HUBEI AGRICULTURAL SCIENCES ›› 2023, Vol. 62 ›› Issue (9): 175-180.doi: 10.14088/j.cnki.issn0439-8114.2023.09.031

• Biological Engineering • Previous Articles     Next Articles

Protective effect of ampelopsin on injury of A549 cells based on AMPK/mTORC1 autophagy pathway

ZHU Hai-bin, ZENG Chun-hui, TANG Mu-lan, CHI Xin-yu, DENG Hao-jian, LI Yu-jun, FANG Jing, XU Hao-liang, YANG Ke   

  1. College of Pharmacy/Key Laboratory of Traditional Chinese Medicine Pharmacology of Guangxi University, Guangxi University of Chinese Medicine, Nanning 530200, Guangxi, China
  • Received:2022-05-23 Online:2023-09-25 Published:2023-10-24

Abstract: To investigate the protective effect of ampelopsin (APS) on lipopolysaccharide (LPS)-induced A549 cell injury model and its impact on the AMPK/mTORC1 autophagy pathway, the cells were randomly divided into the following groups: normal group, model group, APS high concentration group, APS medium concentration group, and APS low concentration group (80, 40, 20 μg/mL). Except for the normal group, all other groups were treated with 30 μg/mL LPS to induce A549 cell injury model for 8 hours. After 24 hours of APS intervention, the survival rate of A549 cells was measured using the MTT method. The leakage of LDH in the supernatant was detected using the fluorescence method, and the TNF-α content in the supernatant was detected using the Elisa method. Autophagosome formation in the cells was assessed using the MDC method, while cell autophagy flow was measured using autophagy double-labeled mRFP-GFP-LC3. The p-AMPK/AMPK, p-mTOR/mTOR, autophagy-related protein LC3-II/LC3-I, and P62 protein expression levels were detected using the Western blot method. Additionally, an intervention validation trial using an AMPK inhibitor (CC) was conducted. The trial involved randomly dividing the subjects into different groups, including the normal group, model group, LPS+APS group, LPS+CC group, and LPS+APS+CC group. After co-culturing APS and CC for 24 hours, Western blot analysis was performed to measure the expression levels of p-AMPK/AMPK, p-mTOR/mTOR, and the autophagy-related protein LC3-Ⅱ/LC3-Ⅰ, as well as P62 protein.The results indicated that APS significantly increased the survival rate of A549 cells damaged by LPS, while reducing LDH leakage and TNF-α content. Moreover, APS led to a significant increase in the number of autophagosomes and lysosomes in the cells, along with a downregulation of P62 protein and p-mTOR/mTOR expression. Additionally, the expression of LC3-Ⅱ/LC3-Ⅰ was significantly upregulated. Furthermore, when the AMPK/mTORC1 autophagy pathway was inhibited using CC, APS was found to upregulate the expression of P62 protein, p-AMPK/AMPK and LC3-Ⅱ/LC3-Ⅰ proteins, while downregulating p-mTOR/mTOR protein expression. These findings suggested that APS promoted autophagy by activating the AMPK/mTORC1 autophagy pathway, thereby enhancing autophagy flow and exerting a protective effect against LPS-induced damage to A549 cells.

Key words: ampelopsin(APS), lipopolysaccharide(LPS), A549 cells, autophagy, AMPK/mTORC1 signaling pathway

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