N-Butanol Extract of Gastrodia elata Suppresses Inflammatory Responses in Lipopolysaccharide-Stimulated Macrophages and Complete Freund's Adjuvant- (CFA-) Induced Arthritis Rats via Inhibition of MAPK Signaling Pathway.

Gastrodia elata is a traditional herbal medicine that has been used for centuries to treat rheumatism. Previous studies have confirmed that ethanol extracts of Gastrodia elata have anti-inflammatory and antioxidant activities, and the n-butanol fraction exerts a higher inhibitory effect. However, the in vivo anti-inflammatory effects of Gastrodia elata have not been evaluated. Thus, we assessed the therapeutic effect of the n-butanol extract of Gastrodia elata (BGE) on complete Freund's adjuvant- (CFA-) induced arthritis rats which were separated into six groups (NOR; MODEL; CFA + dexamethasone (DEX); CFA + 25, 50, 100 mg/kg BGE). The paw swelling, joint radiology, and histology were used to analyze the effect of BGE on delaying the progression of rheumatoid arthritis. Furthermore, serum levels of inflammatory cytokines were analyzed via ELISA. In addition, the effect of BGE on nitric oxide (NO) production, expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2(COX-2), and inflammatory cytokines were detected in lipopolysaccharide- (LPS-) stimulated RAW264.7 macrophage cells. Lastly, the impacts of BGE on the activation of the mitogen-activated protein kinases (MAPK) pathway in CFA rats and LPS-stimulated RAW264.7 macrophage were examined by western blot analysis. The results show that BGE can significantly reduce paw swelling without losing the body weight of rats. Imaging assessment confirms that BGE can protect cartilage from destruction, as well as reducing inflammatory cell infiltration and synovial proliferation. Moreover, BGE suppresses the production of inflammatory cytokines in serum and inhibits the activation of the phosphorylation of p38 and ERK in CFA rats. BGE was also demonstrated to decrease the production of NO and inflammatory cytokines in LPS-stimulated RAW264.7 cells. The effect of BGE in LPS-induced expression leads to reduced p38 and ERK phosphorylation and also downregulates the protein expression of iNOS and COX-2. Taken together, BGE exhibits a potential therapeutic effect on CFA rats, and its anti-inflammatory and antioxidant effects were possibly exerted by regulation of ERK/p38MAPK.

microRNA­96 regulates the proliferation of nucleus pulposus cells by targeting ARID2/AKT signaling.

The aberrant proliferation of nucleus pulposus (NP) cells has been reported to be implicated in the pathogenesis of intervertebral disc degeneration (IDD). Previous studies have demonstrated that microRNAs (miRNAs), which are a group of small noncoding RNAs, are critical regulators of cell proliferation in various pathologies. However, the role of miRNA­96 (miR­96) in the proliferation of NP cells remains to be determined. In the present study, reverse transcription­quantitative polymerase chain reaction was used to investigate the expression of miR­96 in NP tissues from patients with IDD and healthy tissues from patients with traumatic lumbar fracture as the control. A dual­luciferase reporter assay was used to investigate whether AT­rich interaction domain 2 (ARID2) may be a direct target gene for miR­96. Furthermore, isolated NP cells from patients with IDD were transfected with miR­96 mimics and ARID2­targeting small interfering RNAs; cell proliferation, and the protein expression of Akt, phosphorylated Akt and ARID2 were examined, whereas the effects of an Akt inhibitor on NP cell proliferation were also evaluated. The present results demonstrated that miR­96 expression was significantly upregulated in IDD samples, and the level of miR­96 expression was positively associated with disc degeneration grade, which was evaluated by a modified Pfirrmann grading system. In addition, the current study identified ARID2 as a direct gene target of miR­96. Furthermore, it was demonstrated that ARID2 mRNA expression was inversely correlated with the expression of miR­96 in NP tissues. In addition, miR­96 overexpression promoted NP cell proliferation and induced Akt phosphorylation, which led to increased cyclin D1 translation. Notably, overexpression of ARID2 or treatment with an Akt inhibitor decreased the effect of miR­96 on NP cell proliferation. In conclusion, the results of the present study indicate that miR­96 may promote the proliferation of human degenerated NP cells by targeting ARID2 via activation of the Akt pathway, and potentially serves as a therapeutic target for IDD.