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Inhibition of syndecan-4 reduces cartilage degradation in murine models of osteoarthritis through the downregulation of HIF-2α by miR-96-5p.

ABSTRACT: The membranous receptor syndecan-4 (SDC-4) and the nuclear transcription factor hypoxia-induced factor-2α (HIF-2α) play critical roles in the pathogenesis of osteoarthritis (OA). The aim of this study was to determine whether SDC-4 inhibition downregulates HIF-2a expression by microRNA-96-5p (miR-96-5p) in murine chondrocyte and cartilage tissue. The OA model was induced surgically in mice, and SDC-4 polyclonal antibody, HIF-2α small interfering RNA (siRNA) and its control, miR-96-5p mimics and its scrambled controls or anti-miR-96-5p and its control were then injected into the knee joints. At 2 and 4 weeks after surgery, OA progression was evaluated microscopically, histologically, radiographically and immunohistochemically in these mice. Real-time polymerase chain reaction (RT-PCR) and western blotting were performed after treating with antibody and transfecting with miRNA mimic or siRNA to determine their effects on OA-related mediators. The potential miRNAs related to OA development were identified by using miRNA microarray analysis. Whether miRNAs play a pivotal role in OA development in vivo or in vitro was also investigated. MiR-96-5p expression was upregulated by SDC-4-specific antibodies in chondrocytes and cartilage tissue, and miR-96-5p directly targeted the 3'-UTR of HIF-2α to inhibit HIF-2α signaling in murine chondrocytes. Moreover, we demonstrated that anti-SDC-4-attenuated IL-1β-induced chondrocyte hypertrophy and cartilage degradation by inhibiting HIF-2α signaling by a miR-96-5p-dependent mechanism. Our study revealed that the inhibition of SDC-4 exerts its effects on both cartilage homeostasis and the chondrocyte hypertrophy phenotype by inducing miR-96-5p expression, which results in targeting HIF-2α 3'-UTR sequences and inhibiting HIF-2α in murine cartilage tissue and chondrocytes.

SUBMITTER: Zhou K

PROVIDER: S-EPMC8292145 | biostudies-literature | 2021 Aug

REPOSITORIES: biostudies-literature

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Inhibition of syndecan-4 reduces cartilage degradation in murine models of osteoarthritis through the downregulation of HIF-2α by miR-96-5p.

Zhou Kai K He Sirong S Yu Haoda H Pei Fuxing F Zhou Zongke Z

Laboratory investigation; a journal of technical methods and pathology 20210413 8

The membranous receptor syndecan-4 (SDC-4) and the nuclear transcription factor hypoxia-induced factor-2α (HIF-2α) play critical roles in the pathogenesis of osteoarthritis (OA). The aim of this study was to determine whether SDC-4 inhibition downregulates HIF-2a expression by microRNA-96-5p (miR-96-5p) in murine chondrocyte and cartilage tissue. The OA model was induced surgically in mice, and SDC-4 polyclonal antibody, HIF-2α small interfering RNA (siRNA) and its control, miR-96-5p mimics and ...[more]

PMID: 33850295

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Project description:The disorders of cartilage homeostasis and chondrocyte apoptosis are major events in the pathogenesis of osteoarthritis (OA). Herein, we aim to assess the chondroprotective effect and underlying mechanisms of a novel chemically modified curcumin, CMC2.24, in modulating extracellular matrix (ECM) homeostasis and inhibiting chondrocyte apoptosis. Rats underwent the anterior cruciate ligament transection, and medial menisci resection was treated by intra-articular injection with CMC2.24. In an in vitro study, rat chondrocytes were pretreated with CMC2.24 before stimulation with sodium nitroprusside (SNP). Results from in vivo studies demonstrated that the intra-articular administration of CMC2.24 ameliorated osteoarthritic cartilage destruction by promoting collagen 2a1 production and inhibited cartilage degradation and apoptosis by suppressing hypoxia-inducible factor-2a (Hif-2α), matrix metalloproteinase-3, runt-related transcription factor 2, cleaved caspase-3, and vascular endothelial growth factor and the phosphorylation of IκBα and NF-κB p65. The in vitro results revealed that CMC2.24 exhibited a strong inhibitory effect on SNP-induced chondrocyte catabolism and apoptosis. The SNP-enhanced expression of Hif-2α, a catabolic and apoptotic factor, decreased in a dose-dependent manner after CMC2.24 treatment. CMC2.24 pretreatment effectively inhibited SNP-induced IκBα and NF-κB p65 phosphorylation in rat chondrocytes, whereas pretreatment with the NF-κB antagonist BMS-345541 significantly enhanced the effects of CMC2.24. Overall, these results demonstrated that CMC2.24 attenuates OA progression by modulating ECM homeostasis and chondrocyte apoptosis by suppressing the NF-κB/Hif-2α axis, thus providing a new perspective for therapeutic strategies in OA. KEY MESSAGES: • Intra-articular injection of CMC2.24 ameliorated osteoarthritic cartilage destruction. • CMC2.24 promoted cell viability and decreased SNP-induced apoptotic gene expression. • SNP-induced activation of Hif-2α is inhibited by CMC2.24. • CMC2.24 inhibits NF-κB/Hif-2α axis activation to modulate ECM homeostasis and inhibit chondrocyte apoptosis.

Project description:BackgroundChondrocyte hypertrophy is a potential target for osteoarthritis (OA) treatment, with Indian hedgehog (IHH), glioma-associated oncogene homolog (GLI), and hypoxia-inducible factor-2α (HIF-2α) being closely associated with chondrocyte hypertrophy during OA progression. Whereas IHH can modulate chondrocyte hypertrophy, interference with IHH signalling has not achieved the anticipated therapeutic effects and poses safety concerns, necessitating further clarification of the specific mechanisms by which IHH affects articular cartilage degeneration. Inhibition of the HIF-2α overexpression in cartilage slows the progression of early OA, but the mechanisms underlying HIF-2α accumulation in OA cartilage remain unclear. The aim of this study was to determine the function of Ihh, as well as its downstream factors, in chondrocytes, based on an early osteoarthritis (OA) mouse model and in vitro chondrocyte model.MethodsInvestigated the expression levels and locations of IHH-GLI-1 pathway in normal and early degenerated human cartilage, comparing them with HIF-2α and its downstream factors. RT-qPCR, Western blotting, Crystal violet staining, and EdU assays were used to evaluate the pecific regulatory mechanisms of the IHH-GLI-1-HIF-2α signalling axis in normal chondrocytes and in chondrocytes under inflammatory conditions. Validated the impact of IHH on early cartilage degeneration and the relationship between the IHH-GLI-1 pathway and the expression levels and expression locations of HIF-2α and its downstream factors in Col2a1-CreERT2;Ihhfl/fl mice.ResultsIn early-stage degenerative joint cartilage, the GLI-1 pathway in hypertrophic chondrocytes exhibited similar changes in location and levels to HIF-2α and its downstream factor vascular endothelial growth factor (VEGF). In vitro, IHH-GLI-1-HIF-2α signalling activation in chondrocytes under physiological hypoxic conditions inhibited chondrocyte proliferation. In chondrocytes stimulated by inflammatory environments, IHH inhibited the degradation of HIF-2α via the GLI-1 pathway, thereby promoting HIF-2α protein expression. Elevated HIF-2α expression further enhanced intracellular IHH-GLI-1 levels, generating a positive feedback loop to collectively regulate the expression of downstream hypertrophic factors and matrix-degradation factors. In vivo, conditional Ihh knockout in mouse chondrocytes downregulated Hif-2α protein expression in early degenerative cartilage tissue and affected the expression of downstream Vegf and hypertrophic factors.ConclusionsDuring OA progression, the IHH-GLI-1-HIF-2α axis mainly operates within hypertrophic chondrocytes, exacerbating cartilage degeneration by regulating hypertrophic chondrocyte functions, cartilage matrix degradation, and microvascular invasion.The translational potential of this articleThis study identifies the IHH-GLI-1-HIF-2α signalling axis and reveals its potential as a therapeutic target for OA.

Dataset Information

Inhibition of syndecan-4 reduces cartilage degradation in murine models of osteoarthritis through the downregulation of HIF-2α by miR-96-5p.

Publications

Inhibition of syndecan-4 reduces cartilage degradation in murine models of osteoarthritis through the downregulation of HIF-2α by miR-96-5p.

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