Qichao Hu: State Key Laboratory of Southwestern Chinese Medicine Resources, School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, China; Department of Pharmacy, Chinese PLA General Hospital, Beijing, China. Electronic address: huqichaotcm@stu.cdutcm.edu.cn.
Yuan Chen: State Key Laboratory of Southwestern Chinese Medicine Resources, School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, China. Electronic address: tochenyuan@163.com.
Wenwen Zhang: State Key Laboratory of Southwestern Chinese Medicine Resources, School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, China. Electronic address: zhangwenwen0211@163.com.
Yubing Li: State Key Laboratory of Southwestern Chinese Medicine Resources, School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, China. Electronic address: liyubing940@163.com.
Xinyu Deng: State Key Laboratory of Southwestern Chinese Medicine Resources, School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, China. Electronic address: julydxytcm@163.com.
Lisheng Chen: State Key Laboratory of Southwestern Chinese Medicine Resources, School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, China; Department of Pharmacy, Chinese PLA General Hospital, Beijing, China. Electronic address: chenlsls27@163.com.
Zongyuan Zhou: Sichuan Industrial Institute of Antibiotics, School of Pharmacy, Chengdu University, China. Electronic address: zhouzongyuan@cdu.edu.cn.
Xiao Ma: State Key Laboratory of Southwestern Chinese Medicine Resources, School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu, China; Department of Pharmacy, Chinese PLA General Hospital, Beijing, China. Electronic address: tobymaxiao@163.com.
Jinhao Zeng: TCM Regulating Metabolic Diseases Key Laboratory of Sichuan Province, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, China; Department of Gastroenterology, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu , China. Electronic address: zengjinhao@cdutcm.edu.cn.
Yanling Zhao: Department of Pharmacy, Chinese PLA General Hospital, Beijing, China. Electronic address: zhaoyl2855@126.com.
BACKGROUND: Acute gastric injury, a common and recurring global digestive disorder, significantly impairs patient quality of life and overall health. Dehydroevodiamine (DHE), a bioactive natural product derived from Tetradium ruticarpum (A. Juss.) Hartley, shows potential therapeutic effects on acute gastric injury. This study investigates the underlying mechanisms of DHE's alleviating effects on acute gastric injury. METHODS: The gastric mucosal protective effect of DHE was confirmed through in vivo and in vitro acute gastric injury models. Biotin pulldown MS and molecular dynamics simulations identified DHE's target. CETSA and SPR assays validated DHE's affinity for IKK��. Protein site mutation validation and MST pinpointed the direct binding sites of DHE on IKK��. Additionally, the potential mechanism by which DHE ameliorates acute gastric injury was elucidated using WB, IHC, and IF methods, and further confirmed by rescue experiments. RESULTS: DHE effectively ameliorated IDO-induced gastric injury in GES-1 cells and rat gastric mucosa, both in vitro and in vivo. Biotin pulldown MS identified IKK�� as the target of DHE in alleviating gastric injury. CETSA and SPR assays confirmed DHE's direct binding to IKK��. Molecular dynamics simulations, protein mutation experiments, and MST results pinpointed GLU-149, GLU-49, and ASP-103 in the ATP-binding pocket as the binding sites of DHE on IKK��. Notably, DHE was found to competitively bind to IKK�� with ATP. Mechanistically, DHE attenuated IDO-induced gastric injury by inhibiting the IKK��-p65/NLRP3 signaling pathway. Importantly, exogenous activation of IKK�� reversed the therapeutic effect of DHE, indicating that DHE's efficacy depends on IKK��. CONCLUSION: DHE attenuated IDO-induced gastric injury by inhibiting the IKK��-p65/NLRP3 signaling pathway. Notably, DHE is a novel ATP-competitive IKK�� inhibitor that prevents phosphorylation by targeting GLU-149, GLU-49, and ASP-103 in the ATP-binding pocket. This study reveals new targets of action for DHE, providing a new molecular basis for using DHE in treating inflammation-related diseases.
