DC-STAMP Is an Osteoclast Fusogen Engaged in Periodontal Bone Resorption

Abstract

Dendritic cell-specific transmembrane protein (DC-STAMP) plays a key role in the induction of osteoclast (OC) cell fusion, as well as DC-mediated immune regulation. While DC-STAMP gene expression is upregulated in the gingival tissue with periodontitis, its pathophysiological roles in periodontitis remain unclear. To evaluate the effects of DC-STAMP in periodontitis, anti-DC-STAMP-monoclonal antibody (mAb) was tested in a mouse model of ligature-induced periodontitis ( n = 6-7/group) where Pasteurella pneumotropica ( Pp)-reactive immune response activated T cells to produce receptor activator of nuclear factor kappa-B ligand (RANKL), which, in turn, promotes the periodontal bone loss via upregulation of osteoclastogenesis. DC-STAMP was expressed on the cell surface of mature multinuclear OCs, as well as immature mononuclear OCs, in primary cultures of RANKL-stimulated bone marrow cells. Anti-DC-STAMP-mAb suppressed the emergence of large, but not small, multinuclear OCs, suggesting that DC-STAMP is engaged in the late stage of cell fusion. Anti-DC-STAMP-mAb also inhibited pit formation caused by RANKL-stimulated bone marrow cells. Attachment of ligature to a second maxillary molar induced DC-STAMP messenger RNA and protein, along with elevated tartrate-resistant acid phosphatase-positive (TRAP+) OCs and alveolar bone loss. As we expected, systemic administration of anti-DC-STAMP-mAb downregulated the ligature-induced alveolar bone loss. Importantly, local injection of anti-DC-STAMP-mAb also suppressed alveolar bone loss and reduced the total number of multinucleated TRAP+ cells in mice that received ligature attachment. Attachment of ligature induced significantly elevated tumor necrosis factor-α, interleukin-1β, and RANKL in the gingival tissue compared with the control site without ligature ( P < 0.05), which was unaffected by local injection with either anti-DC-STAMP-mAb or control-mAb. Neither in vivo anti- Pp IgG antibody nor in vitro anti- Pp T-cell response and resultant production of RANKL was affected by anti-DC-STAMP-mAb. This study illustrated the roles of DC-STAMP in promoting local OC cell fusion without affecting adaptive immune responses to oral bacteria. Therefore, it is plausible that a novel therapeutic regimen targeting DC-STAMP could suppress periodontal bone loss.

OC‐STAMP promotes osteoclast fusion for pathogenic bone resorption in periodontitis via up‐regulation of permissive fusogen CD9

“…We have recently developed an anti-DC-STAMP-mAb (mouse IgG2a) that can neutralize the cell fusion event in RANKLinduced OCs (12). Anti-OC-STAMP-mAb (mouse IgG3) was generated by the methods previously reported (21)(22)(23).…”

“…Then, at 0, 24, 48, and 72 h after the addition of soluble RANKL, the cells were analyzed by flow cytometry. Cells cultured in a 25 cm 2 flask were removed by incubation with 1 mM EDTA for 1 h. After blocking FcR with anti-CD16/32 mAb (BioLegend), the cells were stained with FITC-labeled anti-DC-STAMP-mAb (15), either Alexa Fluor 546-or Alexa Fluor 647-labeled anti-OC-STAMP, phycoerythrin-labeled anti-CD9-PE-mAb (BD Pharmingen), and APC-labeled anti-CD115-mAb (BioLegend) for 1 h. OCps reacted with respective mAb were then subjected to flow cytometry analysis using the FACSAria III system (BD Biosciences, San Jose, CA, USA).…”

“…While OCps isolated from both DC-STAMPknockout (KO) and OC-STAMP-KO mice are incapable of generating mature multinucleated OCs in response to RANKL stimulation in vitro, only DC-STAMP-KO mice, but not OC-STAMP-KO mice, show the osteopetrotic bone phenotype in adult mice (5). Furthermore, anti-DC-STAMP-mAb administered to the mice with induced periodontitis showed partial suppression (20-30%) of periodontal bone resorption by down-regulating the cell fusion of OCps (15), suggesting the presence of some additional mechanism that up-regulates the fusion of OCps. Collectively, these studies indicate that each STAMP plays a distinct role in OC-genesis-mediated bone remodeling.…”

“…Among a number of bone resorptive diseases, we are particularly interested in the pathophysiology of alveolar bone resorption caused in periodontitis which is mediated by a locally elevated osteoclastogenesis (OC-genesis) factor, namely receptor activator of NF-kB ligand (RANKL) (8)(9)(10), and which affects the 46% of the adult U.S. population (11). To understand the molecular mechanisms underlying pathogenic OC-genesis relative to OCp fusion in periodontitis, we showed that DC-STAMP is engaged in OCp fusion during OC-genesis, which in turn promotes local OC differentiation and bone resorption induced in a mouse model of periodontitis (12). Although increased CD9 immunostaining patterns were demonstrated in neutrophils and epithelium in gingival crevice of patients with periodontitis (13), the possible functions of CD9 expressed by OCs in the alveolar bone of periodontal lesion are largely unknown.…”

Pressure Cycling Technology Assisted Mass Spectrometric Quantification of Gingival Tissue Reveals Proteome Dynamics during the Initiation and Progression of Inflammatory Periodontal Disease

“…There is yet no broad‐scale quantitative and integrative analysis of gingival tissue proteomes in experimental periodontitis models. As the periodontitis model allows the dissection of the cause‐and‐effect relationship between bacterial accumulation and tissue destruction in a longitudinal manner, the main goal of the present study was to examine protein expression and the respective signaling pathways during the establishment of periodontitis in mice. Collectively, our data indicated that PCT‐assisted label‐free quantitative proteomics workflow is an efficient pipeline to reproducibly dissect the gingival tissue proteome and leads to a reliable quantification and catalog of more than 1600 proteins in a 0.5 mg tissue.…”