Proinflammatory M1 Macrophages Inhibit RANKL-Induced Osteoclastogenesis

Abstract

In response to a defined panel of stimuli, immature macrophages can be classified into two major phenotypes: proinflammatory (M1) and anti-inflammatory (M2). Although both phenotypes have been implicated in several chronic inflammatory diseases, their direct role in bone resorption remains unclear. The present study investigated the possible effects of M1 and M2 macrophages on RANKL-induced osteoclastogenesis. In osteoclastogenesis assays using RAW264.7 cells or bone marrow cells as osteoclast precursors, addition of M1 macrophages significantly suppressed RANKL-induced osteoclastogenesis compared to nonstimulated conditions (M0), addition of M2 macrophages, or no macrophage addition (P < 0.05), suggesting that M1 macrophages can downregulate osteoclastogenesis. This effect was maintained when direct contact between M1 and osteoclast precursors was interrupted by cell culture insertion, indicating engagement of soluble factors released from M1. Macrophages which originate from monocytes not only are the key effector cells in innate immunity but also play a pivotal role in the initiation of adaptive immunity (1). It is well documented that polarized macrophages can be classified mainly into two different phenotypes: proinflammatory (M1) and antiinflammatory (M2). The production of inflammatory cytokines, such as tumor necrosis factor alpha (TNF-␣) and interleukin-6 (IL-6), by M1 macrophages promotes inflammation in the context of innate immune response, whereas the production of antiinflammatory cytokines and arginase by M2 macrophages leads to the resolution of inflammation (2). On the other hand, it is also true that osteoclasts that are engaged in bone resorption also belong to monocyte-lineage cells. Although macrophages and osteoclasts share the same precursor, macrophage colony-stimulating factor (M-CSF)-stimulated monocytes, the possible influence of macrophages, and especially the difference between M1 and M2, on osteoclastogenesis is largely unknown.Bone is a unique mineralized tissue which constantly undergoes a physiological remodeling process, and its homeostasis is achieved by the tuned balance between osteoclasts and boneforming cells (osteoblasts). As such, aberrantly promoted osteoclastogenesis is attributed to the bone destruction found in bone lytic diseases such as periodontitis and rheumatoid arthritis, which affects more than 50 million people in the United States alone (3). Of importance to this study, recent research has revealed that osteoclastogenesis is regulated by the immune system. For instance, a recent study (4) revealed that ␥␦-T cells inhibit osteoclastogenesis by their production of interferon gamma (IFN-␥), whereas B and T cells can produce RANKL under inflammatory conditions, thus working toward the promotion of osteoclastogenesis (5). However, in the context of bone lytic diseases involving chronic inflammation, such as periodontitis and rheumatoid arthritis, infiltrations of not only B and T cells but also of macrophages are observed (6, 7). It is true that macrophages are t...

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“…The sense and antisense primers used in these experiments are shown in Table III. Fold changes of the gene of interest were calculated using the ΔΔCt method and either β-actin (for human transcripts) (40) or GAPDH (for mouse transcripts) (41) as a reference (42). …”

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