Fig 2: Anti-VEGF neutralizing antibody inhibits the expression of endothelial markers in differentiating EPCs in vitro. A VEGF neutralizing antibody (100 ng/ml) was added to MSCCM. MSCCM with mouse IgG (100 ng/ml) was used as the control. EPCs were cultured in LG-DMEM, MSCCM, MSCCM with IgG and MSCCM with anti-VEGF antibody for 48 h. (A-D) Immunofluorescence staining demonstrating CD31 expression in the (A) LG-DMEM, (B) MSCCM, (C) MSCCM with IgG and (D) MSCCM with anti-VEGF groups. (E-H) Immunofluorescence staining demonstrating vWF expression in the (E) LG-DMEM, (F) MSCCM, (G) MSCCM with IgG and (H) MSCCM with anti-VEGF groups. Scale bar, 50 µm. (I and J) Reverse transcription-quantitative polymerase chain reaction was used to assess the expression of CD31 and vWF mRNA in the LG-DMEM, MSCCM, MSCCM with IgG and MSCCM with anti-VEGF groups. (K and L) Western blotting demonstrated the protein expression levels of (K) CD31 and (L) vWF in the LG-DMEM, MSCCM, MSCCM with IgG and MSCCM with anti-VEGF groups. Data are expressed as the mean ± standard error of the mean. *P<0.05, **P<0.01, as indicated. VEGF, vascular endothelial growth factor; EPC, endothelial progenitor cell; MSC, mesenchymal stem cell; MSCCM, MSC conditioned media; Ig, immunoglobulin; LG-DMEM, low-glucose Dulbecco's modified Eagle's medium; CD, cluster of differentiation; vWF, von Willebrand factor; n.s., not significant; IOD, integrated optical density.

Fig 3: ?3 PUFAs via FFAR4 suppress neointimal hyperplasia. A, Representative H&E-stained images of control CCA without FeCl3 injury. B, Representative image of CCA at 28 days after FeCl3 injury. Bar indicates 100 µm. C, Representative images show neointima, using immunofluorescence labeling of a-smooth muscle actin (green) and nuclei (blue) in CCA of 4 strains of mice at 28 days after FeCl3 injury. D, Immunostained CD31 (green) as a marker for endothelial cells. E, H&E staining. Bars indicate 100 µm. F, Calculated I/M ratio. *P<0.05 and **P<0.01 determined by 1-way ANOVA post hoc analysis. Values are given as mean±SEM, n=6 to 7 as indicated in parentheses. CCA indicates common carotid artery; E, endothelial cells; EEL, external elastic lamina; H&E, hematoxylin and eosin; IEL, internal elastic lamina; I/M, intima:media; M, media; N, neointima; PUFAs, polyunsaturated fatty acids; PVAT, perivascular adipose tissue; WT, wild type.

Fig 4: CD105pos and CD105neg fibroblasts are identified in normal and tumor-bearing tissues(A) MC analysis of in-vitro-expanded primary fibroblasts. Plots show PDPN and CD105 levels. LIN, EpCAM CD31 CD45.(B) UMAP projection of CAFs from KPC pancreatic (n = 4), KPN colorectal (n = 5), MMTV-PyMT mammary (n = 4), KP lung (n = 4), and BRAFV600E melanoma (n = 3) GEMMs. FlowSOM clusters are color coded. Total of 5 × 105 cell displayed.(C) Stacked bar graphs of GEMM CAF (GCAF) clusters displayed as a fraction of total CAFs. FlowSOM colors based on (B).(D) Heatmap of marker median mass intensities (MMIs) displayed as Z scores. Each GCAF FlowSOM cluster is grouped by unsupervised hierarchical clustering based on marker MMIs. Cell-type annotations based on canonical phenotypic markers are listed. Tumor type/s that the GCAF clusters predominantly arise from are listed.(E) UMAP projection from (B) displaying overlaid signal intensity of CD105 with annotated tumor types.(F) UMAP projection from (B) displaying overlaid signal intensity of example markers. The tumor types of origin are highlighted: Pa, pancreatic; Co, colorectal; Ma, mammary; Lu, lung; Me, melanoma.(G–I) Representative IHC analysis of human colorectal (n = 9), breast (n = 8), and lung adenocarcinoma (n = 6) tumor samples stained for pan-cytokeratin (PCK) (green), vimentin (VIM) (purple), CD105 (yellow), and DAPI (blue). Scale bar, 500 µm.See also Figure S8 and Table S7.