Signaling Pathway In Chondrocytes Research Articles

TBK1 pharmacological inhibition mitigates osteoarthritis through attenuating inflammation and cellular senescence in chondrocytes

ObjectivesTANK-binding kinase 1 (TBK1) is pivotal in autoimmune and inflammatory diseases, yet its role in osteoarthritis (OA) remains elusive. This study sought to elucidate the effect of the TBK1 inhibitor BX795 on OA and to delineate the underlying mechanism by which it mitigates OA. MethodsInterleukin-1 Beta (IL-1β) was utilized to simulate inflammatory responses and extracellular matrix degradation in vitro. In vivo, OA was induced in 8-week-old mice through destabilization of the medial meniscus surgery. The impact of BX795 on OA was evaluated using histological analysis, X-ray, micro-CT, and the von Frey test. Additionally, Western blot, RT-qPCR, and immunofluorescence assays were conducted to investigate the underlying mechanisms of BX795. ResultsPhosphorylated TBK1 (P-TBK1) levels were found to be elevated in OA knee cartilage of both human and mice. Furthermore, intra-articular injection of BX795 ameliorated cartilage degeneration and alleviated OA-associated pain. BX795 also counteracted the suppression of anabolic processes and the augmentation of catabolic activity, inflammation, and senescence observed in the OA mice. In vitro studies revealed that BX795 reduced P-TBK1 levels and reversed the effects of anabolism inhibition, catabolism promotion, and senescence induction triggered by IL-1β. Mechanistically, BX795 inhibited the IL-1β-induced activation of the cGAS–STING and TLR3–TRIF signaling pathways in chondrocytes. ConclusionsPharmacological inhibition of TBK1 with BX795 protects articular cartilage by inhibiting the activation of the cGAS–STING and TLR3–TRIF signaling pathways. This action attenuates inflammatory responses and cellular senescence, positioning BX795 as a promising therapeutic candidate for OA treatment. The translational potential of this articleThis study furnishes experimental evidence and offers a potential mechanistic explanation supporting the efficacy of BX795 as a promising candidate for OA treatment.

Polygonatum sibiricum polysaccharides protect against knee osteoarthritis by inhibiting the TLR2/NF-κB signaling pathway in vivo and in vitro

Polygonatum sibiricum polysaccharides (PSP), the primary constituent of Polygonatum sibiricum, have been shown to exhibit a wide range of pharmacological effects, but their impact on osteoarthritis (OA) remains unclear. The objective of this study was to investigate the protective effects of PSP against OA and to elucidate its underlying molecular mechanism. In our in vitro experiments, PSP not only inhibited the IL-1β-induced inflammatory responses and the nuclear factor kappa-B (NF-κB) signaling pathway in chondrocytes but also regulated the cartilage matrix metabolism. In addition, we detected 394 significantly differentially expressed genes through RNA-seq analysis on PSP-intervened chondrocytes, and the toll-like receptor 2 (TLR2) was identified as the most important feature by functional network analysis and qRT-PCR. It was also revealed that PSP treatment significantly reversed the IL-1-induced up-regulation of TLR2 expression in chondrocytes, while TLR2 overexpression partially inhibited the regulatory effects of PSP on inflammation, NF-κB signaling pathway and matrix metabolism. In our in vivo experiments, PSP treatment alleviated the development of destabilization of medial meniscus (DMM)-induced OA in mouse knee joints, inhibited the DMM-induced activation of the TLR2/NF-κB signaling pathway in mouse knee joint cartilage, and reduced the serum levels of inflammatory cytokines. In conclusion, PSP exerts its anti-inflammatory, matrix synthesis-promoting and matrix catabolism-suppressing effects in knee OA by inhibiting the TLR2/NF-κB signaling pathway, suggesting that PSP may be potentially targeted as a novel all-natural, low-toxicity drug for OA prevention and treatment.

Controlled mechanical loading affects the osteocyte transcriptome in porcine trabecular bone in situ

IntroductionOsteocytes modulate bone adaptation in response to mechanical stimuli imparted by the deforming bone tissue in which they are encased by communicating with osteoclasts and osteoblasts as well as other osteocytes in the lacuna-canalicular network through secreted cytokines and chemokines. Understanding the transcriptional response of osteocytes to mechanical stimulation in situ could identify new targets to inhibit bone loss or enhance bone formation in the presence of diseases like osteoporosis or metastatic cancer. We compared the mechanically regulated transcriptional response of osteocytes in trabecular bone following one or three days of controlled mechanical loading. MethodsPorcine trabecular bone explants were cultured in a bioreactor for 48 h and subsequently loaded twice a day for one day or 3 days. RNA was isolated and sequenced, and the Tuxedo suite was used to identify differentially expressed genes and pathway analysis was conducted using Ingenuity Pathway Analysis (IPA). ResultsThere were about 4000 differentially expressed genes following in situ culture relative to fresh bone. One hundred six genes were differentially expressed between the loaded and non-loaded groups following one day of loading compared to 913 genes after 3 d of loading. Only 45 of these were coincident between the two time points, indicating an evolving transcriptome. Clustering and principal component analysis indicated differences between the loaded and non-loaded groups after 3 d of loading. DiscussionWith sustained loading, there was a nine-fold increase in the number of differentially expressed genes, suggesting that osteocytes respond to loading through sequential activation of downstream genes in the same pathways. The differentially expressed genes were related to osteoarthritis, osteocyte, and chondrocyte signaling pathways. We noted that NFkB and TNF signaling are affected by early loading and this may drive downstream effects on the mechanobiological response. Moreover, these genes may regulate catabolic effects of mechanical disuse through their actions on pre-osteoclasts in the bone marrow niche.

Bisdemethoxycurcumin, a curcumin, protects chondrocytes, and reduces cartilage inflammation via the NRF2/HO-1/NLRP3 pathway.

The objective of this thesis is to evaluate the effect of bisdemethoxycurcumin (BDMC) on osteoarthritis (OA) and comprehensively evaluate the role of the Nuclear Factor erythroid 2-Related Factor 2 (Nrf2) signalling pathway in chondrocytes. In our study, we treated chondrocytes with BDMC in an in vitro chondrocyte assay and measured its influence on extracellular matrix (ECM) expression, downstream heme oxygenase-1 (HO-1) and NOD-like receptor thermal protein domain associated protein 3 (NLRP3) levels. Our study indicates that BDMC significantly activates the Nrf2 signaling pathway in chondrocytes in vitro. Furthermore, the expression of matrix metalloproteinase 3, interleukin 1β, recombinant a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)4 and (ADAMTS)5 was significantly suppressed by BDMC. This study confirms the potential for BDMC to activate the Nrf2/HO-1/NLRP3 signalling pathway and alleviate OA symptoms. Therefore, BDMC is a promising therapeutic agent for OA that offers new insights and treatment methods.

Sirt6 promotes DNA damage repair in osteoarthritis chondrocytes by activating the Keap1/Nrf2/HO-1 signaling pathway

ABSTRACT The aim of this study was to explore the effect and mechanism of Sirt6 on DNA damage repair in OA chondrocytes. Cartilage tissues were collected from OA patients with knee arthroplasty and traumatic amputation patients without OA. Besides, 7-week-old male C57BL/6 mice were randomly divided into Control and OA groups; CHON-001 cells of corresponding groups were treated with 10 ng/ml interleukin (IL)-1β, respectively. Subsequently, Sirt6 or siNrf2 was over-expressed in CHON-001 cells to observe the effect of Sirt6 on DNA damage and senescence of chondrocytes by IL-1β through the nuclear factor E2-related factor 2 (Nrf2) signaling pathway. The expression level of Sirt6 in human and mouse OA cartilage tissues was significantly decreased. However, 24 h of treatment with IL-1β significantly decreased the expression of Sirt6 in chondrocytes, induced DNA damage, and promoted cellular senescence. In addition, over-expression of Sirt6 promoted DNA damage repair and inhibited cellular senescence in IL-1β-induced chondrocytes. Moreover, the overexpression of Sirt6 activated the Keap1/Nrf2/HO-1 signaling pathway in chondrocytes, while knockdown of Nrf2 expression inhibited the DNA damage repair and anti-senescence effects of Sirt6 on IL-1β-treated chondrocytes. Sirt6 may reduce DNA damage and cellular senescence in OA chondrocytes induced by IL-1β through activating the Keap1/Nrf2/HO-1 signaling pathway.

Suramin ameliorates osteoarthritis by acting on the Nrf2/HO-1 and NF-κB signaling pathways in chondrocytes and promoting M2 polarization in macrophages

Osteoarthritis (OA)—the most prevalent of arthritis diseases—is a complicated pathogenesis caused by cartilage degeneration and synovial inflammation. Suramin has been reported to enhance chondrogenic differentiation. However, the therapeutic effect of suramin on OA-induced cartilage destruction has remained unclear. Suramin is an anti-parasitic drug that has potent anti-purinergic properties. This study investigated the protective effects and underlying mechanisms of suramin on articular cartilage degradation using an in vitro study and mice model with post-traumatic OA. We found that suramin markedly suppressed the IL-1β increased expression of matrix destruction proteases—such as ADAMT4, ADAMTS5, MMP3, MMP13, and inflammatory mediators—including the iNOS, COX2, TNFα, and IL-1β; while greatly enhancing the synthesis of cartilage anabolic factors—such as COL2A1, Aggrecan and SOX9 in IL-1β-induced porcine chondrocytes. In vivo experiments showed that intra-articular injection of suramin ameliorated cartilage degeneration and inhibited synovial inflammation in an anterior cruciate ligament transection (ACLT)-induced OA mouse model. In mechanistic studies, we found that exogenous supplementation of suramin can activate Nrf2, and accordingly inhibit the nuclear factor kappa-light-chain-enhancer of activated B cells (NF- κB) and mitogen-activated protein kinase (MAPK) pathways, thereby alleviating the inflammation and ECM degeneration of chondrocytes stimulated by IL-1β. In addition, suramin also repolarized M1 macrophages to the M2 phenotype, further reducing the apoptosis of chondrocytes. Collectively, the results of the study suggests that suramin is a potential drugs which could serve as a facilitating drug for the application of OA therapy toward clinical treatment.

Identification of the osteoarthritis signature gene PDK1 by machine learning and its regulatory mechanisms on chondrocyte autophagy and apoptosis.

Osteoarthritis (OA) is a degenerative joint disease frequently diagnosed in the elderly and middle-aged population. However, its specific pathogenesis has not been clarified. This study aimed to identify biomarkers for OA diagnosis and elucidate their potential mechanisms for restoring OA-dysregulated autophagy and inhibiting chondrocyte apoptosis in vitro. Two publicly available transcriptomic mRNA OA-related datasets (GSE10575 and GSE51588) were explored for biomarker identification by least absolute shrinkage and selection operator (LASSO) regression, weighted gene co-expression network analysis (WGCNA), and support vector machine recursive feature elimination (SVM-RFE). We applied the GSE32317 and GSE55457 cohorts to validate the markers' efficacy for diagnosis. The connections of markers to chondrocyte autophagy and apoptosis in OA were also comprehensively explored in vitro using molecular biology approaches, including qRT-PCR and Western blot. We identified 286 differentially expressed genes (DEGs). These DEGs were enriched in the ECM-receptor interaction and PI3K/AKT signaling pathway. After external cohort validation and protein-protein interaction (PPI) network construction, PDK1 was finally identified as a diagnostic marker for OA. The pharmacological properties of BX795-downregulated PDK1 expression inhibited LPS-induced chondrocyte inflammation and apoptosis and rescued OA-dysregulated autophagy. Additionally, the phosphorylation of the mediators associated with the MAPK and PI3K/AKT pathways was significantly downregulated, indicating the regulatory function of PDK1 in apoptosis and autophagy via MAPK and PI3K/AKT-associated signaling pathways in chondrocytes. A significantly positive association between the PDK1 expression and Neutrophils, Eosinophils, Plasma cells, and activated CD4 memory T cells, as well as an evident negative correlation between T cells follicular helper and CD4 naive T cells, were detected in the immune cell infiltration analysis. PDK1 can be used as a diagnostic marker for OA. Inhibition of its expression can rescue OA-dysregulated autophagy and inhibit apoptosis by reducing the phosphorylation of PI3K/AKT and MAPK signaling pathways.

Oroxin B alleviates osteoarthritis through anti-inflammation and inhibition of PI3K/AKT/mTOR signaling pathway and enhancement of autophagy.

Osteoarthritis (OA) is a common aging-related degenerative joint disease with chronic inflammation as its possible pathogenesis. Oroxin B (OB), a flavonoid isolated from traditional Chinese herbal medicine, possesses anti-inflammation properties which may be involved in regulating the pathogenesis of OA, but its mechanism has not been elucidated. Our study was the first to explore the potential chondroprotective effect and elucidate the underlying mechanism of OB in OA. In vitro, primary mice chondrocytes were stimulated with IL-1β along with or without the administration of OB or autophagy inhibitor 3-methyladenine (3-MA). Cell viability assay was measured with a cell counting kit-8 (CCK-8). The phenotypes of anabolic-related (Aggrecan and Collagen II), catabolic-related (MMP3, MMP13, and ADAMTS5), inflammation-related (iNOS, COX-2, TNF-α, IL-6, and IL-1β), and markers of related signaling pathways in chondrocytes with different treatment were detected through western blot, RT-qPCR, and immunofluorescent staining. In vivo, the destabilized medial meniscus (DMM) operation was performed to establish the OA mice model. After knee intra-articular injection with OB for 8 weeks, the mice's knee joints were obtained for subsequent histological staining and analysis. OB reversed the expression level of anabolic-related proteins (Aggrecan and Collagen II) and catabolic-related (MMP3, MMP13, and ADAMTS5) in IL-1β-induced chondrocytes. Mechanistically, OB suppressed the inflammatory response stimulated by IL-1β, as the inflammation-related (iNOS, COX-2, TNF-α, IL-6, and IL-1β) markers were downregulated after the administration of OB in IL-1β-induced chondrocytes. Besides, the activation of PI3K/AKT/mTOR signaling pathway induced by IL-1β could be inhibited by OB. Additionally, the autophagy process impaired by IL-1β could be rescued by OB. What's more, the introduction of 3-MA to specifically inhibit the autophagic process impairs the protective effect of OB on cartilage. In vivo, histological staining revealed that intra-articular injection of OB attenuated the cartilage degradation, as well as reversed the expression level of anabolic and catabolic-related proteins such as Aggrecan, Collagen II, and MMP13 induced in DMM-induced OA models. The study verified that OB exhibited the chondroprotective effect by anti-inflammatory, inhibiting the PI3K/AKT/mTOR signaling pathway, and enhancing the autophagy process, indicating that OB might be a promising agent for the treatment of OA.

Selumetinib - a potential small molecule inhibitor for osteoarthritis treatment.

Objectives: Osteoarthritis (OA) is a common disease that mainly manifests as inflammation and destruction of cartilage and subchondral bone. Recently, necroptosis has been reported to play an important role in the development of OA. Selumetinib displays a contrasting expression pattern to necroptosis-related proteins. The present study aimed to investigate the potential therapeutic effects of selumetinib in OA process. Methods: In vitro experiments, interleukin-1β (IL-1β) was used to induce necroptosis of chondrocytes. We used high-density cell culture, Western Blot and PT-PCR to observe the effect of different concentrations of selumetinib on the extracellular matrix of cartilage. Afterwards, we visualized the effect of selumetinib on osteoclast formation by TRAP staining and F-actin rings. In vivo experiment, we induced experimental osteoarthritis in mice by surgically destabilizing the medial meniscus (DMM) while administering different concentrations of selumetinib intraperitoneally. Results: Selumetinib promoted cartilage matrix synthesis and inhibited matrix decomposition. We found that selumetinib exerted a protective function by inhibiting the activation of RIP1/RIP3/MLKL signaling pathways in chondrocytes. Selumetinib also inhibited the activation of RANKL-induced NF-κB and MAPK signaling pathways in BMMs, thereby interfering with the expression of osteoclast marker genes. In the DMM-induced OA model, a postsurgical injection of selumetinib inhibited cartilage destruction and lessened the formation of TRAP-positive osteoclasts in subchondral bone. Conclusion: Selumetinib can protect chondrocytes by regulating necroptosis to prevent the progression of OA and reduce osteoclast formation. In summary, our findings suggest that selumetinib has potential as a therapeutic agent for OA.

Primary Cilia: A Cellular Regulator of Articular Cartilage Degeneration.

Osteoarthritis (OA) is the most common joint disease that can cause pain and disability in adults. The main pathological characteristic of OA is cartilage degeneration, which is caused by chondrocyte apoptosis, cartilage matrix degradation, and inflammatory factor destruction. The current treatment for patients with OA focuses on delaying its progression, such as oral anti-inflammatory analgesics or injection of sodium gluconate into the joint cavity. Primary cilia are an important structure involved in cellular signal transduction. Thus, they are very sensitive to mechanical and physicochemical stimuli. It is reported that the primary cilia may play an important role in the development of OA. Here, we review the correlation between the morphology (location, length, incidence, and orientation) of chondrocyte primary cilia and OA and summarize the relevant signaling pathways in chondrocytes that could regulate the OA process through primary cilia, including Hedgehog, Wnt, and inflammation-related signaling pathways. These data provide new ideas for OA treatment.

Circular RNA circNFKB1 promotes osteoarthritis progression through interacting with ENO1 and sustaining NF-κB signaling

Inflammatory cytokines-induced activation of the nuclear factor κB (NF-κB) pathway plays a critical role in the pathogenesis of osteoarthritis (OA). Circular RNA (circRNA) has been identified as important epigenetic factor in numerous diseases. However, the biological roles of inflammation-related circRNAs in regulating OA pathogenesis remain elusive. Here, we revealed circRNA expression profiles in human primary chondrocytes with interleukin-1β (IL-1β) stimulation by circRNA sequencing. We identified a highly upregulated circRNA, termed as circNFKB1 in inflamed chondrocytes and osteoarthritic cartilage. As a circRNA derived from exon 2–5 of NFKB1, circNFKB1 is located in both cytoplasm and nucleus of chondrocytes. Furthermore, knockdown of circNFKB1 inhibited extracellular matrix (ECM) catabolism and rescued IL-1β impaired ECM anabolism whereas ectopic expression of circNFKB1 significantly promoted chondrocytes degradation in vitro. Moreover, intraarticular injection of adenovirus-circNFKB1 in mouse joints triggered spontaneous cartilage loss and OA development. Mechanistically, circNFKB1 interacted with α-enolase (ENO1), regulated the expression of its parental gene NFKB1 and sustained the activation of NF-κB signaling pathway in chondrocytes. Therefore, this study highlights a novel ENO1-interacting circNFKB1 in OA pathogenesis, and provides valuable insights into understanding the regulatory mechanism of NF-κB signaling in chondrocytes and a promising therapeutic target for the treatment of OA.

Polygonatum sibiricum polysaccharide inhibits IL-1β-induced inflammation in human chondrocytes

Osteoarthritis (OA) is a degenerative joint disease associated with inflammation. Polygonatum sibiricum polysaccharide (PSP) is a major group of active components isolated from Polygonatum sibiricum with broad activities including anti-inflammatory effect. However, the role of PSP in OA is unclear. In the present study, we aimed to investigate the effects of PSP on IL-1β-induced inflammatory response in primary human OA chondrocytes. The results showed that PSP improved cell viability of chondrocytes in response to IL-1β induction. The increased levels of several inflammatory mediators including nitric oxide (NO), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in IL-1β-induced chondrocytes were attenuated by PSP in a dose-dependent manner. The IL-1β-induced production of pro-inflammatory cytokines, TNF-α and IL-6, were mitigated by PSP in chondrocytes. Besides, PSP also suppressed the production of matrix metalloproteinases (MMPs), including MMP-1, MMP-3 and MMP-13 in IL-1β-stimulated chondrocytes. Furthermore, the IL-1β-induced activation of NF-κB signaling pathway in chondrocytes was also prevented by PSP. These findings suggested that PSP alleviated IL-1β-induced inflammatory response in chondrocytes via inhibiting NF-κB signaling pathway.

TMEM165 a new player in proteoglycan synthesis: loss of TMEM165 impairs elongation of chondroitin- and heparan-sulfate glycosaminoglycan chains of proteoglycans and triggers early chondrocyte differentiation and hypertrophy

TMEM165 deficiency leads to skeletal disorder characterized by major skeletal dysplasia and pronounced dwarfism. However, the molecular mechanisms involved have not been fully understood. Here, we uncover that TMEM165 deficiency impairs the synthesis of proteoglycans by producing a blockage in the elongation of chondroitin-and heparan-sulfate glycosaminoglycan chains leading to the synthesis of proteoglycans with shorter glycosaminoglycan chains. We demonstrated that the blockage in elongation of glycosaminoglycan chains is not due to defect in the Golgi elongating enzymes but rather to availability of the co-factor Mn2+. Supplementation of cell with Mn2+ rescue the elongation process, confirming a role of TMEM165 in Mn2+ Golgi homeostasis. Additionally, we showed that TMEM165 deficiency functionally impairs TGFβ and BMP signaling pathways in chondrocytes and in fibroblast cells of TMEM165 deficient patients. Finally, we found that loss of TMEM165 impairs chondrogenic differentiation by accelerating the timing of Ihh expression and promoting early chondrocyte maturation and hypertrophy. Collectively, our results indicate that TMEM165 plays an important role in proteoglycan synthesis and underline the critical role of glycosaminoglycan chains structure in the regulation of chondrogenesis. Our data also suggest that Mn2+ supplementation may be a promising therapeutic strategy in the treatment of TMEM165 deficient patients.

Chronic Circadian Rhythm Disturbance Accelerates Knee Cartilage Degeneration in Rats Accompanied by the Activation of the Canonical Wnt/β-Catenin Signaling Pathway.

With the gradual deepening of understanding of systemic health and quality of life, the factors affecting osteoarthritis (OA) are not limited to mechanical injury, metabolic abnormality, age and obesity, etc., but circadian rhythm, which plays a non-negligible role in human daily life. The purpose of this study was to explore the molecular mechanism of chronic circadian rhythm disturbance (CRD) inducing cartilage OA-like degeneration. Rats with the anterior cruciate ligament excision transection (ACLT) were used to establish the early-stage OA model (6-week). The light/dark (LD) cycle shifted 12 h per week for 22 weeks in order to establish a chronic CRD model. BMAL1 knockdown (KD) and Wnt/β-catenin pathway inhibition were performed in chondrocytes. The contents of proinflammatory factors and OA biomarkers in serum and chondrocyte secretions were detected by ELISA. Pathological and immunohistochemical staining of articular cartilage indicated the deterioration of cartilage. WB and qPCR were used to evaluate the relationship between matrix degradation and the activation of Wnt/β-catenin signaling pathway in chondrocytes. We found that chronic CRD could cause OA-like pathological changes in knee cartilage of rats, accelerating cartilage matrix degradation and synovial inflammation. The expression of MMP-3, MMP-13, ADAMTS-4, and β-catenin increased significantly; BMAL1, Aggrecan, and COL2A1 decreased significantly in either LD-shifted cartilage or BMAL1-KD chondrocytes. The expression of β-catenin and p-GSK-3β elevated, while p-β-catenin and GSK-3β diminished. The inhibitor XAV-939 was able to mitigated the increased inflammation produced by transfected siBMAL1. Our study demonstrates that chronic CRD disrupts the balance of matrix synthesis and catabolic metabolism in cartilage and chondrocytes, and it is related to the activation of the canonical Wnt/β-catenin signaling pathway.

Cudratricusxanthone O Inhibits H2O2-Induced Cell Damage by Activating Nrf2/HO-1 Pathway in Human Chondrocytes.

Osteoarthritis (OA) is a common joint degenerative disease induced by oxidative stress in chondrocytes. Although induced-heme oxygenase-1 (HO-1) has been found to protect cells against oxygen radical damage, little information is available regarding the use of bioactive compounds from natural sources for regulating the HO-1 pathway to treat OA. In this study, we explored the inhibitory effects of cudratricusxanthone O (CTO) isolated from the Maclura tricuspidata Bureau (Moraceae) on H2O2-induced damage of SW1353 chondrocytes via regulation of the HO-1 pathway. CTO promoted HO-1 expression by enhancing the translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) into the nucleus without inducing toxicity. Pretreatment with CTO-regulated reactive oxygen species (ROS) production by inducing expression of antioxidant enzymes in H2O2-treated cells and maintained the functions of H2O2-damaged chondrocytes. Furthermore, CTO prevented H2O2-induced apoptosis by regulating the expression of anti-apoptotic proteins. Treatment with the HO-1 inhibitor tin-protoporphyrin IX revealed that these protective effects were exerted due to an increase in HO-1 expression induced by CTO. In conclusion, CTO protects chondrocytes from H2O2-induced damages—including ROS accumulation, dysfunction, and apoptosis through activation of the Nrf2/HO-1 signaling pathway in chondrocytes and, therefore, is a potential therapeutic agent for OA treatment.

Silencing UHRF1 enhances cell autophagy to prevent articular chondrocytes from apoptosis in osteoarthritis through PI3K/AKT/mTOR signaling pathway

Osteoarthritis (OA) is a common chronic degenerative joint disease, and chondrocyte apoptosis is one of most important pathological changes of OA pathogenesis. Growing studies have shown that Ubiquitin-like with PHD and RING finger domains 1 (UHRF1) is an important epigenetic regulatory factor that regulates cell proliferation and apoptosis of various tumors, but its role in OA remains ill-defined. In the present study, we found that UHRF1 expression was increased in human OA cartilage tissues, compared with normal cartilage tissues. Interleukin-1β (IL-1β), a major inflammatory cytokine that promotes cartilage degradation in OA, was used to stimulate primary human chondrocytes in vitro. The expression of UHRF1 was also enhanced in IL-1β-induced chondrocytes. Moreover, down-regulation of UHRF1 induced an increase on cell proliferation and autophagy, and a decrease on apoptosis of chondrocytes after IL-1β treatment. Further data indicated that silencing UHRF1 attenuated the up-regulation of IL-1β on phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway in chondrocytes. Then, an activator of PI3K weakened the effect of UHRF1 silencing on cell proliferation, autophagy, apoptosis of IL-1β-induced chondrocytes, and the cell autophagy special inhibitor 3-methyladenine (3-MA) also showed a same impact on UHRF1, hence suggesting that knockdown of UHRF1 enhances cell autophagy to protect chondrocytes from apoptosis in OA through PI3K/AKT/mTOR signaling pathway. In conclusion, our study suggests that UHRF1 may be a potential regulator of chondrocyte apoptosis in the pathogenesis of OA.

UFL1 attenuates IL-1β-induced inflammatory response in human osteoarthritis chondrocytes

Osteoarthritis (OA) is a chronic inflammatory joint disease characterized by degradation of articular cartilage. Ubiquitin-fold modifier 1 (UFM1)-specific ligase 1 (UFL1) is an UFM1 E3 ligase that has been identified as a regulator of inflammatory response. However, the role of UFL1 in OA remains unknown. The aim of the present study was to explore the function of UFL1 in an in vitro OA system in chondrocytes. Our results showed that UFL1 was lowly expressed in both OA articular tissues and chondrocytes with IL-1β induction. Ectopic expression of UFL1 improved cell viability of IL-1β-induced chondrocytes. UFL1 suppressed the production of NO and PGE2, as well the expression levels of iNOS and COX-2 in IL-1β-induced chondrocytes. The IL-1β-induced increases in TNF-α and IL-6 levels were attenuated by UFL1. Ectopic expression of UFL1 inhibited the production of extracellular matrix (ECM) degrading enzymes including matrix metalloproteinase 3 (MMP-3), MMP-13, ADAMTS-4 and ADAMTS-5 in chondrocytes with IL-1β induction. Additionally, UFL1 suppressed IL-1β-induced activation of NF-κB signaling pathway in chondrocytes. In conclusion, these findings indicated that UFL1 exerted protective effect on IL-1β-induced chondrocytes. Thus, UFL1 might be a potential target for the treatment of OA.

A chondroprotective effect of moracin on IL-1β-induced primary rat chondrocytes and an osteoarthritis rat model through Nrf2/HO-1 and NF-κB axes.

Osteoarthritis (OA) is a common joint disease characterized by cartilage degeneration and inflammation. Although moracin is known to play a role in anti-inflammation and anti-oxidation in several inflammatory diseases, its anti-inflammatory effect on OA remains largely unknown. Therefore, in order to explore the role of moracin in OA, we investigated the anti-inflammatory effect of moracin on interleukin (IL)-β-induced rat chondrocytes in vitro and surgically induced OA rat models in vivo. Rat chondrocytes were pretreated using moracin (0, 5, 10, 15 μmol L-1) and then stimulated with IL-β (10 ng ml-1). Results showed that moracin reduced the expression of IL-1β-induced nitric oxide (NO), prostaglandin E2 (PGE2), tumor necrosis factor-α (TNF-α), IL-6, inducible nitric oxide synthase (iNOS), and cyclooxygenase (COX)-2 in both rat chondrocytes and cell culture supernatants. Besides, IL-1β-induced degradation of aggrecan and collage II, and the high expression of matrix metalloproteinase-13 (MMP-13) and a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS)-5 were also reversed by moracin. Moreover, moracin inhibited the translocation of p65 from the cytoplasm to nucleus induced by IL-1β and activated the Nrf2/HO-1 signaling pathway in chondrocytes. In OA rat models, moracin prevented cartilage of rats from destruction. All these findings above indicated that moracin could be a potentially effective drug for treating OA.

Effect of primary cilia expression rate on Wnt/β signaling pathway in tibial growth plate chondrocytes from chronic renal insufficiency young rats

Objective: To observe the effect of primary cilia expression rate on Wnt/β signaling pathway in tibial growth plate chondrocytes from chronic renal insufficiency (CRI) young rats. Methods: Male 2-week-old SD rats were randomly divided into two groups: (1) Sham group: only the left ureter was exposed (n=6); (2) CRI group: the left ureter was ligated (n=6). Rats were sacrificed 2 weeks after the operation and the primary cilia expression rate of growth plate chondrocytes and key protein β-catenin in Wnt/β signaling pathway were observed in histological section of tibia specimen. Chondrocytes isolated from growth plate in two groups were cultured in vitro to P3 generation. The primary cilia expression rate and the level of β-catenin were measured. The primary cilia expression rate was detected by agonists and antagonists Wnt/β signaling pathway in chondrocytes of CRI group. The level of β-catenin was detected by using serum-free culture and chloral hydrate to intervene chondrocytes in CRI group. Results: The primary cilia expression rate of growth plate chondrocytes in histological section of tibia specimen in CRI group was higher than that in Sham group [(17.5±7.7)% vs (8.7±3.6)%, t=3.237, P=0.005], and the level of β-catenin was higher in CRI group (5.1±0.7 vs 1.9±0.8, t=6.731, P<0.001). The primary cilia expression rate of growth plate chondrocytes cultured in vitro in CRI group was higher than that in Sham group [(20.9±8.1)% vs (11.8±4.7)%, t=3.073, P=0.007], and the level of β-catenin was higher in CRI group (0.49±0.12 vs 0.25±0.11, t=3.297, P=0.011). There was no significant change in primary cilia expression rate after intervention by using Wnt/β signaling agonists and antagonists to change the level of β-catenin [agonists group: (21.3±7.6)%, control: (20.6±6.8)%, antagonists group: (22.4±6.2)%, F=0.173, P=0.842]. The level of β-catenin was significantly changed after intervention by using serum-free culture, chloral hydrate to alter the primary cilia expression rate (serum-free culture group: 0.61±0.23, control: 0.39±0.24, chloral hydrate group: 0.15±0.11, F=6.476, P=0.012). There was a positive correlation between the level of β-catenin and primary cilia expression rate. Conclusion: The primary cilia expression rate and the level of Wnt/β signaling pathway were higher in tibial growth plate chondrocytes in CRI rats model, and primary cilia might have positive regulatory effects on the Wnt/β signaling pathway.

P-Coumaric Acid Attenuates IL-1β-Induced Inflammatory Responses and Cellular Senescence in Rat Chondrocytes.

Osteoarthritis (OA) is a common chronic inflammatory joint disease characterized by cartilage degradation. p-Coumaric acid (PCA), a dietary phenolic compound, has exerted anti-inflammatory and anti-oxidative activities in various diseases. However, the effects of PCA on OA have not been reported. In the present study, we aimed to investigate the effects of PCA on interleukin-1β(IL-1β)-induced inflammatory responses and cellular senescence in rat chondrocytes. Our results revealed that PCA remarkably downregulated IL-1β-induced inflammatory factors such as COX2 and iNOS and cartilage-degrading enzymes like matrix metalloproteinases (MMP1, MMP3, and MMP13) and aggrecanases (ADAMTS4 and ADAMTS5) in chondrocytes. The IL-1β-induced degradation of cartilage matrix (collagen II and aggrecan) could also be suppressed by PCA. Besides, PCA treatment effectively inhibited the IL-1β-induced p16INK4a protein expression and SAβ-gal activities in vitro. Mechanism analysis showed that PCA suppressed IL-1β-induced activation of mitogen-activated protein kinase (MAPK) and nuclear factor-kappaB (NF-κB) pathways. In vivo, we also found that PCA could alleviate the development of OA in a rat model. Altogether, our findings implicate that p-coumaric acid attenuates IL-1β-induced inflammatory responses and cellular senescence via inhibition of the MAPK and NF-κB signaling pathway in chondrocytes, and p-coumaric acid may be a promising candidate for the treatment of osteoarthritis.