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Targeting Extracellular miR-21-TLR7 Signaling Provides Long-Lasting Analgesia in Osteoarthritis.

ABSTRACT: Osteoarthritis (OA) is the most prevalent joint disorder associated with severe chronic pain. Although synovial inflammation is well correlated with pain severity, the molecular mechanism responsible for OA pain remains unclear. Here, we show that extracellular miR-21 released from synovial tissue mediates knee OA pain in surgical OA model rats. miR-21 was the most abundant among increased microRNAs (miRNAs) in the synovial tissue. miR-21 was released into extracellular space from the synovial tissue and increased in the synovial fluid. A single intra-articular injection of miR-21 inhibitor exerted long-term analgesia of knee OA pain, whereas miR-21 injection in naive rats caused knee joint pain. miR-21 mutant, which lacks the Toll-like receptor (TLR) binding motif, but not in the seed sequence, did not cause joint pain, suggesting a non-canonical mode of action different from translational repression. Consistent with this, the algesic effect of miR-21 was blocked by antagonizing TLR7. The TLR7 antagonist also exerted a long-lasting analgesic effect on knee OA pain. Therefore, extracellular miR-21 released from synovial tissue mediates knee OA pain through TLR7 activation in surgical OA model rats. Extracellular miRNA in the joint may be a plausible target for pain therapy, providing a novel analgesic strategy for OA.

SUBMITTER: Hoshikawa N

PROVIDER: S-EPMC6920297 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Targeting Extracellular miR-21-TLR7 Signaling Provides Long-Lasting Analgesia in Osteoarthritis.

Hoshikawa Naoya N Sakai Atsushi A Takai Shinro S Suzuki Hidenori H

Molecular therapy. Nucleic acids 20191120

Osteoarthritis (OA) is the most prevalent joint disorder associated with severe chronic pain. Although synovial inflammation is well correlated with pain severity, the molecular mechanism responsible for OA pain remains unclear. Here, we show that extracellular miR-21 released from synovial tissue mediates knee OA pain in surgical OA model rats. miR-21 was the most abundant among increased microRNAs (miRNAs) in the synovial tissue. miR-21 was released into extracellular space from the synovial t ...[more]

PMID: 31841992

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Project description:BackgroundDue to the lack of research on the pathological mechanism of temporomandibular joint osteoarthritis (TMJOA), there are few effective treatment measures in the clinic. In recent years, microRNAs (miRs) have been demonstrated to play an important role in the pathogenesis of osteoarthritis (OA) by regulating a variety of target genes, and the latest evidence shows that miR-21-5p is specifically overexpressed in OA. The purpose of this project was to clarify whether miR-21-5p can regulate the TMJOA process by targeting Spry1.MethodsTMJOA was induced by a unilateral anterior crossbite (UAC) model, and the effect of miR-21-5p knockout on TMJOA was evaluated by toluidine blue (TB), immunohistochemical (IHC) staining, Western blotting (WB) and RT-qPCR. Primary mouse condylar chondrocytes (MCCs) were isolated, cultured and transfected with a series of mimics, inhibitors, siRNA-Spry1 or cDNA Spry1. WB, RT-qPCR, IHC and TB were used to detect the effect of miR-21-5p and its target gene Spry1 on the expression of MMP-13, VEGF and p-ERK1/2 in TMJOA. The effect of miR-21-5p on angiogenesis was evaluated by chick embryo chorioallantoic membrane (CAM) assay and WB.ResultsIn the UAC model, the cartilage thickness and extracellular matrix of miR-21-5p knockout mice were less damaged, and miR-21-5p and UAC model were shown to affect the expression of Spry1, IL-1β, MMP-13, and VEGF. Luciferase experiments confirmed that Spry1 was the direct target of miR-21-5p. The expression levels of Spry1, MMP-13, VEGF and p-ERK1/2 in MCCs transfected with miR-21-5p mimic were higher than those in the inhibitor group. Under the simulated inflammatory environment of IL-1β, the expression levels of MMP-13, VEGF and p-ERK1/2 were positively correlated with miR-21-5p, while Spry1 was negatively correlated with miR-21-5p. Inhibition of miR-21-5p expression and overexpression of Spry1 enhanced the inhibition of MMP-13, VEGF and p-ERK1/2 expression. MiR-21-5p had a significant role in promoting angiogenesis in the chick embryo CAM assay, and this role was clearly mediated by the ERK-MAPK signalling pathway.ConclusionThis study verified that miR-21-5p can promote the process of TMJOA by targeting Spry1, which provides a new direction for future research on the treatment of this disease.

Project description:BackgroundOur previous study identified miR-99a as a negative regulator of early chondrogenic differentiation. However, the functional role of miR-99a in the pathogenesis of osteoarthritis (OA) remains unclear.MethodsWe examined the levels of miR-99a and Frizzled 8 (FZD8) expression in tissue specimens. Human SW1353 chondrosarcoma cells were stimulated with IL-6 and TNF-α to construct an in vitro OA environment. A luciferase reporter assay was performed to analyze the relationship between miR-99a and FZD8. CCK-8 assays, flow cytometry, and ELISA assays were used to assess cell viability, apoptosis, and inflammatory molecule expression, respectively. Percutaneous intra-spinal injections of papain mixed solution were performed to create an OA Sprague-Dawley rat model. Alcian Blue staining, Safranin O Fast Green staining, and Toluidine Blue O staining were performed to detect the degrees of cartilage injury.ResultsMiR-99a expression was downregulated in the severe spine OA patients when compared with the mild spine OA patients, and was also decreased in the experimentally induced in vitro OA environment when compared with the control environment. Functionally, overexpression of miR-99a significantly suppressed cell apoptosis and extracellular matrix degradation stimulated by IL-6 and TNF-α. FZD8 was identified as a target gene of miR-99a. Furthermore, the suppressive effects of miR-99a on cell injury induced by IL-6 and TNF-α were reversed by FZD8 overexpression. Moreover, the levels of miR-99a expression were also reduced in the induced OA model rats, and miR-99a agomir injection relieved the cartilage damage. At the molecular level, miR-99a overexpression downregulated the levels of MMP13, β-catenin, Bax, and caspase-3 protein expression and upregulated the levels of COL2A1 and Bcl-2 protein expression in the in vitro OA-like chondrocyte model and also in the experimental OA model rats.ConclusionsOur data showed that miR-99a alleviated apoptosis and extracellular matrix degradation by targeting FZD8, and thereby suppressed the development and progression of experimentally induced spine osteoarthritis.

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Targeting Extracellular miR-21-TLR7 Signaling Provides Long-Lasting Analgesia in Osteoarthritis.

Publications

Targeting Extracellular miR-21-TLR7 Signaling Provides Long-Lasting Analgesia in Osteoarthritis.

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