulation of osteoclastogenesis by immature cells of the osteoblastic lineage is mediated through receptor activator of NF kappa B (RANK), its ligand RANKL, and osteoprotegerin (OPG), a natural decoy receptor for RANKL. Here, the expression of OPG and RANKL in primary mouse osteoblastic cultures was investigated to determine whether the osteoclastogenic stimulus depended on the stage of osteoblastic differentiation and the presence of the calciotrophic hormone 1,25-dihydroxyvitamin D(3) (1,25-(OH)(2)D(3)). OPG mRNA expression was increased in osteoblastic cultures after the onset of mineralisation relative to less mature cultures, but did not alter in response to 1,25-(OH)(2)D(3) treatment. In contrast, basal RANK L mRNA expression did not change during differentiation but was significantly enhanced by 1,25-(OH)(2)D(3) treatment at all times. The stimulatory effects of 1,25-(OH)(2)D(3) on RANKL were lessened in more mature cultures, however. The RANKL/OPG ratio, an index of osteoclastogenic stimulus, was therefore increased by 1,25-(OH)(2)D(3) treatment at all stages of osteoblastic differentiation, but to a lesser degree in cultures after the onset of mineralisation. Thus the 1,25-(OH)(2)D(3)-driven increase in osteoclastogenic potential of immature osteoblasts appears to be mediated by increased RANKL mRNA expression, with mature osteoblasts having relatively decreased osteoclastogenic activity due to increased OPG mRNA expression. These findings suggest a possible mechanism for the recently proposed negative regulatory role of mature osteoblasts on osteoclastogenesis and indicate that the relative proportions of immature and mature osteoblasts in the local microenvironment may control the degree of resorption at each specific bone site.
authors.affiliation: Bone and Mineral Research Program, Garvan Institute of Medical Research, Darlinghurst, New South Wales 2010, Australia. g.thomas@garvan.org.au
authors.forename: G P
authors.initials: GP
authors.lastname: Thomas
authors.forename: S U
authors.initials: SU
authors.lastname: Baker
authors.forename: J A
authors.initials: JA
authors.lastname: Eisman
authors.forename: E M
authors.initials: EM
authors.lastname: Gardiner
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D002117:Calcitriol / Q000494:pharmacology; D002352:Carrier Proteins / Q000235:genetics*; D002454:Cell Differentiation; D002478:Cells, Cultured; D015870:Gene Expression / Q000187:drug effects; D006023:Glycoproteins / Q000235:genetics*; D008562:Membrane Glycoproteins / Q000235:genetics*; D051379:Mice; D008815:Mice, Inbred Strains; D010006:Osteoblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D053244:Osteoprotegerin; D053245:RANK Ligand; D012333:RNA, Messenger / Q000032:analysis*; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics*; D018124:Receptors, Tumor Necrosis Factor; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D002352:Carrier Proteins; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D053244:Osteoprotegerin; D053245:RANK Ligand; D012333:RNA, Messenger; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear; D018124:Receptors, Tumor Necrosis Factor; C505687:Tnfrsf11a protein, mouse; C506031:Tnfrsf11b protein, mouse; C505689:Tnfsf11 protein, mouse; D002117:Calcitriol