Vindoline Attenuates Osteoarthritis Progression Through Suppressing the NF-κB and ERK Pathways in Both Chondrocytes and Subchondral Osteoclasts.
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ABSTRACT:
SUBMITTER: Zhu M
PROVIDER: S-EPMC8790248 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:This study explored the mechanism by which metformin (Met) inhibits osteoclast activation and determined its effects on osteoarthritis (OA) mice. Bone marrow-derived macrophages were isolated. Osteoclastogenesis was detected using tartrate-resistant acid phosphatase (TRAP) staining. Cell proliferation was evaluated using CCK-8, F-actin rings were detected by immunofluorescence staining, and bone resorption was detected using bone slices. Nuclear factor kappa-B (NF-κB) and nuclear factor of activated T-cell cytoplasmic 1 (NFATc1) were detected using luciferase assays, and the adenosine monophosphate-activated protein kinase (AMPK), NF-κB, and mitogen-activated protein kinase (MAPK) signaling pathways were detected using western blotting. Finally, expression of genes involved in osteoclastogenesis was measured using quantitative polymerase chain reaction. A knee OA mouse model was established by destabilization of the medial meniscus (DMM). Male C57BL/6J mice were assigned to sham-operated, DMM+vehicle, and DMM+Met groups. Met (100 mg/kg/d) or vehicle was administered from the first day postoperative until sacrifice. At 4- and 8-week post OA induction, micro-computed tomography was performed to analyze microstructural changes in the subchondral bone, hematoxylin and eosin staining and Safranin-O/Fast Green staining were performed to evaluate the degenerated cartilage, TRAP-stained osteoclasts were enumerated, and receptor activator of nuclear factor κB ligand (RANKL), AMPK, and NF-κB were detected using immunohistochemistry. BMM proliferation was not affected by Met treatment below 2 mM. Met inhibited osteoclast formation and bone resorption in a dose-dependent manner in vitro. Met suppressed RANKL-induced activation of p-AMPK, NF-κB, phosphorylated extracellular regulated protein kinases (p-ERK) and up-regulation of genes involved in osteoclastogenesis. Met reversed decreases in BV/TV, Tb.Th, Tb.N, and CD, and an increase in Tb.Sp at 4 weeks postoperatively. The number of osteoclasts and OARSI score were decreased by Met without effect on body weight or blood glucose levels. Met inhibited RANKL, p-AMPK, and NF-κB expression in early OA. The mechanism by which Met inhibits osteoclast activation may be associated with AMPK/NF-κB/ERK signaling pathway, indicating a novel strategy for OA treatment.
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Project description:Osteolytic diseases such as osteoporosis and neoplastic bone metastases are caused by the excessive activation of osteoclasts. Inhibiting the excessive activation of osteoclasts is a crucial strategy for treating osteolytic diseases. This study investigated the roles and mechanisms of regorafenib, a tyrosine kinase inhibitor, on osteoclasts and osteolytic diseases. We first identified the potential targets and mechanisms of regorafenib on osteoclast-related osteolytic diseases using network pharmacological analysis and molecular docking techniques. Then, we verified its role and mechanism on osteoclasts via cellular and animal experiments. Network pharmacology analysis identified 89 common targets shared by regorafenib and osteoclast-related osteolytic diseases. Enrichment analysis suggested that regorafenib may act on osteoclast-related osteolytic diseases by modulating targets such as AKT1, CASP3, MMP9, and MAPK3, regulating biological processes such as cell proliferation, apoptosis, and phosphorylation regulation, and influencing signaling pathways such as MAPK, PI3K/AKT, and osteoclast differentiation. The molecular docking results indicated that regorafenib and AKT1, CASP3, MMP9, MAPK3, and MAPK14 were stably docked. Cell experiments demonstrated that regorafenib significantly inhibited osteoclast differentiation and bone resorption in RAW 264.7 cells and bone marrow macrophages in a dose-dependent manner, with up to 50% reduction at 800 nM concentration without exhibiting cytotoxic effects. Furthermore, Western blot and RT-qPCR results demonstrated that regorafenib inhibited osteoclast differentiation by blocking the transduction of RANKL-induced NF-κB, p38, ERK, and NFAT signaling pathways. In vivo studies using an ovariectomized mouse model showed that regorafenib significantly improved bone volume fraction (BV/TV), bone surface to total volume (BS/TV), and number of trabeculae (TB.N), as well as reduced trabecular separation (Tb.Sp) compared to the OVX groups (P < 0.05). TRAcP staining results revealed a reduction in the number of osteoclasts with regorafenib treatment (P < 0.01). These results indicate that regorafenib exerts its protective effects against osteoclast-related osteolytic disease by inhibiting the RANKL-induced NF-κB, NFAT, ERK, and p38 signaling pathways. This study proves that regorafenib may serve as a potential therapeutic agent for osteoclast-related osteolytic diseases.
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Project description:This study explored the mechanism of curcumin (CUR) suppressing osteoclastogenesis and evaluated its effects on osteoarthritis (OA) mouse. Bone marrow-derived macrophages were isolated as osteoclast precursors. In the presence or absence of CUR, cell proliferation was detected by CCK-8, osteoclastogenesis was detected by tartrate-resistant acid phosphatase (TRAP) staining, F-actin rings formation was detected by immunofluorescence, bone resorption was detected by bone slices, IκBα, nuclear factor kappa-B (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways were detected using western blot, osteoclastogenesis-related gens were measured using quantitative polymerase chain reaction. A knee OA mouse model was designed by destabilizing the medial meniscus (DMM). Thirty-six male mice were divided into sham+vehicle, OA+vehicle, and OA+CUR groups. Mice were administered with or without CUR at 25 mg/kg/d from the first post-operative day until sacrifice. After 4 and 8 weeks of OA induction, micro-computed tomography was performed to analyze microstructure changes in subchondral bone, hematoxylin and eosin staining was performed to calculate the thickness of the calcified and hyaline cartilage layers, toluidine blue O staining was performed to assess the degenerated cartilage, TRAP-stained osteoclasts were counted, and NF-κB, phosphorylated Jun N-terminal Kinases (p-JNK), and receptor activator of nuclear factor κB ligand (RANKL) were detected using immunohistochemistry. CUR suppressed osteoclastogenesis and bone resorption without cytotoxicity. CUR restrained RANKL-induced activation of NF-κB, p-JNK and up-regulation of osteoclastogenesis-related genes. CUR delayed cartilage degeneration by suppressing osteoclastogenesis and bone resorption in early OA. The mechanism of CUR inhibiting osteoclastogenesis might be associated with NF-κB/JNK signaling pathway, indicating a novel strategy for OA treatment.
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Project description:Endplate osteochondritis is considered one of the major causes of intervertebral disc degeneration (IVDD) and low back pain. Menopausal women have a higher rate of endplate cartilage degeneration than similarly aged men, but the related mechanisms are still unclear. Subchondral bone changes, mainly mediated by osteoblasts and osteoclasts, are considered an important reason for the degeneration of cartilage. This work explored the role of osteoclasts in endplate cartilage degeneration, as well as its underlying mechanisms. A rat ovariectomy (OVX) model was used to induce estrogen deficiency. Our experiments indicated that OVX significantly promoted osteoclastogenesis and anabolism and catabolism changes in endplate chondrocytes. OVX osteoclasts cause an imbalance between anabolism and catabolism in endplate chondrocytes, as shown by a decrease in anabolic markers such as Aggrecan and Collagen II, and an increase in catabolic markers such as a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5) and matrix metalloproteinases (MMP13). Osteoclasts were also confirmed in this study to be able to secrete HtrA serine peptidase 1 (HTRA1), which resulted in increased catabolism in endplate chondrocytes through the NF-κB pathway under estrogen deficiency. This study demonstrated the involvement and mechanism of osteoclasts in the anabolism and catabolism changes of endplate cartilage under estrogen deficiency, and proposed a new strategy for the treatment of endplate osteochondritis and IVDD by targeting HTRA1.
| S-EPMC10320255 | biostudies-literature
Project description:Osteoarthritis (OA) is a degenerative joint disease associated with inflammatory response. Tripartite motif 8 (TRIM8) is a member of TRIM family that has been found to regulate inflammation. The present study was aimed to evaluate the role of TRIM8 in OA chondrocytes. Our results showed that TRIM8 expression was significantly increased in interleukin 1 beta (IL-1β)-stimulated OA chondrocytes. To knock down the TRIM8 expression in chondrocytes, the chondrocytes were transfected with si-TRIM8. Knockdown of TRIM8 attenuated IL-1β-induced production of inflammatory mediators including nitric oxide and prostaglandin E2. The increased expression levels of inducible nitric oxide synthase and cyclooxygenase-2 in IL-1β-induced chondrocytes were suppressed by TRIM8 knockdown. The IL-1β-induced production of proinflammatory cytokines including TNF-α and IL-6 was significantly decreased after transfection with si-TRIM8. Besides, knockdown of TRIM8 mitigated the IL-1β-induced decrease in aggrecan and collagen-II proteins expression and increase in matrix-degrading enzymes in chondrocytes. Furthermore, TRIM8 knockdown prevented IL-1β-induced nuclear factor kappa B (NF-κB) activation in chondrocytes. Taken together, these findings indicated that knockdown of TRIM8 attenuates IL-1β-induced inflammatory response in OA chondrocytes through the inactivation of NF-κB pathway. Thus, targeting TRIM8 might provide therapeutic treatment for OA.
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Project description:BackgroundOsteoarthritis (OA) is a common joint disorder worldwide which causes great health and economic burden. However, there remains an unmet goal to develop an effective therapeutic method to prevent or delay OA. Chondrocytes, as the major cells involved in OA progression, may serve as a promising therapeutic target.ResultsA kind of carbon dots (CDs) with excellent biocompatibility was fabricated from folic acid via hydrothermal method and could effectively attenuate osteoarthritis. It was demonstrated that CDs treatment could rescue IL1β-induced proinflammatory responses, oxidative stress, cartilage degeneration and extracellular matrix degradation. Moreover, CDs reprogrammed lipopolysaccharide (LPS)-induced macrophage inflammation and polarization. Conditioned medium (CM) from CDs-treated macrophages could attenuate IL1β-induced chondrocyte injury. Also, CM from CDs-treated chondrocytes had immunoregulatory functions on macrophages. Mechanistically, CDs inhibited the activation of nuclear factor-κB (NF-κB) and mitogen-activated protein kinases (MAPK) signaling pathways in IL1β-stimulated chondrocytes. In vivo, anterior cruciate ligament transection (ACLT) mice model was adopted and it was indicated that intra-articular injection of CDs effectively delays OA pathogenesis.ConclusionsTaken together, these findings indicated CDs could mediate OA via promoting cartilage repair and immunomodulating macrophages within local microenvironment, which may provide evidences for utilizing CDs as a novel nanomaterial for OA treatment.
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Project description:Joint pain is the defining symptom of osteoarthritis (OA) but its origin and mechanisms remain unclear. Here, we investigated an unprecedented role of osteoclast-initiated subchondral bone remodeling in sensory innervation for OA pain. We show that osteoclasts secrete netrin-1 to induce sensory nerve axonal growth in subchondral bone. Reduction of osteoclast formation by knockout of receptor activator of nuclear factor kappa-B ligand (Rankl) in osteocytes inhibited the growth of sensory nerves into subchondral bone, dorsal root ganglion neuron hyperexcitability, and behavioral measures of pain hypersensitivity in OA mice. Moreover, we demonstrated a possible role for netrin-1 secreted by osteoclasts during aberrant subchondral bone remodeling in inducing sensory innervation and OA pain through its receptor DCC (deleted in colorectal cancer). Importantly, knockout of Netrin1 in tartrate-resistant acid phosphatase-positive (TRAP-positive) osteoclasts or knockdown of Dcc reduces OA pain behavior. In particular, inhibition of osteoclast activity by alendronate modifies aberrant subchondral bone remodeling and reduces innervation and pain behavior at the early stage of OA. These results suggest that intervention of the axonal guidance molecules (e.g., netrin-1) derived from aberrant subchondral bone remodeling may have therapeutic potential for OA pain.
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Project description:Prostaglandin E2 (PGE2), a major cyclooxygenase-2 (COX-2) product, is highly secreted by the osteoblast lineage in the subchondral bone tissue of osteoarthritis (OA) patients. However, NSAIDs, including COX-2 inhibitors, have severe side effects during OA treatment. Therefore, the identification of novel drug targets of PGE2 signaling in OA progression is urgently needed. Osteoclasts play a critical role in subchondral bone homeostasis and OA-related pain. However, the mechanisms by which PGE2 regulates osteoclast function and subsequently subchondral bone homeostasis are largely unknown. Here, we show that PGE2 acts via EP4 receptors on osteoclasts during the progression of OA and OA-related pain. Our data show that while PGE2 mediates migration and osteoclastogenesis via its EP2 and EP4 receptors, tissue-specific knockout of only the EP4 receptor in osteoclasts (EP4LysM) reduced disease progression and osteophyte formation in a murine model of OA. Furthermore, OA-related pain was alleviated in the EP4LysM mice, with reduced Netrin-1 secretion and CGRP-positive sensory innervation of the subchondral bone. The expression of platelet-derived growth factor-BB (PDGF-BB) was also lower in the EP4LysM mice, which resulted in reduced type H blood vessel formation in subchondral bone. Importantly, we identified a novel potent EP4 antagonist, HL-43, which showed in vitro and in vivo effects consistent with those observed in the EP4LysM mice. Finally, we showed that the Gαs/PI3K/AKT/MAPK signaling pathway is downstream of EP4 activation via PGE2 in osteoclasts. Together, our data demonstrate that PGE2/EP4 signaling in osteoclasts mediates angiogenesis and sensory neuron innervation in subchondral bone, promoting OA progression and pain, and that inhibition of EP4 with HL-43 has therapeutic potential in OA.
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Project description:Osteoarthritis (OA) is a chronic progressive osteoarthropathy in the elderly. Osteoclast activation plays a crucial role in the occurrence of subchondral bone loss in early OA. However, the specific mechanism of osteoclast differentiation in OA remains unclear. In our study, gene expression profiles related to OA disease progression and osteoclast activation were screened from the Gene Expression Omnibus (GEO) repository. GEO2R and Funrich analysis tools were employed to find differentially expressed genes (DEGs). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses demonstrated that chemical carcinogenesis, reactive oxygen species (ROS), and response to oxidative stress were mainly involved in osteoclast differentiation in OA subchondral bone. Furthermore, fourteen DEGs that are associated with oxidative stress were identified. The first ranked differential gene, heme oxygenase 1 (HMOX1), was selected for further validation. Related results showed that osteoclast activation in the pathogenesis of OA subchondral bone is accompanied by the downregulation of HMOX1. Carnosol was revealed to inhibit osteoclastogenesis by targeting HMOX1 and upregulating the expression of antioxidant protein in vitro. Meanwhile, carnosol was found to alleviate the severity of OA by inhibiting the activation of subchondral osteoclasts in vivo. Our research indicated that the activation of osteoclasts due to subchondral bone redox dysplasia may serve as a significant pathway for the advancement of OA. Targeting HMOX1 in subchondral osteoclasts may offer novel insights for the treatment of early OA.
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Project description:Cigarette smoke (CS) is the main cause of chronic obstructive pulmonary disease (COPD), and continuous CS exposure causes lung inflammation and deterioration. To investigate the protective effects of Artemisia gmelinii against lung inflammation in this study, cigarette smoke extract (CSE)/lipopolysaccharide (LPS)-treated alveolar macrophages (AMs) and mice stimulated with CSE/porcine pancreas elastase (PPE) were used. Artemisia gmelinii ethanol extract (AGE) was effective in decreasing the levels of cytokines, chemokine, inducible nitric oxide synthase, and cyclooxygenase-2 by inhibiting mitogen-activated protein (MAP) kinases/nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway in AMs. Additionally, oral administration of AGE suppressed inflammatory cells' infiltration and secretion of inflammatory cytokines, chemokines, matrix metallopeptidase 9, and neutrophil extracellular traps in bronchoalveolar lavage fluid from the COPD model. Moreover, the obstruction of small airways, the destruction of the lung parenchyma, and expression of IL-6, TNF-α, IL-1β, and MIP-2 were suppressed by inhibiting NF-κB activation in the lung tissues of the AGE group. These effects are associated with scopolin, chlorogenic acid, hyperoside, 3,4-di-O-caffeoylquinic acid, 3,5-di-O-caffeoylquinic acid, and 4,5-di-O-caffeoylquinic acid, which are the main components of AGE. These data demonstrate the mitigation effect of AGE on lung inflammation via inhibition of MAPK and NF-κB pathways, suggesting that AGE may be instrumental in improving respiratory and lung health.
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