Fig 2: The angiogenic effect of HSYA was related to PI3K/Akt/mTOR signaling pathway. Representative western blots of VEGF-A (A), CD34 (B), and HIF-1α (C), phospho-Akt and total Akt (D), phospho-p70S6k and total p70S6k (E), phospho-mTOR and total mTOR along with their quantification in sham, CCI, 10 mg/kg HSYA, and 20 mg/kg on day 14. n = 4, *p < 0.05, **p < 0.01, ***p < 0.001

Fig 3: HSYA promoted angiogenesis through activating PI3K/Akt/mTOR signaling pathway in TBI. Western blotting showing the protein expression of A phospho-Akt and total Akt, B phosphor-p70S6k and total p70S6k, C phospho-mTOR and total mTOR, D VEGF-A, E CD34, and F HIF-1α in CCI, 20 HSYA, and 20 HSYA + LY294002. n = 4. Molecular docking of HSYA with PI3k in 3D (G) and 2D (H). *p < 0.05, **p < 0.01

Fig 4: IL‐33 upregulates the expression of related adhesion molecules in ECs and EGC. (A) KEGG bubble map of DEGs between IL‐33‐knockdown ECs and negative control ECs. (B) RT‐qPCR was used to verify the DEGs in ECs after knockdown or overexpression of IL‐33. (C) WB was used to verify the gene expression of ECs after knockdown or overexpression of IL‐33. (D) ‌Tissue immunofluorescence showed that the expressions of IL‐33 (p = 0.045) and CD34 (p = 0.004) were higher in submucosal carcinoma than in intramucosal carcinoma. EGC with high expression of IL‐33 (p = 0.048) and CD34 (p = 0.011) was more malignant. (E) ‌DEG volcano plot of EGC organoids after the addition of IL‐33. (F) ‌GSEA revealed that DEGs were enriched in the cell adhesion and MAPK signaling pathways after ECs overexpressed IL‐33. (G) RT‒qPCR was used to verify the DEGs of EGC organoids after IL‐33 overexpression. (H) WB was used to verify the gene expression of organoids after IL‐33 overexpression. (I) Proposed a model of the mechanism in this study. IL‐33 activates the IL‐3/ST2 signaling pathway in an autocrine manner, enhances the expression of PECAM1 and CD34 in ECs, and promotes angiogenesis. High levels of IL‐33 also bind to ST2 in EGC organoids in a paracrine manner and activate the MAPK signaling pathway after the expression of KRT17 is upregulated, promoting the growth of EGC. IHC of IL‐33 at the boundary of normal gastric mucosa and EGC. Scale bar, 200 µm. The data are presented as the means ± SDs of three independent experiments. *p < 0.05; ****p < 0.0001 versus empty vector control (two‐sided Wilcoxon rank‐sum test). DEGs, differentially expressed genes; ECs, endothelial cells; EGC, early gastric cancer; GSEA, gene set enrichment analysis; KEGG, Kyoto Encyclopedia of Genes and Genomes; MAPK, mitogen‐activated protein kinase; NC, negative control; OE‒IL‒33, overexpressed IL‒33; RT‐qPCR, real‐time quantitative polymerase chain reaction; sh‒IL‒33, knocked down IL‒33; WB, western blot.