C/ebpα controls osteoclast terminal differentiation, activation, function, and postnatal bone homeostasis through direct regulation of Nfatc1

Abstract

Osteoclast lineage commitment and differentiation have been studied extensively, although the mechanism by which transcription factor(s) control osteoclast terminal differentiation, activation and function remains unclear. CCAAT/enhancer-binding protein α (C/ebpα) has been reported to be a key regulator of osteoclast cell lineage commitment, yet C/ebpα’s roles in osteoclast terminal differentiation, activation and function and bone homeostasis, under physiological or pathological conditions, have not been studied because newborn C/ebpα null mice die within several hours after birth. Furthermore, the function of C/ebpα in osteoclast terminal differentiation, activation and function are largely unknown. Herein, we generated and analyzed an osteoclast-specific C/ebpα conditional knockout (CKO) mouse model via Ctsk-Cre mice and found that C/ebpα-deficient mice exhibited a severe osteopetrosis phenotype due to impaired osteoclast terminal differentiation, activation and function, including mildly reduced osteoclast number, impaired osteoclast polarization, actin formation, and bone resorption, which demonstrated the novel function of C/ebpα in cell function and terminal differentiation. Interestingly, C/ebpα deficiency did not affect bone formation or monocyte/macrophage development. Our results further demonstrated that C/ebpα deficiency suppressed the expression of osteoclast functional genes, e.g. encoding Cathepsin K (Ctsk), Atp6i (Tcirg1) and osteoclast regulator genes, e.g. encoding c-fos (Fos), and nuclear factor of activated T-cells 1 (Nfatc1), while having no effect on Pu.1 (Spi1) expression. Promoter activity mapping and ChIP assay defined the critical cis-regulatory element (CCRE) in the promoter region of Nfatc1, and also showed that the CCREs were directly associated with C/ebpα, which enhanced the promoter’s activity. The deficiency of C/ebpα in osteoclasts completely blocked ovariectomy-induced bone loss, indicating C/ebpα is a promising new target for the treatment of osteolytic diseases.

Interleukin‐20 differentially regulates bone mesenchymal stem cell activities in RANKL‐induced osteoclastogenesis through the OPG/RANKL/RANK axis and the NF‐κB, MAPK and AKT signalling pathways

“…While exploring the molecular mechanism(s) underlying the differential regulation of osteoclastogenesis by IL‐20, we found that IL‐20 dose‐dependently upregulated and downregulated the expression of osteoclast‐specific genes and transcriptional activators to perform bone resorption function, such as RANK, CK, TRAP, MT1‐MMP, MMP9, NFATc1 and c‐Fos. In signal transduction, activation of the transcription factors NFATc1 and c‐Fos is mainly regulated by several key signalling pathways including NF‐κB, p38, ERK, JNK and AKT pathways, which are involved in proliferation, apoptosis, differentiation and function in osteoclasts . In this case, NF‐κB, p38 and JNK pathways act as positive regulation in osteoclastogenesis, whereas ERK and AKT pathways are involved in negative regulation .…”

At the Crossroads of the Adipocyte and Osteoclast Differentiation Programs: Future Therapeutic Perspectives

“…The intracellular osteoclastogenic pathways linked to these stimuli are mediated through distinct RANK-mediated activation of tumor necrosis factor receptor-associated factors (TRAFs) that leads to the feed forward expression and activation of nuclear factor-kappa B (NFκB), c-fos and nuclear factor of activated T cells 1 (NFATc1) in HSCs, ultimately inducing the osteoblast-dependent osteoclast differentiation [40][41][42]. However, several recent studies have addressed the intracellular mechanisms that are interwoven between the adipogenic and osteoclastogenic programs [27][28][29][30][31][32]. In this context, it has been reported that some of the key players known to regulate MSC adipocyte differentiation are expressed and crucial for HSC osteoclast differentiation and likewise, the major osteoclast differentiation factor RANKL is highly expressed in bone marrow adipocytes [27][28][29][30][31][32][33][34][43][44][45][46].…”

“…Recent studies have revealed that adipogenic transcription factors including PPARγ, C/EBPα and CEBPβ are also expressed and activated in hematopoietic lineage cells during osteoclast differentiation and are critical for the lineage priming, differentiation and activity of osteoclasts [27][28][29][30][31][32][33][34][35]. Furthermore, several adipokines are reported to influence the osteoclastogenic program of HSCs suggesting that increased bone marrow adipogenesis could exert further detrimental effects on bone health by uncoupling osteoblast-osteoclast communication in a fashion that promotes osteoclastogenesis and bone resorption [6,7,[36][37][38].…”

F2r negatively regulates osteoclastogenesis through inhibiting the Akt and NFκB signaling pathways

“…We used 10% sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE) to perform western blot as described [24,28,29]. MBMs were transfected lentivirus and induced to 5 days for western blot analysis.…”

“…Next, we observed the formation of the F-actin ring, which is an important structure for osteoclast maturation [29]. Equal numbers of MBM cells were seeded on each bone slice and induced by M-CSF and RANKL to generate osteoclasts.…”