Yanghua Tang1, Yafeng Mo2, Dawei Xin1, Linru Zeng1, Zhenshuang Yue1, Canda Xu1. 1. Department of Orthopedics, Xiaoshan Hospital of Traditional Chinese Medicine No. 156 Yucai Road, Xiaoshan District, Hangzhou, Zhejiang Province, China. 2. Department of Orthopedics, Third Clinical College of Zhejiang Chinese Medical University No. 548 Binwen Road, Binjiang District, Hangzhou, Zhejiang Province, China.
Abstract
PURPOSE: To investigate the therapeutic effects of β-ecdysterone on osteoarthritis (OA) and the underlying mechanism. METHODS: OA model was established on rats by injecting MIA. ELSA was used to determine the concentration of IL-1β, IL-6, NO and TNF-α in the chondrocytes and cartilage tissues. Immunofluorescence assay was used to determine the expression of collagen II in the chondrocytes. The survival rate of chondrocytes was evaluated by MTT assay. The apoptosis of chondrocytes was checked by AO/PI staining and flow cytometry assay. The expression level of Atg7, PI3K and caspase-3 was evaluated by qRT-PCR. Western Blot was used determine the expression of PI3K, p-AKT1, AKT1, p-mTOR, mTOR, p70S6K, p-p70S6K, LC3I, LC3II and caspase-3. HE staining was used to check the pathological state of cartilage tissues. RESULTS: Chondrocytes were tolerable to rapamycin, 3-methyladenine and β-ecdysterone at the concentration of 10 mM, 100 nM and 40 μM, respectively. The apoptosis of chondrocytes was inhibited by rapamycin and β-ecdysterone, and induced by 3-methyladenine. PI3K, p-AKT1, p-mTOR, p-p70S6K and caspase-3 were down-regulated by rapamycin and β-ecdysterone, and up-regulated by 3-methyladenine in both the chondrocytes and the cartilage tissues. The expression of Atg7 and LC3II/LC3I were regulated in a opposite way. The inflammation state was improved by rapamycin and β-ecdysterone both the chondrocytes and the cartilage tissues. HE staining results showed that the pathological state of cartilage tissues was alleviated by β-ecdysterone. CONCLUSION: β-ecdysterone might alleviate osteoarthritis by activating autophagy in chondrocytes through regulating PI3K/AKT/mTOR signal pathway. AJTR
PURPOSE: To investigate the therapeutic effects of β-ecdysterone on osteoarthritis (OA) and the underlying mechanism. METHODS: OA model was established on rats by injecting MIA. ELSA was used to determine the concentration of IL-1β, IL-6, NO and TNF-α in the chondrocytes and cartilage tissues. Immunofluorescence assay was used to determine the expression of collagen II in the chondrocytes. The survival rate of chondrocytes was evaluated by MTT assay. The apoptosis of chondrocytes was checked by AO/PI staining and flow cytometry assay. The expression level of Atg7, PI3K and caspase-3 was evaluated by qRT-PCR. Western Blot was used determine the expression of PI3K, p-AKT1, AKT1, p-mTOR, mTOR, p70S6K, p-p70S6K, LC3I, LC3II and caspase-3. HE staining was used to check the pathological state of cartilage tissues. RESULTS: Chondrocytes were tolerable to rapamycin, 3-methyladenine and β-ecdysterone at the concentration of 10 mM, 100 nM and 40 μM, respectively. The apoptosis of chondrocytes was inhibited by rapamycin and β-ecdysterone, and induced by 3-methyladenine. PI3K, p-AKT1, p-mTOR, p-p70S6K and caspase-3 were down-regulated by rapamycin and β-ecdysterone, and up-regulated by 3-methyladenine in both the chondrocytes and the cartilage tissues. The expression of Atg7 and LC3II/LC3I were regulated in a opposite way. The inflammation state was improved by rapamycin and β-ecdysterone both the chondrocytes and the cartilage tissues. HE staining results showed that the pathological state of cartilage tissues was alleviated by β-ecdysterone. CONCLUSION: β-ecdysterone might alleviate osteoarthritis by activating autophagy in chondrocytes through regulating PI3K/AKT/mTOR signal pathway. AJTR