Dihydromikanolide Inhibits ROS-Mediated NLRP3 Inflammation via Antioxidant Nrf2 Activation and Mitophagy Induction in LPS/ATP-Stimulated Macrophages.

Dihydromikanolide (DHK) is a natural product in Mikania species. We examined the anti-inflammatory molecular mechanisms of DHK employing in vitro RAW264.7 macrophages and in vivo BALB/c mice under LPS/ATP stimulation. We found that DHK suppressed NLRP3 inflammasome, procaspase-1 activation and then pro-inflammatory IL1β expression in LPS/ATP-stimulated RAW264.7 cells. Notably, DHK-triggered autophagy in RAW264.7 cells was demonstrated by increased LC3-II accumulation, p62/SQSTM1 expression, Beclin-1/Bcl-2 ratio and PI3K/AKT/mTOR phosphorylation. Besides, DHK increased Parkin and Pink-1 protein expressions implying mitophagy induction in RAW264.7 cells. Interestingly, DHK enhanced Nrf2 nuclear translocation and provoked antioxidant HO-1, NQO-1 and γ-GCLC expressions in RAW264.7 cells. Nrf2 knockdown reversed DHK-inhibited LPS/ATP-stimulated IL1β expression in RAW264.7 cells. Interestingly, LPS/ATP-stimulated NLRP3 inflammasome and IL1β expression were inhibited by DHK, Mito-TEMPO (a mitochondrial ROS inhibitor), or N-acetylcysteine (a ROS inhibitor). In vivo study revealed that DHK attenuated wet/dry weight ratio of lung tissue, lung neutrophil intrusions and pulmonary oedema, and reduced the increased total cells, neutrophils, TNFα and IL1β expression in bronchoalveolar lavage fluid (BALF) in LPS-stimulated BALB/c mice. DHK alleviated LPS-induced pathological alterations of lung through inhibiting NLRP3 inflammation, enhancing antioxidant Nrf2 pathway and inducing mitophagy in LPS-stimulated BALB/c mice. Dihydromikanolide may be a potential therapeutic agent for inflammatory diseases.