Nicole Joy Horwood

IL-12 Alone and in Synergy with IL-18 Inhibits Osteoclast Formation In Vitro

IL-12, like IL-18, was shown to potently inhibit osteoclast formation in cultures of cocultures of murine osteoblast and spleen cells, as well as in adult spleen cells treated with M-CSF and receptor activator of NF-κB ligand (RANKL). Neither IL-12 nor IL-18 was able to inhibit RANKL-induced osteoclast formation in cultured RAW264.7 cells, demonstrating that IL-12, like IL-18, was unable to act directly on osteoclastic precursors. IL-12, like IL-18, was found to act by T cells, since depletion of T cells from the adult spleen cell cultures ablated the inhibitory action of IL-12 and addition of either CD4 or CD8 T cells from C57BL/6 mice to RANKL-stimulated RAW264.7 cultures permitted IL-12 or IL-18 to be inhibitory. Additionally, IL-12 was still able to inhibit osteoclast formation in cocultures with osteoblasts and spleen cells from either GM-CSF R−/− mice or IFN-γ R−/− mice, indicating that neither GM-CSF nor IFN-γ was mediating osteoclast inhibition in these cultures. Combined, IL-18 and IL-12 synergistically inhibited osteoclast formation at concentrations 20- to 1000-fold less, respectively, than when added individually. A candidate inhibitor could not be demonstrated using neutralizing Abs to IL-4, IL-10, or IL-13 or from mRNA expression profiles among known cytokine inhibitors of osteoclastogenesis in response to IL-12 and IL-18 treatment, although the unknown inhibitory molecule was determined to be secreted from T cells.

Fibroblastic stromal cells express receptor activator of NF-kappa B ligand and support osteoclast differentiation.

Osteoclast formation in bone is supported by osteoblasts expressing receptor activator of NF‐κB ligand (RANKL) and macrophage colony‐stimulating factor (M‐CSF) expression. Numerous osteotropic factors regulate expression levels of RANKL and the RANKL decoy receptor osteoprotegerin (OPG) in osteoblasts, thereby affecting osteoclast differentiation. However, not only is RANKL widely expressed in soft tissues, but osteoclasts have been noted in extraskeletal lesions. We found that cultured skin fibroblastic cells express RANKL, M‐CSF, and OPG messenger (mRNA). Stimulation by 1α,25 dihydroxyvitamin D3 [1,25(OH)2D3] plus dexamethasone (Dex) augmented RANKL and diminished OPG mRNA expression in fibroblastic cells and caused the formation of numerous osteoclasts in cocultures of skin fibroblastic cells with hemopoietic cells or monocytes. The osteoclasts thus formed expressed tartrate‐resistant acid phosphatase (TRAP) and calcitonin (CT) receptors and formed resorption pits in cortical bone. Osteoclast formation also was stimulated (in the presence of Dex) by prostaglandin E2 (PGE2), interleukin‐11 (IL‐11), IL‐1, tumor necrosis factor‐α (TNF‐α), and parathyroid hormone‐related protein (PTHrP), factors which also stimulate osteoclast formation supported by osteoblasts. In addition, granulocyte‐macrophage‐CSF (GM‐CSF), transforming growth factor‐β (TGF‐β), and OPG inhibited osteoclast formation in skin fibroblastic cell‐hemopoietic cell cocultures; CT reduced only osteoclast nuclearity. Fibroblastic stromal cells from other tissues (lung, respiratory diaphragm, spleen, and tumor) also supported osteoclast formation. Thus, RANKL‐positive fibroblastic cells in extraskeletal tissues can support osteoclastogenesis if osteolytic factors and osteoclast precursors are present. Such mesenchymally derived cells may play a role in pathological osteolysis and may be involved in osteoclast formation in extraskeletal tissues.

Secreted frizzled-related protein-1 inhibits RANKL-dependent osteoclast formation.

We determined that sFRP‐1 mRNA was differentially expressed by osteoblast/stromal cell lines and that sFRP‐1 neutralizing antibodies and siRNA complementary to sFRP‐1 coding sequence enhanced, while recombinant sFRP‐1 inhibited, osteoclast formation. In studying the mechanism of action for sFRP‐1, we found that sFRP‐1 could bind recombinant RANKL. These results suggest potential cross‐talk between Wnt and RANKL pathways.

Interleukin-18: perspectives on the newest interleukin.

Just over two years ago the newest member of the interleukin family of cytokines, IL-18, was molecularly cloned. IL-18 was originally identified as a result of its ability to induce interferon gamma production, however with the advent of its cloning and the production of recombinant protein a number of other biological actions have since been identified. Recently the receptor for IL-18 was also characterised. Due to the structural and biological properties shared between IL-18 and IL-1 and their respective receptors, questions relating to IL-18 activities are being answered at a rapid pace. This article addresses the biology of IL-18 in both disease and non-disease states.

T-cells mediate an inhibitory effect of interleukin-4 on osteoclastogenesis.

IL‐4 is an important cytokine that can influence bone. We identified two distinct actions of IL‐4 to inhibit osteoclast formation: one direct on osteoclast progenitors and the second through the production of a novel T‐cell surface‐associated molecule(s). These data show a new link between the immune system and bone.