Abstract

Lymphangiogenesis drives gastric cancer lymph node (GC LN) metastasis via neuro-immune-cancer interactions in the tumor microenvironment. However, the underlying mechanisms remain incompletely understood. Here, we demonstrate that chronic stress (CS) in GC patients correlates with poorer short-term prognosis, perineural/lymphovascular invasion, and advanced LN staging. Mechanistically, CS-induced neutrophil extracellular traps (NETs) are key mediators of LN metastasis. Inhibiting NETs suppressed popliteal LN metastasis and lymphangiogenesis. In vitro and in vivo experiments revealed that CS upregulated acetylcholine (ACh) and choline acetyltransferase (ChAT) levels, with ACh directly inducing NETs formation in a dose-dependently. Exogenous ACh increased LN metastasis (volume), whereas gastric vagotomy attenuated CS-mediated orthotopic tumor growth. Functional assays confirmed that ACh-driven NETs enhanced GC cell malignant behaviors, accelerated adhesive suspension state transformation via extracellular matrix remodeling, and activated a feedforward ACh-NGF axis. This triggered PKC-γ/STAT3-S100A8/A9-ROS cascade, mediating NETs formation and upregulating VEGFA/C. Protein analysis showed CD300ld overexpression in CS + GC tissues, independent of ACh signaling. Notably, combined targeting of the NGF/TrkA and CD300ld exerted synergistic anti-LN metastatic and lymphangiogenic effects. Collectively, this study elucidates a CS-driven mechanism regulating NETs to promote GC LN metastasis and identifies NGF/TrkA and CD300ld as therapeutic targets for GC LN metastasis.