IL-3 inhibits rat osteoclast differentiation induced by TNF-α and other pro-osteoclastogenic cytokines.

IL-3, a haematopoiesis regulatory factor, has previously been shown to inhibit both mouse and human osteoclast differentiation and bone resorption. Here, the role of rat IL-3 on rat osteoclast differentiation was evaluated to address whether the inhibitory action of IL-3 on osteoclastogenesis is conserved in various species. It was observed that IL-3 inhibited rat osteoclast differentiation induced by both TNF-α and receptor activator of NF-ĸB ligand (RANKL). TNF-α is known to induce bone loss in postmenopausal osteoporotic women and it also synergise with many pro-osteoclastogenic cytokines to cause huge pathological bone loss. Importantly, it was found that rat IL-3 inhibits the synergistic action of TNF-α with RANKL and IL-1ß, TGFß1 and TGF-ß3. IL-3 downregulates the TNF-α-induced nuclear translocation of NF-ĸB-p65 and c-fos without affecting c-jun. Interestingly, we observed that IL-3 also inhibits osteoclast differentiation in vivo in rats induced by TNF-α. All these results suggest that inhibitory action of IL-3 on osteoclastogenesis is conserved in various species including mice, rats and humans. Thus, our results clearly indicate that IL-3 has therapeutic potential to treat pathological bone loss in important skeletal diseases.

IL-3 Differentially Regulates Membrane and Soluble RANKL in Osteoblasts through Metalloproteases and the JAK2/STAT5 Pathway and Improves the RANKL/OPG Ratio in Adult Mice.

Bone remodeling comprises balanced activities between osteoclasts and osteoblasts, which is regulated by various factors, including hormones and cytokines. We previously reported that IL-3 inhibits osteoclast differentiation and pathological bone loss. IL-3 also enhances osteoblast differentiation and bone formation from mesenchymal stem cells. However, the role of IL-3 in regulation of osteoblast-osteoclast interactions and underlying mechanisms is not yet delineated. In this study, we investigated the role of IL-3 on the regulation of osteoblast-specific molecules, receptor activator of NF-κB ligand (RANKL), and osteoprotegerin (OPG) that modulate bone homeostasis. We found that IL-3 increases RANKL expression at both the transcriptional and translational levels, and it showed no effect on OPG expression in calvarial osteoblasts. The increased RANKL expression by IL-3 induces mononuclear osteoclasts; however, it does not induce multinuclear osteoclasts. Interestingly, IL-3 decreases soluble RANKL by reducing ectodomain shedding of membrane RANKL through downregulation of metalloproteases mainly a disintegrin and metalloproteinase (ADAM)10, ADAM17, ADAM19, and MMP3. Moreover, IL-3 increases membrane RANKL by activating the JAK2/STAT5 pathway. Furthermore, IL-3 enhances RANKL expression in mesenchymal stem cells of wild-type mice but not in STAT5a knockout mice. Interestingly, IL-3 restores RANKL expression in adult mice by enhancing bone-specific RANKL and decreasing serum RANKL. Furthermore, IL-3 increases the serum OPG level in adult mice. Thus, our results reveal, to our knowledge for the first time, that IL-3 differentially regulates two functional forms of RANKL through metalloproteases and the JAK2/STAT5 pathway, and it helps in restoring the decreased RANKL/OPG ratio in adult mice. Notably, our studies indicate the novel role of IL-3 in regulating bone homeostasis in important skeletal disorders.

Adipose-Derived Mesenchymal Stem Cells Prevent Systemic Bone Loss in Collagen-Induced Arthritis.

Rheumatoid arthritis (RA) is an autoimmune disease characterized by chronic inflammatory synovitis leading to joint destruction and systemic bone loss. The inflammation-induced bone loss is mediated by increased osteoclast formation and function. Current antirheumatic therapies primarily target suppression of inflammatory cascade with limited or no success in controlling progression of bone destruction. Mesenchymal stem cells (MSCs) by virtue of their tissue repair and immunomodulatory properties have shown promising results in various autoimmune and degenerative diseases. However, the role of MSCs in prevention of bone destruction in RA is not yet understood. In this study, we investigated the effect of adipose-derived MSCs (ASCs) on in vitro formation of bone-resorbing osteoclasts and pathological bone loss in the mouse collagen-induced arthritis (CIA) model of RA. We observed that ASCs significantly inhibited receptor activator of NF-κB ligand (RANKL)-induced osteoclastogenesis in both a contact-dependent and -independent manner. Additionally, ASCs inhibited RANKL-induced osteoclastogenesis in the presence of proinflammatory cytokines such as TNF-α, IL-17, and IL-1ß. Furthermore, treatment with ASCs at the onset of CIA significantly reduced clinical symptoms and joint pathology. Interestingly, ASCs protected periarticular and systemic bone loss in CIA mice by maintaining trabecular bone structure. We further observed that treatment with ASCs reduced osteoclast precursors in bone marrow, resulting in decreased osteoclastogenesis. Moreover, ASCs suppressed autoimmune T cell responses and increased the percentages of peripheral regulatory T and B cells. Thus, we provide strong evidence that ASCs ameliorate inflammation-induced systemic bone loss in CIA mice by reducing osteoclast precursors and promoting immune tolerance.