Interleukin-4 (IL-4) is an important immune regulatory protein that possesses potent anti-osteoclastogenic properties, and does so via the transcription factor STAT6. Previous studies have shown that IL-4 selectively blocks RANKL-induced activation of NF-κB and mitogen-activated protein kinase (MAPK) pathway molecules, suggesting that the cytokine arrests osteoclastogenesis by blockade of these signaling cascades. However, the fact that the inhibitory effect on these pathways requires prolonged IL-4 pretreatment, and that the cytokine fails to exert an anti-osteoclastogenic effect after short-term pre-exposure of RANKL to osteoclast precursors, suggests that an additional, more immediate mechanism may also be involved. In this study, we found that simultaneous exposure of IL-4 did not alter RANKL-dependent activation of NF-κB or MAPKs, whereas the cytokine did block RANKL-induced nuclear factor activated T cells c1 (NFATc1), a master osteoclastogenic transcription factor. This inhibitory effect of IL-4 required STAT6, consistent with its functional role in osteoclastogenesis. In addition, the cytokine also partially impaired RANKL-stimulated bone resorption. Furthermore, IL-4 suppressed expression of RANKL-induced osteoclast specific genes in a STAT6-dependent manner, but failed to do so when osteoclast precursors were pre-exposed to RANKL. Thus, we provide the first evidence that IL-4 inhibits osteoclast formation by inhibiting RANKL induction of NFATc1 via STAT6 as an early event, in addition to its suppression of other signaling pathways. The inhibitory effect is ultimately regulated at the gene expression transcriptional level. © 2011 Wiley Periodicals, Inc.