Bub1 negatively regulates osteoclast activation through NF-κB suppression in inflammatory arthritis
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ABSTRACT:
PROVIDER: PRJNA1008093 | ENA |
REPOSITORIES: ENA
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Project description:Rheumatoid arthritis (RA) is an inflammatory autoimmune disease characterized by synovitis and, bone and cartilage destruction. Although disease-modifying anti-rheumatic drugs (DMARDs) have dramatically improved clinical outcomes, these therapies are not universally effective in all patients due to the heterogeneity of RA pathogenesis. Therefore, there is still a need to elucidate molecular mechanisms underlying RA pathogenesis to identify novel therapeutic targets. In this study, we identified Budding uninhibited by benzimidazoles 1 (BUB1) was highly expressed in RA patients’ synovium and murine ankle tissue with arthritis. Since we revealed CD45+CD11b+ myeloid cells as Bub1 highly expressing population among synovial cells, myeloid cell-specific Bub1 conditional knockout (cKO) mice were generated. Obtained cKO mice showed no significant phenotypes in healthy conditions while they exhibited reduced bone mineral density under K/BxN serum-transfer arthritis. Moreover, histomorphometric analysis suggested that the reduced bone mass was caused by increased osteoclast differentiation and activation. RNA-seq and subsequent Gene set enrichment analysis (GSEA) demonstrated significantly enriched NF-κB pathway among up-regulated genes in RANKL-stimulated bone marrow macrophages obtained from Bub1 cKO, suggesting Bub1 KO macrophages are sensitive to inflammatory conditions. Indeed, osteoclastogenesis in Bub1 KO macrophages was enhanced by RANKL and TNFα treatment and localization of NF-κB component p65 was observed. Finally, osteoclastogenesis was increased by Bub1 inhibitor BAY1816032 treatment. These data suggested that BUB1 in myeloid cells plays a protective role against bone loss under inflammatory arthritis.
2024-04-12 | GSE241409 | GEO
Project description:In this study, analysis of miRNA expression changes in osteoclast differentiation from human primary monocytes revealed the rapid upregulation of two miRNA clusters, miR-212/132 and miR-99b/let-7e/125a. We demonstrate that they negatively target monocyte-specific and immunomodulatory genes like TNFAIP3, IGF1R and IL15. Depletion of these miRNAs inhibits osteoclast differentiation and upregulates their targets. These miRNAs are also upregulated in other inflammatory monocytic differentiation processes. Most importantly, we demonstrate for the first time the direct involvement of Nuclear Factor kappa B (NF-κB) in the regulation of these miRNAs, as well as with their targets, whereby NF-κB p65 binds the promoters of these two miRNA clusters and NF-κB inhibition or depletion results in impaired upregulation of their expression. Human primary CD14+ cells isolated from three different donors, and differentiated into osteoclast with MCSF and RANKL for 21 days. miRNA expression analyzed in paired samples at day 0, 2 and 21
2014-12-03 | E-GEOD-63773 | biostudies-arrayexpress
Project description:In this study, analysis of miRNA expression changes in osteoclast differentiation from human primary monocytes revealed the rapid upregulation of two miRNA clusters, miR-212/132 and miR-99b/let-7e/125a. We demonstrate that they negatively target monocyte-specific and immunomodulatory genes like TNFAIP3, IGF1R and IL15. Depletion of these miRNAs inhibits osteoclast differentiation and upregulates their targets. These miRNAs are also upregulated in other inflammatory monocytic differentiation processes. Most importantly, we demonstrate for the first time the direct involvement of Nuclear Factor kappa B (NF-κB) in the regulation of these miRNAs, as well as with their targets, whereby NF-κB p65 binds the promoters of these two miRNA clusters and NF-κB inhibition or depletion results in impaired upregulation of their expression.
2014-12-03 | GSE63773 | GEO
Project description:Osteoclast differentiation is crucial for bone absorption and osteoclast is involved in bone destruction in rheumatoid arthritis. The aim of this study was to investigate the inhibitory effect of MJ2 on osteoclast differentiation and to elucidate its mechanism. Murine macrophage cell line, Raw 264.7 cells and collagen-induced arthritis mouse model were used for in vitro and in vivo study, respectively. MJ2-treated cells significantly inhibited osteoclast differentiation and decreased arthritic score. Surface proteins (SP) extracted from MJ2 also showed inhibitory effect on osteoclast differentiation by upregulating lipocalin 2 (lcn2) expression. Specifically, heat shock protein 60 (hsp60) in SP was revealed as an active component of MJ2. Hsp60 inhibited binding of receptor activator of nuclear factor-κB ligand (RANKL) to receptor activator of nuclear factor κB (RANK). In conclusion, MJ2 inhibited osteoclast formation and differentiation through lcn2 and RANKL-binding property and the effective component of MJ2 might be hsp60 present in surface layer.
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Project description:N4BP1 negatively regulates NF-κB by binding and inhibiting NEMO oligomerization
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Project description:Transcriptional profiling of human control and Néstor-Guillermo Progeria Syndrome (NGPS) fibroblasts and induced pluripotent stem cells (iPSCs). Somatic cell reprogramming involves rejuvenation of adult cells and relies on the ability to erase age-associated molecular marks. Accordingly, reprogramming efficiency declines with ageing, and age-associated features such as genetic instability, cell senescence or telomere shortening negatively affect this process. However, the regulatory mechanisms that constitute age-associated barriers for cell reprogramming remain largely unknown. Here, by using cells from patients with premature ageing, we demonstrate that NF-κB activation is a critical barrier for the generation of induced pluripotent stem cells (iPSCs) in ageing. We show that NF-κB repression occurs during cell reprogramming towards a pluripotent state. Conversely, ageing-associated NF-κB hyperactivation impairs generation of iPSCs by eliciting reprogramming repressors DOT1L and YY1, reinforcing cell senescence signals and down-regulating pluripotency genes. We also show that genetic and pharmacological NF-κB inhibitory strategies significantly increase the reprogramming efficiency of fibroblasts from Néstor-Guillermo Progeria Syndrome (NGPS) and Hutchinson-Gilford Progeria Syndrome (HGPS) patients, as well as from normal aged donors. Finally, we demonstrate that DOT1L inhibition in vivo ameliorates the accelerated ageing phenotype and extends lifespan in a progeroid animal model. Collectively, our results provide evidence for a novel role of NF-κB in the control of cell fate transitions and reinforce the interest of studying age-associated molecular impairments to implement cell reprogramming methodologies, and to identify new targets of rejuvenation strategies. Control and NGPS fibroblasts were reprogrammed. RNA was extracted and transcriptional profiling was obtained with GeneChip Human Exon 1.0 ST Arrays.
2015-07-01 | E-GEOD-65170 | biostudies-arrayexpress
Project description:Transcriptional profiling of human control and Néstor-Guillermo Progeria Syndrome (NGPS) mesenchymal stem cells (MSCs). Somatic cell reprogramming involves rejuvenation of adult cells and relies on the ability to erase age-associated molecular marks. Accordingly, reprogramming efficiency declines with ageing, and age-associated features such as genetic instability, cell senescence or telomere shortening negatively affect this process. However, the regulatory mechanisms that constitute age-associated barriers for cell reprogramming remain largely unknown. Here, by using cells from patients with premature ageing, we demonstrate that NF-κB activation is a critical barrier for the generation of induced pluripotent stem cells (iPSCs) in ageing. We show that NF-κB repression occurs during cell reprogramming towards a pluripotent state. Conversely, ageing-associated NF-κB hyperactivation impairs generation of iPSCs by eliciting reprogramming repressors DOT1L and YY1, reinforcing cell senescence signals and down-regulating pluripotency genes. We also show that genetic and pharmacological NF-κB inhibitory strategies significantly increase the reprogramming efficiency of fibroblasts from Néstor-Guillermo Progeria Syndrome (NGPS) and Hutchinson-Gilford Progeria Syndrome (HGPS) patients, as well as from normal aged donors. Finally, we demonstrate that DOT1L inhibition in vivo ameliorates the accelerated ageing phenotype and extends lifespan in a progeroid animal model. Collectively, our results provide evidence for a novel role of NF-κB in the control of cell fate transitions and reinforce the interest of studying age-associated molecular impairments to implement cell reprogramming methodologies, and to identify new targets of rejuvenation strategies. Control and NGPS MSCs were differentiated into bone in the presence or absence of sodium salicylate. Total RNA was extracted and global gene expression was analyzed.
2015-07-01 | E-GEOD-65172 | biostudies-arrayexpress
Project description:Many immune responses depend upon activation of NF-κB, a key transcription factor in the elicitation of a cytokine response. Here we show that N4BP1 inhibits TLR-dependent activation of NF-κB by interacting with the NF-κB signaling essential modulator (NEMO, also known as IκB kinase γ) to attenuate NEMO-NEMO dimerization or oligomerization. The UBA-like (ubiquitin associated-like) and CUE-like (ubiquitin conjugation to ER degradation) domains in N4BP1 mediate the interaction with the NEMO COZI domain. Both in vitro and in mice, N4bp1 deficiency specifically enhances TRIF-independent (TLR2, TLR7, or TLR9-mediated), but not TRIF-dependent (TLR3 or TLR4-mediated), NF-κB activation leading to increased production of proinflammatory cytokines. In response to TLR4 or TLR3 activation, TRIF causes activation of caspase-8, which cleaves N4BP1 distal to residues D424 and D490 and abolishes its inhibitory effect. N4bp1-/- mice also exhibit diminished numbers of T cells in the peripheral blood. Our work identifies N4BP1 as an inhibitory checkpoint protein that must be overcome to activate NF-κB, and a TRIF-initiated caspase-8-dependent mechanism by which this is accomplished.
2021-02-08 | GSE165916 | GEO
Project description:Paget's disease of bone (PDB) is a common disease characterized by osteoclast activation that leads to various skeletal complications. Susceptibility to PDB is mediated by a common variant at the optineurin (OPTN) locus, which is associated with reduced levels of mRNA. However, it is unclear how this leads to the development of PDB. Here, we show that OPTN acts as a negative regulator of osteoclast differentiation in vitro and that mice with a loss-of-function mutation in Optn have increased osteoclast activity and bone turnover. Osteoclasts derived from Optn mutant mice have an increase in NF-κB activation and a reduction in interferon beta expression in response to RANKL when compared to wild-type mice. These studies identify OPTN as a regulator of bone resorption and are consistent with a model whereby genetically determined reductions in OPTN expression predispose to PDB by enhancing osteoclast differentiation.
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Project description:Osteoporosis is a serious public health problem that results in fragility fractures, especially in postmenopausal women. Because the current therapeutic strategy for osteoporosis has various side effects, a safer and more effective treatment is worth exploring. It is important to examine natural plant extracts during new drug design due to low toxicity. Mogrol is an aglycon of mogroside, which is the active component of Siraitia grosvenorii and exhibits anti-inflammatory, anticancer and neuroprotective effects. Here, we demonstrated that mogrol dose-dependently inhibited osteoclast formation and function. To confirm the mechanism, RNA sequencing (RNA-seq), real-time PCR (RT–PCR), immunofluorescence and Western blotting were performed. The RNA-seq data revealed that mogrol had an effect on genes involved in osteoclastogenesis. Furthermore, RT–PCR indicated that mogrol suppressed osteoclastogenesis-related gene expression, including CTSK, ACP5, MMP9 and DC-STAMP, in RANKL-induced bone marrow macrophages (BMMs). Western blotting demonstrated that mogrol suppressed osteoclast formation by blocking TNF receptor-associated factor 6 (TRAF6)-dependent activation of the mitogen-activated protein kinase (MAPK)/nuclear factor-B (NF-κB) signaling pathway, which decreased two vital downstream transcription factors, the nuclear factor of activated T cells calcineurin-dependent 1 (NFATc1) and c-Fos proteins expression. Furthermore, mogrol dramatically reduced bone mass loss in postmenopausal mice. In conclusion, these data showed that mogrol may be a promising procedure for osteoporosis prevention or therapy.
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