Multi-immunometabolomics mining: NP prevents hyperimmune in ALI by inhibiting Leucine/PI3K/Akt/mTOR signaling pathway

Acute lung injury (ALI) is currently a global health concern. Nicandra physalodes (L.) Gaertn. (NP) holds an important position in traditional Chinese medicine and nutrition. The potential protective mechanisms of NP against ALI remain unknown. The purpose of this study was to investigate the protective effects and molecular mechanisms of NP extract (NPE) on lipopolysaccharide (LPS)-induced ALI in mice. By utilizing network pharmacology to forecast the active ingredients in NP as well as possible signaling pathways. The composition of the NPE was analyzed using UPLC-Q-TOF-MS/MS. In addition, ¹H-NMR immunometabolomics was employed to identify alterations in primary metabolic pathways and metabolites in the lung, serum, and fecal tissues. Finally, the protein and gene expression of key pathways were verified by IHC, IF, RT-qPCR, and ELISA. It was found that the main ingredients of NPE were revealed to be nicandrenone, withanolide A, and baicalin. NPE significantly improved lung injury, pulmonary edema, and inflammatory cell infiltration in mice with ALI. In addition, NPE improved autophagic activity and alleviated Th1 and Th17 cell-induced lung inflammation by suppressing the PI3K/Akt/mTOR signaling pathway. Importantly, immunometabolomic analysis of fecal, serum, and lung tissues revealed that NPE reversed ALI-induced leucine resistance by remodeling immunometabolism. We confirmed NPE prevents ALI by remodeling immunometabolism, regulating the Leucine/PI3K/Akt/mTOR signaling pathway, inhibiting Th1/Th17 cell differentiation, and providing a scientific immunological basis for the clinical application of NPE.