Journal: Translational Oncology

Article Title: Spatial and single-cell multi-omics reveal pro-angiogenic THY1 ⁺ fibroblast subtypes predicting prognosis in prostate cancer

doi: 10.1016/j.tranon.2026.102664

Figure Lengend Snippet: Single-cell and immunohistochemical validation of CAF subpopulations. (A) t-SNE visualization of major cell types in the GSE176031 single-cell RNA-seq dataset ( n = 21,743 cells). (B) Heatmap showing the mean expression levels of the 8 signature genes across 9 major cell types. (C) UMAP projection of 948 CAFs from our single-cell RNA-seq dataset2 ( GSE141445 ), colored according to the expression levels of ACTA2, VIM, COL1A1, and THY1. (D–G) Multiplex immunofluorescence staining of a prostate cancer TMA (84 tumor samples; four markers: DAPI [blue], COL1A1 [green], THY1 [yellow], α-SMA [red]): (D) Representative image from a BCR-positive, GS 4 + 3 sample with 46.9 % THY1 ⁺ CAFs (scale bar: 300 μm); (E) Representative image from a BCR-negative, GS 4 + 3 sample with 7.7 % THY1 ⁺ CAFs (scale bar: 300 μm); (F) enlarged views from 5 BCR-positive cases (scale bar: 100 μm); (G) enlarged views from 5 BCR-negative cases (scale bar: 100 μm). (H) Violin plot comparing the percentage of THY1 ⁺ CAFs between the GS >7 and GS ≤7 groups. (I) Kaplan–Meier analysis of recurrence-free survival between the high- and low- THY1 ⁺ CAF groups (log-rank test). (J) ROC curves evaluating the predictive accuracy of the THY1 ⁺ CAF percentage for BCR at 1, 3, and 10 years. (K) Schematic diagram illustrating two CAF subtypes: THY1 ⁻ αSMA ⁺ (left) and THY1 ⁺ αSMA ⁺ (right).

Article Snippet: MACS enrichment: Initial enrichment was performed using human THY1 MicroBeads (Miltenyi Biotec, Cat. No. 130–096–253) with LS columns.

Techniques: Immunohistochemical staining, Biomarker Discovery, RNA Sequencing, Expressing, Multiplex Assay, Immunofluorescence, Staining

Journal: Translational Oncology

Article Title: Spatial and single-cell multi-omics reveal pro-angiogenic THY1 ⁺ fibroblast subtypes predicting prognosis in prostate cancer

doi: 10.1016/j.tranon.2026.102664

Figure Lengend Snippet: Transcriptomic and functional analysis reveals THY1 ⁺ CAFs as a proangiogenic subpopulation in prostate cancer. (A) Analysis of our RNA-seq dataset1 (3193 CAFs). Left: UMAP projection colored according to the six identified CAF clusters. Middle: UMAP visualization of THY1 expression. Right: Violin plot showing the distribution of THY1 expression levels across all six clusters. Statistical significance was determined by one-way ANOVA with Tukey's post hoc test ( p < 0.0001 for clusters 1–3 vs. clusters 4–6). (B) Analysis of our single-cell RNA-seq dataset2 (951 CAFs). Left: t-SNE projection colored according to the five identified CAF clusters. Middle: t-SNE visualization of THY1 expression. Right: Violin plot of THY1 expression across clusters. Statistical significance was determined by one-way ANOVA with Tukey's post hoc test (*: p < 0.001 for clusters 1 and 4 vs. clusters 2 and 3). (C) Volcano plot of differentially expressed genes (DEGs) between THY1 -high (clusters 1–3) and THY1 -low (clusters 4–6) CAFs in our RNA-seq dataset1. (D) Volcano plot of DEGs between THY1 -high (clusters 1–4) and THY1 -low (cluster 5) CAFs in our single-cell RNA-seq dataset2. (E) Venn diagram identifying 121 common genes that were upregulated in THY1 -high clusters in both datasets. (F) Functional enrichment analysis (GO biological process) of the 121 common upregulated genes was performed via Metascape. The bar chart shows the top significantly enriched terms ranked by -log10 (p value). (G) Validation of primary CAF isolation. Top: Representative immunofluorescence images of sorted THY1 ⁺ and THY1 ⁻ CAFs. Middle: Flow cytometry analysis confirming the purity (> 99 %) of the sorted populations. Bottom: Western blot analysis confirming THY1 protein expression in the sorted populations (scale bar: 200 μm). (H) Representative images of the HUVEC tube formation assay after treatment with serum-free medium (control), CM from THY1 ⁺ CAFs, or CM from THY1 ⁻ CAFs. The right panels show magnified views (scale bar: 200 μm). (I, J) Quantification of total tube length (I) and the number of branch points (J) from four independent experiments. The data are presented as the means ± SEMs. ***: p < 0.001; ns, not significant (Student’s t-test).

Article Snippet: MACS enrichment: Initial enrichment was performed using human THY1 MicroBeads (Miltenyi Biotec, Cat. No. 130–096–253) with LS columns.

Techniques: Functional Assay, RNA Sequencing, Expressing, Biomarker Discovery, Isolation, Immunofluorescence, Flow Cytometry, Western Blot, HUVEC Tube Formation Assay, Control

Journal: Translational Oncology

Article Title: Spatial and single-cell multi-omics reveal pro-angiogenic THY1 ⁺ fibroblast subtypes predicting prognosis in prostate cancer

doi: 10.1016/j.tranon.2026.102664

Figure Lengend Snippet: Identification and functional validation of secretory proteins from CAFs. (A) Schematic of the antibody arrays used to detect 507 secretory proteins in CAF-conditioned medium. (B) Antibody array results highlighting selected proteins: CXCL6 (yellow), VEGFA (green), MMP2 (red), and MMP19 (blue). (C) Quantification of fluorescence signals for selected proteins from the antibody arrays. (D, E) ELISA validation of CXCL6 (D) and VEGFA (E) secretion levels in conditioned media from THY1⁺ and THY1⁻ CAFs. (F–K) Effects of CXCL6 on HUVEC tube formation: (F) Representative images of tube formation after treatment with different concentrations of recombinant CXCL6; the bottom row shows magnified views (scale bar: 200 μm, enlarged: 100 μm). (G, H) Quantification of tube length and branch points from the four independent experiments shown in (F). (I) Representative images of HUVECs treated with THY1 ⁺ CAF-1-conditioned medium combined with different concentrations of CXCL6; the bottom row shows magnified views. (J, K) Quantification of tube formation from the four independent experiments shown in (I) (scale bar: 200 μm, enlarged: 100 μm). *: p < 0.05, **: p < 0.01, ***: p < 0.001, ****: p < 0.0001; p values were calculated via paired Student’s t tests.

Article Snippet: MACS enrichment: Initial enrichment was performed using human THY1 MicroBeads (Miltenyi Biotec, Cat. No. 130–096–253) with LS columns.

Techniques: Functional Assay, Biomarker Discovery, Ab Array, Fluorescence, Enzyme-linked Immunosorbent Assay, Recombinant

Journal: Translational Oncology

Article Title: Spatial and single-cell multi-omics reveal pro-angiogenic THY1 ⁺ fibroblast subtypes predicting prognosis in prostate cancer

doi: 10.1016/j.tranon.2026.102664

Figure Lengend Snippet: Functional role of THY1 in regulating the proangiogenic CAF phenotype. (A) Representative images of HUVEC tube formation after treatment with THY1⁺ CAF-1-conditioned medium combined with CXCL6 and the CXCR2 inhibitor SB225002; the bottom row shows magnified views (scale bar: 200 μm, enlarged: 100 μm). (B, C) Quantification of tube formation from four independent experiments. (D) Validation of THY1 knockdown in THY1⁺ CAF1 cells via shRNA, as assessed by qPCR and Western blotting. (E) HUVEC tube formation assay after treatment with CM from THY1 ⁺ CAF1-shNC, THY1 ⁺ CAF1- shTHY1 #1, or THY1 ⁺ CAF1- shTHY1 #2 cells. (F, G) Quantification of tube formation from (E). (H, I) qPCR and ELISA analysis of CXCL6 expression and secretion in shRNA-treated CAFs. (J, K) qPCR and ELISA analysis of VEGFA expression and secretion in shRNA-treated CAFs. ns: not significant.

Article Snippet: MACS enrichment: Initial enrichment was performed using human THY1 MicroBeads (Miltenyi Biotec, Cat. No. 130–096–253) with LS columns.

Techniques: Functional Assay, Biomarker Discovery, Knockdown, shRNA, Western Blot, HUVEC Tube Formation Assay, Enzyme-linked Immunosorbent Assay, Expressing