Tumor necrosis factor-alpha (TNF) stimulates RANKL-induced osteoclastogenesis via coupling of TNF type 1 receptor and RANK signaling pathways

Tumor necrosis factor-cw (TNF) and the ligand for receptor activator of NF- KB (RANKL) are abundant in sites of inflammatory bone erosion. Because thes e cytokines are potent osteoclastogenic factors and because their signaling pathways are considerably overlapping, we postulated that under pro-inflam matory conditions RANKL and TNF might synergistically orchestrate enhanced osteoclastogenesis via cooperative mechanisms. We found TNF, via TNF type 1 receptor (TNFr1), prompts robust osteoclastogenesis by osteoclast precurso rs pretreated with RANKL, and deletion of TNFrl abrogates this response. En hanced osteoclastogenesis is associated with high expression of otherwise T NF and RANKL-induced mediators, including c-Src, TRAF2, TRAF6, and MEKK-1, levels of which were notably reduced in TNFrl knockouts. Recruitment of TRA Fs and MEKK1 leads to activation of downstream pathways, primarily I kappaB /NF-kappaB, ERKs, and cJun/AP-1. Consistent with impaired osteoclastogenesi s and reduced expression of TRAFs and MEKK1, we found that phosphorylation and activation of I kappaB, NF-kappaB, ERKs, and cJun/AP-1 are severely red uced in RANKL-treated TNFr1-null osteoclast precursors compared with wild t ype counterparts. Finally, we found that TNF and RANKL synergistically up-r egulate RANK expression in wild type precursors, whereas basal and stimulat ed levels of RANK are significantly lower in TNFrl knockout cells. Our data suggest that exuberant TNF-induced osteoclastogensis is the result of coup ling between RANK and TNFrl and is dependent upon signals transmitted by th e latter receptor.