Escin AInhibits the Progression of Osteoarthritis by Down-regulating the HIF-2α, NF-κB Pathways and Their Antioxidant Activity
WANG Miao,WAN Rui-jie,LIU Wei
Abstract:
Objective To explore the role and related molecular mechanism of Escin A (Escin A, EsA) in mouse osteoarthritis (OA). Methods Isolate and culture the primary chondrocytes of C57BL/6 mice, and treat the cells with 10 mg/L IL-1β and different concentrations (10, 50, 100 μmol/L) of Escin A. CCK-8 detects cell activity.The Enzyme-linked immunosorbent assay was used to detect the content of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in the supernatant of cell culture fluid,real-time quantitative PCR to detect the mRNA expression levels of matrix metalloproteinase-13 (MMP-13) and proteoglycanase (ADAMTS5), western blot detection of collagen type II (Collagen II), matrix metalloproteinase-9 (MMP-9), hypoxia-inducible factor-2 (HIF-2α) and nuclear transcription factor-Kappa B (NF-κB) pathway related protein expression, immunofluorescence staining to detect p65 expression, superoxide dismutase (SOD), catalase (CAT) and malondialdehyde (MDA) kits were used to detect the level of oxidative activity. The 60 C 57 BL/ 6 male mice were randomly divided into sham operation group, OA group, OA+low-dose EsA group and OA+high-dose EsA group, which 15 mice in each group.After establishing the mouse OA model, mice in the OA+low-dose EsA group and OA+high-dose EsA group were injected with EsA intraperitoneally at 5 mg/kg and 10 mg/kg respectively, every other day, and the mice were sacri?ced after 6 weeks. Bone density measuring instrument detects the bone density of the proximal femur in mice,HE staining and safranine O fast green staining were used to detect the pathological morphological changes of bone tissue. Results Under the action of Escin A, it can inhibit the decrease of cell survival rate and the increase of TNF-α and IL-6 content caused by IL-1β stimulation (P<0.05). At the same time, it can make MMP-13 and ADAMTS-5 mRNA expression decreased (P<0.05), Collagen II protein expression increased and MMP-9 protein expression decreased (P<0.05), inhibited IL-1β-induced activation of HIF-2α and NF-κB signaling pathways, and make MDA level drop, SOD and CAT activity increase (P<0.05).After OA mice were treated with Escin A at low and high doses, the bone density of the femur of the mice increased (P<0.05), the bone tissue lesions were improved. Conclusion Escin A can inhibit osteoarthritis in mice, and its mechanism may be related to the HIF-2α and NF-κB pathways and their antioxidant activity.
