Paeoniflorin, the Main Monomer Component of Paeonia lactiflora, Exhibits Anti-inflammatory Properties in Osteoarthritis Synovial Inflammation.

OBJECTIVE: To explore the mechanism of paeoniflorin (PF) on osteoarthritis (OA) synovial inflammation from network pharmacology to experimental pharmacology. METHODS: Targets of OA were constructed by detecting the database of network database platforms (Therapeutic Target database, DrugBank and GeneCards), and the targets of PF were constructed by PubChem and Herbal Ingredients' Targets database. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of these co-targeted genes were conducted via Database for Annotation, Visualization, and Integrated Discovery (DAVID) database, and protein-protein interaction (PPI) networks were conducted via the search tool for the retrieval of interacting genes (STRING) database. Cell counting kit-8 (CCK-8) assay was performed to assess the potential toxicity of PF on human OA fibroblast-like synoviocytes (FLS), quantitative real-time polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA) and Western blot were used to verify the potential mechanism of PF in synovial inflammation. RESULTS: Twenty-six co-targeted genes were identified. GO enrichment results showed that these co-targeted genes were most likely localized in the cytoplasm, and the biological processes mainly involved 'cellular response to hypoxia' 'lipopolysaccharide (LPS)-mediated signaling pathway' and 'positive regulation of gene expression'. KEGG pathway analysis indicated that these co-targeted genes may function through pathways associated with 'hypoxia-inducible factor-1 (HIF-1) signaling pathway' and 'tumor-necrosis factor (TNF) signaling pathway'. The PPI network showed that the top 3 hub genes were TP53, TNF, and CASP3. Molecular docking results showed that PF was well docking with TNF. CCK-8 showed no potential toxicity of 10, 20 and 50 µmol/L PF on human OA FLS. And PF significantly decreased the expression levels of interleukin-1 ß, interleukin-6, TNF-α matrix metalloproteinase 13 (MMP13), and a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5) and TNF-α in LPS-induced OA FLS. CONCLUSION: PF exhibited potent anti-inflammatory effect in OA synovial inflammation.

Cinnamic Aldehyde, the main monomer component of Cinnamon, exhibits anti-inflammatory property in OA synovial fibroblasts via TLR4/MyD88 pathway.

Cinnamon is a wildly used traditional Chinese herbal medicine for osteoarthritis (OA) treatment, but the underlying mechanism remains ambiguous. The purpose of this study is to explore the mechanism of cinnamic aldehyde (CA), a bioactive substance extracted from Cinnamon, on synovial inflammation in OA. A total of 144 CA-OA co-targeted genes were identified by detect databases (PubChem, HIT, TCMSP, TTD, DrugBank and GeneCards). The results of GO enrichment analysis indicated that these co-targeted genes have participated in many biological processes including 'inflammatory response', 'cellular response to lipopolysaccharide', 'response to drug', 'immune response', 'lipopolysaccharide-mediated signalling pathway', etc. KEGG pathway analysis showed these co-targeted genes were mainly enriched in 'Toll-like receptor signalling pathway', 'TNF signalling pathway', 'NF-kappa B signalling pathway', etc. Molecular docking demonstrated that CA could successfully bind to TLR2 and TLR4. The results of in vitro experiments showed no potential toxicity of 10, 20 and 50 µM/L CA on human OA FLS, and CA can significantly inhibit the inflammation in LPS-induced human FLS. Further experimental mechanism evidence confirmed CA can inhibited the inflammation in LPS-induced human OA FLS via blocking the TLR4/MyD88 signalling pathway. Our results demonstrated that CA exhibited strong anti-inflammation effect in OA FLS through blocking the activation of TLR4/MyD88 signalling pathway, suggesting its potential as a hopeful candidate for the development of novel agents for the treatment of OA.

Efficacy and Mechanism of Electroacupuncture Treatment of Rabbits With Different Degrees of Knee Osteoarthritis: A Study Based on Synovial Innate Immune Response.

Knee osteoarthritis (KOA) is a chronic degenerative bone and joint disease, which is often clinically manifested as pain, joint swelling, and deformity. Its pathological manifestations are mainly synovial inflammation and cartilage degeneration. This study aims to investigate the efficacy of electro-acupuncture (EA) on model rabbits with varying degrees of KOA and to study the mechanism of EA on KOA based on the innate immune response. Mild and moderate rabbit KOA models were established using a modified Hluth method, and EA was given to both the mild and moderate model groups. The Lequesne-MG index was used to evaluate the behavioral changes in the rabbits before and after EA treatment. Morphological changes in the synovial membrane and cartilage of each group were observed by H&E staining. The Mankin scoring standard and the Krenn scoring standard were used to score the pathology of the cartilage tissue and synovial tissue, respectively. The inflammatory factors and metalloproteinases were detected in the serum of each group by ELISA. The protein and messenger RNA (mRNA) expressions of important elements related to Toll-like receptors (TLRs)-mediated innate immune response in the synovial tissue were detected by Western blot and quantitative PCR (qPCR). The Lequesne-MG index score of the rabbits gradually increased with the modeling prolonged but decreased significantly after EA treatment, indicating that EA has a better effect on alleviating the pain and improving the dysfunction. The morphological analysis showed that the inflammation of and the damage to the synovial membrane and the cartilage tissue gradually deteriorated with the modeling prolonged. However, the synovial membrane inflammation was significantly relieved after EA treatment, and the cartilage injury showed signs of repair. The ELISA analysis showed that, with the modeling prolonged, the serum-related inflammatory factors and mechanism of metalloproteinases gradually increased but decreased after EA treatment. The tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), and matrix metalloproteinase3 (MMP3) of EA1 group were significantly lower than those of EA2 group. Both Western blot and qPCR results showed that the protein and mRNA expressions of the elements related to the innate immune response in the synovial membrane increased gradually with the modeling prolonged, but decreased significantly after EA treatment. Additionally, the expression of some components in EA1 group was significantly lower than that in EA2 group. These results confirm that synovial inflammation gradually aggravated with time from the early to mid-stage of KOA. EA alleviated the inflammation and histological changes in KOA rabbits by inhibiting the TLRs-mediated innate synovial immune response. This suggests that using EA in the early stage of KOA may achieve a desirable efficacy.

Cinnamic Aldehyde Inhibits Lipopolysaccharide-Induced Chondrocyte Inflammation and Reduces Cartilage Degeneration by Blocking the Nuclear Factor-Kappa B Signaling Pathway.

Osteoarthritis (OA), as one of the top 10 causes of physical disability, is characterized by inflammation of the synovial membrane and progressive destruction of the articular cartilage. Cinnamic aldehyde (CA), an α,ß-unsaturated aldehyde extracted from the traditional Chinese herbal medicine cinnamon (Cinnamomum verum J.Presl), has been reported to have anti-inflammatory, antioxidant, and anticancer properties. However, the anti-inflammatory effect of CA on OA remains unclear. The purpose of the present study was to investigate the effects of CA on inflammation, and cartilage degeneration in OA. A CCK-8 assay was performed to assess the potential toxicity of CA on cultured human OA chondrocytes. Following treatment with lipopolysaccharide (LPS) and CA, the expression of proinflammatory cytokines, including interleukin (IL)-1ß, IL-6, and tumor necrosis factor-alfa (TNF-α), was evaluated using quantitative real-time polymerase chain reaction (RT-qPCR) analysis, enzyme-linked immunosorbent assay, and Western blotting (WB). The production of matrix metalloproteinase-13 (MMP-13) and a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS-5) was also examined using RT-qPCR and WB. Furthermore, to investigate the potential anti-inflammatory mechanism of CA, biomarkers of the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway (p65, IKB-α) were detected using WB. The results demonstrated that CA significantly inhibited the expressions of IL-1ß, IL-6, TNF-α, MMP-13, and ADAMTS-5 in LPS-induced OA chondrocytes. CA dramatically suppressed LPS-stimulated NF-κB activation. Collectively, these results suggest that CA treatment may effectively prevent OA.

[Effect of Tongluo Zhitong (, TLZT) gel preparation on p53 /miR-502-5p /NF-κBp65 in synovial tissue of knee osteoarthritis].

OBJECTIVE: To observe effects of Tongluo Zhitong (, TLZT) gel preparation on p53, miR-502-5p, NF-κBp65 in synovial tissue of knee osteoarthritis (KOA), and to explore mechanism of TLZT gel preparation in treating KOA. METHODS: Thirthy-six Wistar rats aged 8 weeks and weighed 200 to 220 g (meaned 208 g) were randomly divided into normal group, model group and traditional Chinese medicine (TCM) group, 12 rats in each group. KOA model was established by modified Hulth method. After 4 weeks of modeling, TCM group treated with TLZT gel preparation for external use, 3 times daily for 2 weeks;normal group and model group were fed normally without intervention. After treatment, morphological changes of specimens in each group were observed, changes of miR-502-5p in synovial tissue were detected by qPCR, and contents of p53, NF-κBp65, IL-1ß, TNF-α, MMP-13 in synovial tissue were detected by qPCR and Western Blot respectively. RESULTS: (1)Morphological observation of specimens showed that the articular cartilage in model group was hyaline and uneven, the synovial membranes were hypertrophic and proliferative with a large number of inflammatory cells infiltrating, the joint fluid was thicker in texture;the articular cartilage in TCM group was more transparent and smooth, synovial hyperplasia was mild with a small amount of inflammatory cell infiltration, the texture of articular fluid was clear and sparse. (2) Compared with normal group, content of miR-502-5p of synovial tissue in model and TCM group were increased, mRNA and expression of p53 decreased, expression of NF-κBp65, IL-1ß, TNF-α, MMP-13 increased. (3)Compared with model group, content of miR-502-5p in synovial tissue of TCM group decreased (P<0.05), mRNA and protein expression of p53 increased (P<0.05), mRNA and protein expression of NF-κBp65, IL-1ß, TNF-α, MMP-13 decreased (P<0.05). CONCLUSION: Expression of p53, miR-502 -5p, NF -κBp65 in synovial tissue is closely related to synovial hyperplasia and inflammatory reaction, TLZT gel preparation may reduce proliferation and inflammatory reaction of KOA synovium by regulating the expression of p53, miR- 502-5p, NF-κBp65 in synovial tissues.