Tumor Necrosis Factor α Regulatesα vβ5 Integrin Expression by Osteoclast Precursors in Vitro and in Vivo1.

Early osteoclast precursors, in the form of murine bone marrow macrophages (BMMs), while expressing no detectable avb3 integrin, contain abundant avb5 and attach to matrix in an av integrin-dependent manner. Furthermore, avb5 expression by osteoclast precursors progressively falls as they assume the resorptive phenotype. We find the osteoclastogenic agent, tumor necrosis factor-a, (TNF) down-regulates avb5 expression by BMMS via attenuation of b5 messenger RNA (mRNA) t1/2. Using BMMs from TNF receptor knockout mice we establish the p55 receptor transmits the b5 suppressive effect. The functional implications of TNF-mediated avb5 down-regulation are underscored by the capacity of an av inhibitory peptide mimetic to prevent spreading by BMMs expressing abundant avb5 while failing to impact those in which the integrin has been diminished by TNF. Finally, b5 mRNA in BMMs of wild-type mice administered lipopolysaccharide (LPS) progressively falls with time of in vivo treatment. Alternatively, b5 mRNA does not decline in BMMs of LPS-treated mice lacking both TNF receptors, documenting down-regulation of the b5 integrin subunit, in vivo, is mediated by TNF. Thus, matrix attachment of osteoclast precursors and mature osteoclasts are governed by distinct av integrins which are differentially regulated by specific cytokines. (Endocrinology 141: 284–290, 2000) O are physiological polykaryons, and the principal if not exclusive resorptive cells of bone. They are derived from macrophage precursors residing in diverse tissues, principally marrow (1). While the precise means by which osteoclast precursors undergo differentiation are incompletely defined, attachment to matrix is probably essential. For example, macrophage polykaryons, including osteoclasts, fail to develop in suspension culture (Teitelbaum, S. L., unpublished observation) but form in abundance when in contact with matrix. Thus, the capacity of osteoclast precursors to recognize and attach to matrix is pivotal to generation of the resorptive phenotype. Integrins, which are major negotiators of cell matrixattachment, are transmembrane heterodimers consisting of noncovalently linked a and b subunits. Given the importance of matrix recognition to osteoclast recruitment and function, attention has turned to identifying integrins that mediate these events. These experiments establish avb3 as an integrin essential to osteoclast function. For example, antibody blockade of avb3 blunts bone resorption in vitro and in vivo (2–4), and the b3 integrin knockout mouse develops osteosclerosis due to osteoclast failure (5). Given the abundance of avb3 on multinucleated osteoclasts, it seems likely the integrin participates in bone degradation by the mature cell. Consistent with the this posture, avb3 is progressively expressed as osteoclast precursors differentiate in culture (6). Moreover, cytokines and steroids that impact osteoclast differentiation (6) accelerate appearance of avb3 on osteoclast precursors and do so by regulating the b3 subunit (6, 7). While these observations are in keeping with the hypothesis that avb3 plays a pivotal role in the matrix degrading capacity of the mature polykaryon, they fail to address the means by which mononuclear osteoclast precursors recognize and attach to matrix. In this regard, we find avb3 undetectable on early, marrow-derived osteoclast precursors, despite their capacity to bind to extracellular matrix protein recognized by this integrin (6). This conundrum is a reflection of the fact these avb3 negative cells express the closely related integrin avb5, suggesting the latter heterodimer mediates osteoclast precursor-matrix recognition. In fact, as marrow macrophages assume the osteoclast phenotype in culture, avb5 disappears pari passu with emergence of avb3 (6). Given osteoclastogenesis is attended by diminution of avb5, one would expect agents that accelerate osteoclastogenesis to down-regulate the integrin. Tumor necrosis factor (TNF) is a proinflammatory cytokine with potent bone resorptive capacity, exerting its effect by enhancing osteoclast recruitment (8–11). We find this cytokine hastens disappearance, on osteoclast precursors, of avb5, a reflection of accelerated degradation of b5 subunit mRNA. The TNF effect on the b5 integrin is mediated through the p55TNF receptor and, reflecting the in vitro situation, the TNF agonist lipopolysaccharide (LPS), when administered in vivo, rapidly diminishes marrow macrophage b5 mRNA. Materials and Methods