The balance between bone formation and bone resorption in inflammatory diseases is often disturbed. Periodontitis, a chronic inflammation of the tooth gums, leads to unwanted bone loss as a response to inflammatory compounds such as interleukin-1β (IL-1β). This excessive bone loss reflects an increased osteoclast formation and activity. Osteoclast formation is a multistep process driven by osteoclastogenesis supporting cells such as periodontal ligament fibroblasts. The inflammatory factors can induce osteoclastogenesis, probably also by affecting the periodontal ligament fibroblast. In this study we investigated how pre-culture of periodontal ligament fibroblasts with IL-1β affected osteoclastogenesis. Fibroblasts were pre-cultured with IL-1β and/or dexamethasone, a commonly used anti-inflammatory compound, before being co-cultured with peripheral blood mononuclear cells (PBMCs). Pre-culture with IL-1β (1-100 ng/ml) resulted in an increased number of adhered PBMCs as well as an increased mRNA expression of intercellular adhesion molecule-1 (ICAM-1), macrophage colony stimulating factor (M-CSF) and IL-1β. Pre-culture with IL-1β also caused retraction of fibroblasts and an augmented formation of TRACP(+) multinucleated cells. Our data suggest that stimulation of fibroblasts with IL-1β has a long-lasting effect, leading to a significantly increased osteoclastogenesis. These results provide new insights for understanding excessive bone loss in periodontitis.