Over-activated osteoclasts derived from myeloid or peripheral blood monocytes by inflammatory
cytokines results in
osteoporosis,
osteoarthritis, and other bone erosion-related diseases.
Interleukin 35 (IL-35) is a novel anti-inflammatory and immunosuppressive factor. This study investigated the effect of IL-35 on TNF-α-induced osteoclastogenesis. In the presence of IL-35, this process was detected by
Tartrate-Resistant Acid Phosphatase (TRAP) staining,
F-actin staining, and
bone resorption assays. The effects of IL-35 on TNF-α-induced apoptosis were demonstrated by TUNEL staining, cell viability assays, and flow cytometry. Moreover, a microarray was performed to detect the effect of IL-35 on TNF-α-activated
phosphatase kinase. The effect of IL-35 on the TNF-α-mediated activation of NF-κB, MAPK,
TRAF2, RIP1, Fas-associated death domain (FADD), and caspase3 was further investigated. In addition, a murine calvarial
osteolysis model was established via the
subcutaneous injection of TNF-α onto the calvaria, and histological analysis was subsequently performed. As a result, IL-35 inhibited TNF-α-induced osteoclast formation and
bone resorption in vitro and
osteolysis calvaria in vivo. NFATc1, c-fos, and TRAP were downregulated by IL-35 through the inhibition of NF-κB and MAPK, during which JAK1/STAT1 was activated. Moreover, based on TUNEL staining and flow cytometry, IL-35 was shown to enhance TNF-α-induced osteoclast apoptosis. Meanwhile, FADD and cleaved-
caspase 3 were increased in cells treated with TNF-α and IL-35, whereas the
DNA-binding activity of NF-κB was increased in TNF-α-treated cells, but was decreased in cells treated with both TNF-α and IL-35. In conclusion, IL-35 inhibits TNF-α-induced osteoclastogenesis and promotes apoptosis by activating JAK1/STAT1 and shifting activation from
TNF receptor-associated death domain (TRADD)-
TRAF2/RIP1-NF-κB to TRADD-FADD-
caspase 3 signaling.