Osteoarthritis Chondrocytes Research Articles

The Nrf2/HMGB1/NF-κB axis modulates chondrocyte apoptosis and extracellular matrix degradation in osteoarthritis.

Osteoarthritis (OA) is a degenerative or posttraumatic condition of the joints. In OA chondrocytes, Nrf2 functions as a stress response regulator with antioxidant and anti-inflammatory effects. This study aims to investigate the role of Nrf2 and its downstream pathway in the development of osteoarthritis. IL-1β treatment suppresses Nrf2, aggrecan, and COL2A1 levels and cell viability but promotes apoptosis in chondrocytes. IL-1β stimulation induces cell apoptosis, upregulates the mRNA expression of inflammatory factors, decreases aggrecan, COL2A1, and Bcl-2 levels but increases ADAMTS-5, ADAMTS-4, MMP13, cleaved caspase 3, and BAX levels, and promotes p65 phosphorylation. Nrf2 overexpression exerts opposite effects on IL-1β-treated chondrocytes, as demonstrated by the significant attenuation of IL-1β-induced changes in chondrocytes. By binding to the HMGB1 promoter region, Nrf2 suppresses HMGB1 expression. Similar to Nrf2 overexpression, HMGB1 knockdown also attenuates IL-1β-induced changes in chondrocytes. Notably, under IL-1β stimulation, the effects of Nrf2 overexpression or tert-butylhydroquinone (TBHQ, an activator of Nrf2) on apoptosis, inflammatory factor expression, ECM and apoptosis, and NF-κB pathway activity in chondrocytes are remarkably reversed by HMGB1 overexpression or recombinant HMGB1 (rHMGB1). Similarly, rHMGB1 could partially counteract the curative effect of TBHQ on OA damage in mice. In OA cartilage tissue samples, the level of Nrf2 is lower, while the levels of HMGB1, apoptotic, and inflammatory factors are increased compared to normal cartilage tissue samples. In conclusion, for the first time, the Nrf2/HMGB1 axis was found to modulate apoptosis, ECM degradation, inflammation and activation of NF-κB signaling in chondrocytes and OA mice.

Activation of the Mechanosensitive Ion Channels Piezo1 and TRPV4 in Primary Human Healthy and Osteoarthritic Chondrocytes Exhibits Ion Channel Crosstalk and Modulates Gene Expression.

Osteoarthritis (OA) is the most common degenerative joint disease causing pain and functional limitations. Physical activity as a clinically relevant, effective intervention alleviates pain and promotes joint function. In chondrocytes, perception and transmission of mechanical signals are controlled by mechanosensitive ion channels, whose dysfunction in OA chondrocytes is leading to disease progression. Signaling of mechanosensitive ion channels Piezo/TRPV4 was analyzed by Yoda1/GSK1016790A application and calcium-imaging of Fura-2-loaded chondrocytes. Expression analysis was determined by qPCR and immunofluorescence in healthy vs. OA chondrocytes. Chondrocytes were mechanically stimulated using the Flexcell™ technique. Yoda1 and GSK1016790A caused an increase in intracellular calcium [Ca2+]i for Yoda1, depending on extracellularly available Ca2+. When used concomitantly, the agonist applied first inhibited the effect of subsequent agonist application, indicating mutual interference between Piezo/TRPV4. Yoda1 increased the expression of metalloproteinases, bone-morphogenic protein, and interleukins in healthy and OA chondrocytes to a different extent. Flexcell™-induced changes in the expression of MMPs and ILs differed from changes induced by Yoda1. We conclude that Piezo1/TRPV4 communicate with each other, an interference that may be impaired in OA chondrocytes. It is important to consider that mechanical stimulation may have different effects on OA depending on its intensity.

A comprehensive analysis of single-cell RNA transcriptome reveals unique SPP1+ chondrocytes in human osteoarthritis

Osteoarthritis (OA) has become the most common degenerative disease in the world, which brings a serious economic burden to society and the country. Although epidemiological studies have shown that the occurrence of osteoarthritis is associated with obesity, sex, and trauma, the biomolecular mechanisms for the development and progression of osteoarthritis remain ambiguous. Several studies have drawn a connection between SPP1 and osteoarthritis. SPP1 was first found to be highly expressed in osteoarthritic cartilage, and later more studies have shown that SPP1 is also highly expressed in subchondral bone and synovial in OA patients. However, the biological function of SPP1 remains unclear. Single-cell RNA sequencing (scRNA-seq) is a novel technique that reflects gene expression at the cellular level, making it better depict the state of different cells than ordinary transcriptome data. However, most of the existing chondrocyte scRNA-seq studies focus on the occurrence and development of OA chondrocytes and lack analysis of normal chondrocyte development. Therefore, to better understand the mechanism of OA, scRNA-seq analysis of a larger cell volume containing normal and osteoarthritic cartilage is of great importance. Our study identifies a unique cluster of chondrocytes characterized by high SPP1 expression. The metabolic and biological characteristics of these clusters were further investigated. Besides, in animal models, we found that the expression of SPP1 is spatially heterogeneous in cartilage. Overall, our work provides novel insight into the potential role of SPP1 in OA, which sheds light on understanding the role of SPP1 in OA, promoting the progress of the treatment and prevention in the field of OA.

Yes-associated protein nuclear translocation promotes anabolic activity in human articular chondrocytes

Yes-associated protein (YAP) has been widely studied as a mechanotransducer in many cell types, but its function in cartilage is controversial. The aim of this study was to identify the effect of YAP phosphorylation and nuclear translocation on the chondrocyte response to stimuli relevant to osteoarthritis (OA). Cultured normal human articular chondrocytes from 81 donors were treated with increased osmolarity media as an in vitro model of mechanical stimulation, fibronectin fragments (FN-f) or IL-1β as catabolic stimuli, and IGF-1 as an anabolic stimulus. YAP function was assessed with gene knockdown and inhibition by verteporfin. Nuclear translocation of YAP and its transcriptional co-activator TAZ and site-specific YAP phosphorylation were determined by immunoblotting. Immunohistochemistry and immunofluorescence to detect YAP were performed on normal and OA human cartilage with different degrees of damage. Chondrocyte YAP/TAZ nuclear translocation increased under physiological osmolarity (400mOsm) and IGF-1 stimulation, which was associated with YAP phosphorylation at Ser128. In contrast, catabolic stimulation decreased the levels of nuclear YAP/TAZ through YAP phosphorylation at Ser127. Following YAP inhibition, anabolic gene expression and transcriptional activity decreased. Additionally, YAP knockdown reduced proteoglycan staining and levels of type II collagen. Total YAP immunostaining was greater in OA cartilage, but YAP was sequestered in the cytosol in cartilage areas with more severe damage. YAP chondrocyte nuclear translocation is regulated by differential phosphorylation in response to anabolic and catabolic stimuli. Decreased nuclear YAP in OA chondrocytes may contribute to reduced anabolic activity and promotion of further cartilage loss.

Tannic acid, an IL-1β-direct binding compound, ameliorates IL-1β-induced inflammation and cartilage degradation by hindering IL-1β-IL-1R1 interaction.

Interleukin-1β (IL-1β) is one of the most potent pro-inflammatory cytokines implicated in a wide range of autoinflammatory, autoimmune, infectious, and degenerative diseases. Therefore, many researchers have focused on developing therapeutic molecules that inhibit IL-1β-IL-1 receptor 1 (IL-1R1) interaction for the treatment of IL-1-related diseases. Among IL-1-related diseases, osteoarthritis (OA), is characterized by progressive cartilage destruction, chondrocyte inflammation, and extracellular matrix (ECM) degradation. Tannic acid (TA) has been proposed to have multiple beneficial effects, including anti-inflammatory, anti-oxidant, and anti-tumor activities. However, it is unclear whether TA plays a role in anti-IL-1β activity by blocking IL-1β-IL-1R1 interaction in OA. In this study, we report the anti-IL-1β activity of TA in the progression of OA in both in vitro human OA chondrocytes and in vivo rat OA models. Herein, using-ELISA-based screening, natural compound candidates capable of inhibiting the IL-1β-IL-1R1 interaction were identified. Among selected candidates, TA showed hindering IL-1β-IL-1R1 interaction by direct binding to IL-1β using surface plasmon resonance (SPR) assay. In addition, TA inhibited IL-1β bioactivity in HEK-Blue IL-1-dependent reporter cell line. TA also inhibited IL-1β-induced expression of inducible nitric oxide synthase (NOS2), cyclooxygenase-2 (COX-2), IL-6, tumor necrosis factor-alpha (TNF-α), nitric oxide (NO), and prostaglandin E2 (PGE2) in human OA chondrocytes. Moreover, TA downregulated IL-1β-stimulated matrix metalloproteinase (MMP)3, MMP13, ADAM metallopeptidase with thrombospondin type 1 motif (ADAMTS)4, and ADAMTS5, while upregulating collagen type II (COL2A1) and aggrecan (ACAN). Mechanistically, we confirmed that TA suppressed IL-1β-induced MAPK and NF-κB activation. The protective effects of TA were also observed in a monosodium iodoacetamide (MIA)-induced rat OA model by reducing pain and cartilage degradation and inhibiting IL-1β-mediated inflammation. Collectively, our results provide evidence that TA plays a potential role in OA and IL-1β-related diseases by hindering IL-1β-IL-1R1 interaction and suppressing IL-1β bioactivity.

Differential Effects of Hypoxia versus Hyperoxia or Physoxia on Phenotype and Energy Metabolism in Human Chondrocytes from Osteoarthritic Compared to Macroscopically Normal Cartilage.

Chondrocyte phenotype and energy metabolism are altered in osteoarthritis (OA). However, most studies characterising the change in human chondrocyte behaviour in OA have been conducted in supraphysiological oxygen concentrations. The purpose of this study was to compare phenotype and energy metabolism in chondrocytes from macroscopically normal (MN) and OA cartilage maintained in 18.9% (standard tissue culture), 6% (equivalent to superficial zone of cartilage in vivo) or 1% oxygen (equivalent to deep zone of cartilage in vivo). MMP13 production was higher in chondrocytes from OA compared to MN cartilage in hyperoxia and physoxia but not hypoxia. Hypoxia promoted SOX9, COL2A1 and ACAN protein expression in chondrocytes from MN but not OA cartilage. OA chondrocytes used higher levels of glycolysis regardless of oxygen availability. These results show that differences in phenotype and energy metabolism between chondrocytes from OA and MN cartilage differ depending on oxygen availability. OA chondrocytes show elevated synthesis of cartilage-catabolising enzymes and chondrocytes from MN cartilage show reduced cartilage anabolism in oxygenated conditions. This is relevant as a recent study has shown that oxygen levels are elevated in OA cartilage in vivo. Our findings may indicate that this elevated cartilage oxygenation may promote cartilage loss in OA.

Unravelling the Basic Calcium Phosphate crystal-dependent chondrocyte protein secretome; a role for TGF-β signaling

Basic Calcium Phosphate (BCP) crystals play an active role in the progression of osteoarthritis (OA). However, the cellular consequences remain largely unknown. Therefore, we characterized for the first time the changes in the protein secretome of human OA articular chondrocytes as a result of BCP stimulation using two unbiased proteomic analysis methods. Isolated human OA articular chondrocytes were stimulated with BCP crystals and examined by Quantitative Reverse Transcription PCR (RT-qPCR) and enzyme-linked immune sorbent assay (ELISA) after twenty-four and forty-eight hours. Forty-eight hours conditioned media were analyzed by label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) and an antibody array. The activity of BCP dependent Transforming Growth Factor Beta (TGF-β) signaling was analyzed by RT-qPCR and luciferase reporter assays. The molecular consequences regarding BCP-dependent TGF-β signaling on BCP-dependent Interleukin 6 (IL-6) were investigated using specific pathway inhibitors. Synthesized BCP crystals induced IL-6 expression and secretion upon stimulation of human articular chondrocytes. Concomitant induction of catabolic gene expression was observed. Analysis of conditioned media revealed a complex and diverse response with a large number of proteins involved in TGF-β signaling, both in activation of latent TGF-β and TGF-β superfamily members, which were increased compared to non-stimulated OA chondrocytes. Activity of this BCP driven TGF-β signaling was confirmed by increased activity of expression of TGF-β target genes and luciferase reporters. Inhibition of BCP driven TGF-β signaling resulted in decreased IL-6 expression and secretion with a moderate effect on catabolic gene expression. BCP crystal stimulation resulted in a complex and diverse chondrocyte protein secretome response. An important role for BCP-dependent TGF-β signaling was identified in development of a pro-inflammatory environment.

CELL MORPHOLOGY AS A BIOLOGICAL FINGERPRINT FOR DESCRIBING CHONDROCYTE PHENOTYPE UNDER INFLAMMATORY ATTACK

While cell morphology has been recognized as a fundamental regulator of cell behavior, few studies have measured the complex cell morphological changes of chondrocytes using quantitative cell morphometry descriptors in relation to inflammation and phenotypic outcome.Acute vs. persistent exposure to IL-1β and how IL-1β modulated dynamic changes in cell morphology in relation to the phenotype, donor and OA grade in healthy and osteoarthritis (OA) chondrocytes was investigated. A panel of quantitative cell morphometry descriptors was measured using an automated high-throughput method. Absolute quantification of gene expression was measured by ddPCR followed by correlation analyses.In OA chondrocytes, chronic IL-1β significantly decreased COL2A1, SOX9, and ACAN, increased IL-6 and IL-8 levels and caused chondrocytes to become less wide, smaller, longer, slimmer, less round and more circular, consistent with a de-differentiated phenotype. In healthy chondrocytes, 3 days after acute (72 h) IL-1β exposure, COL1A2 and IL-6 significantly increased but had minor effects on cell morphology. However, in healthy chondrocytes, persistent IL-1β led to more profound effects in all cell morphology descriptors and chondrocytes expressed significantly less COL2A1 and more IL-6 and IL-8 vs. controls and acutely-stimulated chondrocytes. In both OA and healthy chronically-stimulated chondrocytes, area, width and circularity were sensitive to the persistent presence of the IL-1β cytokine. Moreover, there were many significant and strong correlations among the measured parameters, with several indications of an IL-1β-mediated mechanism.Cell morphology combined with gene expression analysis could guide researchers interested in understanding inflammatory effects in the complex domain of cartilage/chondrocyte biology. Use of quantitative cell morphometry could complement classical approaches by providing numerical data on a large number of cells, thereby providing a biological fingerprint for describing chondrocyte phenotype, which could help to understand how changes in cell morphology lead to disease progression.

DNA METHYLATION AND ITS INFLUENCE ON THE PATHOGENESIS OF OSTEOARTHRITIS: A SYSTEMATIC LITERATURE REVIEW

There is an evolving body of evidence that demonstrates the role of epigenetic mechanisms, such as DNA-methylation in the pathogenesis of OA. This systematic review aims to summarize the current evidence of DNA methylation and its influence on the pathogenesis of OA.A pre-defined protocol in alignment with the PRISMA guidelines was employed to systematically review eight bibliographic databases, to identify associations between DNA-methylation of articular chondrocytes and osteoarthritis. A search of Medline (Ovid), Embase, Web-of-Science, Scopus, PubMed, Cinahl (EBSCOhost), Cochrane Central and Google Scholar was performed between 1st January 2015 to 31st January 2021. Data extraction was performed by two independent reviewers.During the observation period, we identified 15 gene specific studies and 24 genome wide methylation analyses. The gene specific studies mostly focused on the expression of pro-inflammatory markers, such as IL8 and MMP13 which are overexpressed in OA chondrocytes. DNA hypomethylation in the promoter region resulted in overexpression, whereas hypermethylation was seen in non-OA chondrocytes. Others reported on the association between OA risk genes and the DNA methylation pattern close to RUNX2, which is an important OA signal. The genome wide methylation studies reported mostly on differentially methylated regions comparing OA chondrocytes and non-OA chondrocytes. Clustering of the regions identified genes that are involved in skeletal morphogenesis and development. Differentially methylated regions were seen in hip OA and knee OA chondrocytes, and even within different regions of an OA affected knee joint, differentially methylated regions were identified depending on the disease stage.This systematic review demonstrates the growing evidence of epigenetic mechanisms, such as DNA methylation, in the pathogenesis of OA. In recent years, there has been a focus on the interplay between OA risk genes and DNA methylation changes which revealed a reactivation of genes responsible for endochondral ossification during development. These are important findings and may help to identify eventual future therapeutic targets. However, the current body of literature is mostly showing the differences in DNA methylation of OA chondrocytes and non-OA chondrocytes, but a true longitudinal analysis demonstrating the DNA methylation changes actually happening is still not available.

Dihydrocaffeic acid improves IL-1β-induced inflammation and cartilage degradation via inhibiting NF-κB and MAPK signalling pathways.

Osteoarthritis (OA) is a prevalent joint disorder with inflammatory response and cartilage deterioration as its main features. Dihydrocaffeic acid (DHCA), a bioactive component extracted from natural plant (gynura bicolor), has demonstrated anti-inflammatory properties in various diseases. We aimed to explore the chondroprotective effect of DHCA on OA and its potential mechanism. In vitro, interleukin-1 beta (IL-1β) was used to establish the mice OA chondrocytes. Cell counting kit-8 evaluated chondrocyte viability. Western blotting analyzed the expression levels of collagen II, aggrecan, SOX9, inducible nitric oxide synthase (iNOS), IL-6, matrix metalloproteinases (MMPs: MMP1, MMP3, and MMP13), and signalling molecules associated with nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways. Immunofluorescence analysis assessed the expression of aggrecan, collagen II, MMP13, and p-P65. In vivo, a destabilized medial meniscus (DMM) surgery was used to induce mice OA knee joints. After injection of DHCA or a vehicle into the injured joints, histological staining gauged the severity of cartilage damage. DHCA prevented iNOS and IL-6 from being upregulated by IL-1β. Moreover, the IL-1β-induced upregulation of MMPs could be inhibited by DHCA. Additionally, the administration of DHCA counteracted IL-1β-induced downregulation of aggrecan, collagen II, and SOX9. DHCA protected articular cartilage by blocking the NF-κB and MAPK pathways. Furthermore, DHCA mitigated the destruction of articular cartilage in vivo. We present evidence that DHCA alleviates inflammation and cartilage degradation in OA chondrocytes via suppressing the NF-κB and MAPK pathways, indicating that DHCA may be a potential agent for OA treatment.

PGC-1α in osteoarthritic chondrocytes: From mechanism to target of action.

Osteoarthritis (OA) is one of the most common degenerative joint diseases, often involving the entire joint. The degeneration of articular cartilage is an important feature of OA, and there is growing evidence that the mitochondrial biogenesis master regulator peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) exert a chondroprotective effect. PGC-1α delays the development and progression of OA by affecting mitochondrial biogenesis, oxidative stress, mitophagy and mitochondrial DNA (mtDNA) replication in chondrocytes. In addition, PGC-1α can regulate the metabolic abnormalities of OA chondrocytes and inhibit chondrocyte apoptosis. In this paper, we review the regulatory mechanisms of PGC-1α and its effects on OA chondrocytes, and introduce potential drugs and novel nanohybrid for the treatment of OA which act by affecting the activity of PGC-1α. This information will help to further elucidate the pathogenesis of OA and provide new ideas for the development of therapeutic strategies for OA.

The Effect of CaV1.2 Inhibitor Nifedipine on Chondrogenic Differentiation of Human Bone Marrow or Menstrual Blood-Derived Mesenchymal Stem Cells and Chondrocytes

Cartilage is an avascular tissue and sensitive to mechanical trauma and/or age-related degenerative processes leading to the development of osteoarthritis (OA). Therefore, it is important to investigate the mesenchymal cell-based chondrogenic regenerating mechanisms and possible their regulation. The aim of this study was to investigate the role of intracellular calcium (iCa2+) and its regulation through voltage-operated calcium channels (VOCC) on chondrogenic differentiation of mesenchymal stem/stromal cells derived from human bone marrow (BMMSCs) and menstrual blood (MenSCs) in comparison to OA chondrocytes. The level of iCa2+ was highest in chondrocytes, whereas iCa2+ store capacity was biggest in MenSCs and they proliferated better as compared to other cells. The level of CaV1.2 channels was also highest in OA chondrocytes than in other cells. CaV1.2 antagonist nifedipine slightly suppressed iCa2+, Cav1.2 and the proliferation of all cells and affected iCa2+ stores, particularly in BMMSCs. The expression of the CaV1.2 gene during 21 days of chondrogenic differentiation was highest in MenSCs, showing the weakest chondrogenic differentiation, which was stimulated by the nifedipine. The best chondrogenic differentiation potential showed BMMSCs (SOX9 and COL2A1 expression); however, purposeful iCa2+ and VOCC regulation by blockers can stimulate a chondrogenic response at least in MenSCs.

Vitamin D reduces pain and cartilage destruction in knee osteoarthritis animals through inhibiting the matrix metalloprotease (MMPs) expression

Aim of the studyIn this study, we investigated the therapeutic potential of vitamin D (VITD) in OA Wistar rats induced by anterior cruciate ligament transection combined with medial meniscectomy (ACLT + MMx). In ACLT + MMx-induced OA rats, pain severity, cartilage destruction, inflammatory cytokines, and MMPs were all measured. Materials and methodsACLT + MMx methods were used to induce OA, and pain behavioral studies such as the weight bearing test and paw withdrawal test were performed while the knee width and body weights were also measured. Furthermore, Hematoxylin and Eosin (H&E) staining was used to determine knee histopathological studies, as well as OARSI scoring, cartilage thickness, cartilage width, and cartilage degradation scores. The enzyme-linked immunosorbent assay (ELISA) studies were used to check the serum levels of VITD, C-telopeptide of Type II collagen (CTX-II), and pro-inflammatory cytokines tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), and anti-inflammatory cytokines interleukin-10 (IL-10), and MMPs (MMP-3, MMP-9, and MMP-13). Finally, the reverse transcription polymerase chain reaction (RT-PCR) test was used to determine the levels of MMPs, nuclear factor-kappa B (NF-κB), TNF-α, IL-6, and IL-10 in IL-1β stimulated chondrocytes. ResultsThe oral VITD supplement significantly reduced OA pain, inflammation, cartilage destruction, and MMPs levels. Furthermore, serum VITD levels increased while CTX-II levels decreased, indicating that VITD reduced cartilage degradation effectively. Moreover, VITD supplementation reduced the expression of pro-inflammatory TNF-α, IL-1β, and IL-6 cytokines while increasing the expression of anti-inflammatory IL-10. The elevation of MMPs after ACLT + MMx surgery contributed to articular cartilage destruction, which was reduced by VITD supplementation. Finally, VITD supplementation significantly reduces serum levels of MMPs, IL-1β, TNF-α, and IL-6 while increasing IL-10 levels. Then, using the in-vitro cytotoxicity (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) MTT assay, examine the cytotoxicity profile of VITD in rat chondrocytes after stimulated with IL-1β, which shows no toxicity in the dose range of VITD 0–500 IU. Finally, RT-PCR studies in IL-1β stimulated rat chondrocytes revealed that VITD (50, 100, and 500 IU) significantly reduced the mRNA levels of MMPs, NF-κB, TNF-α, and IL-6, while increasing IL-10 levels, indicating that VITD reduced chondrocyte destruction and overcame harsh conditions in a dose-dependent manner. ConclusionOverall, the in vivo and in vitro findings show that VITD effectively reduces OA pain, inflammation, and chondrocyte destruction by lowering MMPs levels specifically.

SPRED2 promotes autophagy and attenuates inflammatory response in IL-1β induced osteoarthritis chondrocytes via regulating the p38 MAPK signaling pathway

Osteoarthritis (OA) is an age-related degenerative disease primarily characterized by articular cartilage degeneration. Many inflammatory mediators are upregulated in OA patients. Mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) pathways play a role in the regulation of inflammatory response. Autophagy appears to exhibit a protective mechanism, and alleviate the symptoms of OA in rats. Dysregulation of SPRED2 is associated with various diseases involving inflammatory response. However, the role of SPRED2 in OA development remains to be investigated. The present work demonstrated that SPRED2 promoted autophagy and attenuated inflammatory response in IL-1β induced osteoarthritis chondrocytes via regulating the p38 MAPK signaling pathway. SPRED2 was downregulated in human knee cartilage tissues of OA patients and in IL-1β-induced chondrocytes. SPRED2 enhanced chondrocyte proliferation and prevented cell apoptosis induced by IL-1β. SPRED2 prevented IL-1β-induced chondrocytes autophagy and inflammatory response in chondrocytes. SPRED2 inhibited the activation of p38 MAPK signaling pathway and ameliorated OA injury of cartilage. Thus, SPRED2 promoted autophagy and inhibited inflammatory response by regulation of p38 MAPK signaling pathway in vivo.

Inhibition of miR-182-5p Targets FGF9 to Alleviate Osteoarthritis.

The pathogenesis of osteoarthritis (OA) is complex and there is no specific drug for treatment. The aim of this study was to identify the molecular targets of OA therapy, focusing on the expression and biological functions of miR-182-5p and its target genes in OA. miR-182-5p and fibroblast growth factor 9 (FGF9) were overexpressed or knocked down in IL-1β-induced chondrocytes. An OA knee model was performed by surgically destroying the medial meniscus. The gene expression of miR-182-5p and FGF9 was calculated. The protein FGF9 was tested by western blotting. Cell counting kit-8 (CCK8), plate cloning assay, and flow cytometry were conducted to evaluate cell proliferation and apoptosis. The expression of inflammatory factors, tumor necrosis factor-alpha (TNF-α), interleukin (IL)-6, and interleukin (IL)-8, was evaluated using enzyme-linked immunosorbent assay (ELISA). Dual-luciferase reporter assays validated the targeting relationship between miR-182-5p and FGF9. Hematoxylin-eosin (HE) and safranin O-fast Green (S-O) staining were utilized to access cartilage damage. Ki67 expression in cartilage was detected using immunohistochemistry (IHC). TdT-mediated dUTP nick-end labeling (TUNEL) assays were used to calculate the apoptosis rate of cartilage. The expression of miR-182-5p was upregulated, and FGF9 was downregulated in the IL-1β-induced chondrocytes. OA chondrocytes proliferation ability in the miR-182-5p mimics group was decreased, and the apoptosis rate and inflammatory factor were increased. Transfection with miR-182-5p inhibitor increased the proliferative ability and decreased the apoptosis rate in the IL-1β-induced chondrocytes. Transfection with miR-182-5p inhibitor reversed IL-1β-induced inflammatory factor release in chondrocytes. Targeted binding sites existed between miR-182-5p and FGF9. After overexpression of FGF9, the miR-182-5p effect on OA chondrocytes was reversed. The hyaline cartilage thickness and proteoglycan content decreased in OA rats, and this was reversed by miR-182-5p inhibitor treatment. miR-182-5p expression levels were increased in OA chondrocytes and regulated chondrocyte proliferation, apoptosis, and inflammation by targeting FGF9. miR-182-5p is a potential gene for OA treatment.

Retraction: MicroRNA-181 inhibits proliferation and promotes apoptosis of chondrocytes in osteoarthritis by targeting PTEN.

Retraction: MicroRNA-181 inhibits proliferation and promotes apoptosis of chondrocytes in osteoarthritis by targeting PTEN.

Apigenin Inhibits the Progression of Osteoarthritis by Mediating Macrophage Polarization.

The overall purpose of this study was to investigate the mechanism of macrophage polarization on chondrocyte injury in osteoarthritis and the protective effect of apigenin on chondrocytes in osteoarthritis. Primary chondrocytes were isolated from the knee cartilage of three-day-old mice, and cells positive for Alsine blue staining and type II collagen immunocytochemical staining were identified and used in followup experiments. Transwell coculture was performed. Chondrocytes were inoculated in the inferior compartment, and macrophages were inoculated in the upper compartment. The experimental groups were the N group, LPS group, and LPS+ apigenin group. The effect of macrophage polarization on chondrocyte inflammation and the protective effect of apigenin on chondrocytes were verified by the drug administration. Real-time quantitative PCR (qPCR) and Western blot were used to detect the expression of RNA and protein. Experimental OA was induced by modified Hulth surgery in mice. Modified Hulth surgery was performed on the mouse's right knee to induce experimental osteoarthritis in mice, with the nonoperative right knee serving as an ipsilateral control. The mice were randomly assigned to three groups (six mice per group): the sham group, the modified Hulth group, and the modified Hulth + apigenin group. Animals were given gavage for four weeks. The protective effect of apigenin on articular cartilage was verified by histological staining and immunohistochemical analysis. Histological staining showed that apigenin had a protective effect on cartilage degeneration induced by modified Hulth surgery. The PCR results showed that apigenin significantly reduced the expression levels of IL-1, IL-6, MMP3, and MMP13 in the articular cartilage of OA mice, and it had a protective effect on articular cartilage. Apigenin reduced the levels of IL-1, IL-6, TNF-α, and IL-12 in macrophages and increased the levels of MG-L1, MG-L2, ARG-1, and IL-10, which can inhibit the M1 polarization of macrophages and promote M2 polarization. In the coculture system, apigenin decreased the protein levels of TRPM7, P-mTOR, BAX, and c-caspase3 in macrophages, while significantly increasing the protein levels of Bcl2. The levels of IL-1, IL-6, MMP13, TNF-α, P38, JNK, and ERK phosphorylation were reduced in chondrocytes. Apigenin alleviates cartilage injury in OA mice induced by modified Hulth. Apigenin inhibits chondrocyte inflammation through the MAPK pathway. Apigenin alleviates macrophage-polarization-induced inflammatory response and chondrocyte apoptosis in the macrophage-chondrocyte coculture system through the TRPM7-mTOR pathway.

Promotion of Knee Cartilage Degradation by IκB Kinase ε in the Pathogenesis of Osteoarthritis in Human and Murine Models.

NF-κB signaling is an important modulator in osteoarthritis (OA), and IκB kinase ε (IKKε) regulates the NF-κB pathway. This study was undertaken to identify the functional involvement of IKKε in the pathogenesis of OA and the effectiveness of IKKε inhibition as a modulatory treatment. IKKε expression in normal and OA human knee joints was analyzed immunohistochemically. Gain- or loss-of-function experiments were performed using human chondrocytes. Furthermore, OA was surgically induced in mice, followed by intraarticular injection of BAY-985, an IKKε/TANK-binding kinase 1 inhibitor, into the left knee joint every 5 days for 8 weeks. Mice were subsequently examined for histologic features of cartilage damage and inflammation. IKKε protein expression was increased in human OA cartilage. In vitro, expression levels of OA-related factors were down-regulated following knockdown of IKKε with the use of small interfering RNA in human OA chondrocytes or following treatment with BAY-985. Conversely, IKKε overexpression significantly increased the expression of OA-related catabolic mediators. In Western blot analysis of human chondrocytes, IKKε overexpression increased the phosphorylation of IκBα and p65. In vivo, intraarticular injection of BAY-985 into the knee joints of mice attenuated OA-related cartilage degradation and hyperalgesia via NF-κB signaling. These results suggest that IKKε regulates cartilage degradation through a catabolic response mediated by NF-κB signaling, and this could represent a potential target for OA treatment. Furthermore, BAY-985 may serve as a major disease-modifying compound among the drugs developed for OA.

Neo-cartilage formation using human nondegenerate versus osteoarthritic chondrocyte-derived cartilage organoids in a viscoelastic hydrogel.

Current regenerative cartilage therapies are associated with several drawbacks such as dedifferentiation of chondrocytes during expansion and the formation of fibrocartilage. Optimized chondrocyte expansion and tissue formation could lead to better clinical results of these therapies. In this study, a novel chondrocyte suspension expansion protocol that includes the addition of porcine notochordal cell-derived matrixwas used to self-assemble human chondrocytes from osteoarthritic (OA) and nondegenerate (ND) origin into cartilage organoids containing collagen type II and proteoglycans. Proliferation rate and viability were similar for OA and ND chondrocytes and organoids formed had a similar histologic appearance and gene expression profile. Organoids were then encapsulated in viscoelastic alginate hydrogels to form larger tissues. Chondrocytes on the outer bounds of the organoids produced a proteoglycan-rich matrix to bridge the space between organoids. In hydrogels containing ND organoids some collagen type I was observed between the organoids. Surrounding the bulk of organoids in the center of the gels, in both OA and ND gels a continuous tissue containing cells, proteoglycans and collagen type II had been produced. No difference was observed in sulphated glycosaminoglycan and hydroxyproline content between gels containing organoids from OA or ND origin after 28 days. It was concluded that OA chondrocytes, which can be harvested from leftover surgery tissue, perform similar to ND chondrocytes in terms of human cartilage organoid formation and matrix production in alginate gels. This opens possibilities for their potential to serve as a platform for cartilage regeneration but also as an in vitro model to study pathways, pathology, or drug development.

The Role of Ferroptosis in the Pathogenesis of Osteoarthritis.

Osteoarthritis (OA) is the most common type of arthritis. Its high prevalence, especially in the elderly, and its negative impact on physical function make it a leading cause of disability in the elderly. Joint pain as well joint stiffness are the common classic signs of OA. Chondrocyte death together with loss of articular cartilage integrity are the main pathologic changes in OA. Non-steroidal anti-inflammatory drugs (NSAIDs) and glucocorticoids are commonly used for the management of OA; still, their effectiveness is limited, and no therapeutic strategy is able to fully stop OA progression. Ferroptosis is a kind of cell death, distinct from apoptosis and necroptosis, caused by iron-dependent peroxidation of membrane phospholipids that terminates cell life by disintegrating all plasma membranes. It has been suggested that ferroptosis has a critical role in decreased viability of chondrocytes in OA, and here, we review recent findings regarding the pathologic pathways that lead to chondrocyte ferroptosis, and discuss the possible therapeutic utility of ferroptosis inhibition in OA.