Fig 1: mTOR inhibitors could decrease the tumor stroma and proportion of TipECs. A, Flowchart for screening of commonly used clinical drugs that reduced tumor stroma. Chemo, chemotherapy; SSA, somatostatin analogue; TKI, tyrosine kinase inhibitor. B, The changes in tumor stroma, ApoE, CD31, and PDGFB induced by commonly used clinical drugs could be demonstrated through H&E staining and IHC. Scale bar, 50 μm. C, The ssGSEA analysis was utilized to demonstrate the effects of various drugs on tumor stroma based on transcriptome data. EMT, epithelial–mesenchymal transition; TMZ, temozolomide. D, The impact of different drugs on the tumor stroma score could be assessed based on transcriptome data. E, The gene ontology enrichment analysis of downregulated genes in the everolimus treatment group relative to the control group. F, Experimental design for G and H. Everolimus or the same volume of PBS was administered by intragastric administration in PDX (n = 5 per group). NSG, NOD/SCID gamma. G, The secretion comparison of ApoE and PDGFB between the everolimus-treated group and the PBS-treated group in IF. H, The proportion of CD45−CD31+CXCR4+ cells between the everolimus-treated group and the PBS-treated group. Data in the graph are shown as the means ± SD from three independent experiments. **, P < 0.01; ***, P < 0.001.
Fig 2: The mechanisms by which the TipECs recruited CAFs. A, The SCENIC algorithm was used to predict specific transcription factor activity within various subtypes of ECs. AEC, artery EC; CapEC, capillary EC; ProlifEC, proliferating EC; StrEC, stromal EC. B, Rank for regulons in TipECs based on regulon specificity score according to pySCENIC analysis. C, Transcriptomic analysis demonstrates the impact of ApoE (10 ng/mL) on the predicted transcriptional factors in A and B. D, The effect of representative acid palmitic acid on the changes in predicted transcription factors (20 µmol/L). E, Based on GSEA transcription factor dataset analysis, the normalized enrichment score (NES) of ATF6 transcription factor activated by ApoE is shown. F, The impact of ApoE (10 ng/mL) on the activity of transcription factors at different time points. G and H, Deletion of ATF6 and its effect on the expression of PDGFB in HUVECs influenced by ApoE (10 ng/mL) and palmitic acid (20 µmol/L). KO, knockout; NC, negative control. I, Possible binding site of ATF6 as a transcription factor with the PDGFB promoter. J, ChIP of ATF6 in HUVECs. ATF6 binding to the PDGFB promoter region was quantified by PCR. K and L, The binding of ATF6 to the PDGFB promoter region was enhanced by ApoE (10 ng/mL) and palmitic acid (20 µmol/L). Data in the graph are shown as the means ± SD from three independent experiments. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, not significant.
Fig 3: ApoE mediated EC lipid metabolism to affect the recruitment of fibroblasts. A, Interaction strength coefficients between TipECs and various types of cells in the tumor microenvironment. Mph, macrophage. B, Comparisons of the activity of tumor stroma–related signaling pathways in various cell types. C, The interaction strengths between sender of PDGF and NOTCH signaling and corresponding receiver were compared in various cell types. D, Transcriptomic analysis demonstrated the impact of ApoE (10 ng/mL) on the alteration of genes associated with the PDGF signaling pathway in HUVECs. E, Different concentrations of ApoE resulted in changes in the transcriptional expression of PDGFB. F, Different periods of ApoE (10 ng/mL) resulted in changes in the secretion of PDGFB. G, Free fatty acids metabolomics revealed the impact of different concentrations of ApoE on the levels of various fatty acids. H and I, PCR and ELISA assays demonstrated the effects of different free fatty acids on the expression and secretion of PDGFB in HUVECs. The concentration of 27 free fatty acids administered was 20 µmol/L. J, Experimental design for K and L. K and L, The impact of ApoE (10 ng/mL) and palmitic acid (20 µmol/L) conditioning on the invasive ability of HUVECs toward CAFs. Scale bar, 100 μm. Data in the graph are shown as the means ± SD from three independent experiments. ***, P < 0.001; ns, not significant. AEC, artery EC; CapEC, capillary EC; ProlifEC, proliferating EC; StrEC, stromal EC.
Supplier Page from Abcam for Human PDGF B ELISA Kit, Fluorescent