t24  (ATCC)

Journal: bioRxiv

Article Title: Pan-cancer circular genomics identifies intratumoral Staphylococcus lugdunensis as a metabolic driver in bladder cancer

doi: 10.64898/2026.04.14.717102

Figure Lengend Snippet: (A) Heatmap illustrating Spearman rank correlations between bacterial abundance and the infiltration proportions of diverse immune cell populations within the tumor microenvironment. (B) Real time proliferation curves of T24 BLCA cells following exposure to live S. lugdunensis at a multiplicity of infection of 100 to 1 compared to BHI medium control. Data represent the mean plus or minus standard error of the mean (SEM). (C) Representative confocal fluorescence imaging of T24 tumor cells following co culture with S. lugdunensis . Green fluorescent signals denote HADA labeled S. lugdunensis , which were observed localized within malignant cells visualized by red fluorescent actin tracker staining. (D) Volcano plot showing predicted bacterial metagenomic functional pathways and modules via PICRUSt2. (E and F) Volcano plots of differentially abundant lipid metabolites identified through targeted lipidomics, comparing S. lugdunensis lysates versus BHI medium (E) and S. lugdunensis positive tumors versus NATs (F). (G) Venn diagram depicting the intersection of enriched lipid species between bacterial lysates and tumor tissues, identifying LPC14:0 as a candidate effector molecule. (H) Normalized abundance of LPC14:0 in S. lugdunensis (SL) negative and positive tumor tissues. (I) Box plots showing the accelerated proliferation of T24 cells treated with exogenous LPC14:0 relative to vehicle control (six replicates/group). (J) Functional enrichment analysis of differentially expressed genes reveal that the PPAR signaling pathway was significantly enriched in the LPC14:0 treatment group. (K) Quantitative assessment of cellular fatty acid uptake in T24 cells following treatment with vehicle (NC) or 30 µM LPC14:0 for 18 hours. Data are expressed as relative background-subtracted fluorescence intensity (ΔRFU) normalized to total protein concentration (12 replicates/group). (L) Biochemical analysis of FAO activity in T24 cells under the same treatment conditions (six replicates/group). Two-tailed Student’s t -test: *, P < 0.05; **, P < 0.01; ***, P < 0.001; See also Figure S5 and Table S5.

Article Snippet: FAO capacity was assessed using a Fatty Acid Oxidation (FAO) Colorimetric Assay Kit (Elabscience, Cat# E-BC-K784-M) in T24 cells (treated similarly to those in the fatty acid uptake assay) and in subcutaneous tumors harvested from the in vivo models.

Techniques: Infection, Control, Fluorescence, Imaging, Co-Culture Assay, Labeling, Staining, Functional Assay, Protein Concentration, Activity Assay, Two Tailed Test

Journal: bioRxiv

Article Title: Pan-cancer circular genomics identifies intratumoral Staphylococcus lugdunensis as a metabolic driver in bladder cancer

doi: 10.64898/2026.04.14.717102

Figure Lengend Snippet: (A) Schematic representation of the experimental workflow in balb/c Null hairless mice. T24 bladder cancer cells were subcutaneously injected, and mice were subsequently treated with vehicle, Escherichia coli , or S. lugdunensis at the fifth days. Tumor volume was monitored over 30 days, showing that S. lugdunensis significantly accelerated tumor growth. (B) Representative tumor images of resected tumors at day 30 and corresponding bacterial colony forming units recovered from the tumor tissues, confirming successful bacterial colonization. (C) Quantitative analysis of terminal tumor weight and longitudinal mouse body weight across the three experimental groups (N=8/group). (D) Difference of relative expression of LPC14:0 across the three experimental groups. (E) Representative immunohistochemistry (IHC) images of Ki67 staining in tumor sections. Scale bars represent 100 microns. (F) The bar graph shows the quantification of Ki67 positive cells, indicating enhanced cellular proliferation in the S. lugdunensis group. (G) Biochemical analysis of FAO activity in mice tumor under the S. lugdunensis conditions (N=4/group). (H) Experimental design for the exogenous administration of the LPC14:0. Mice harboring T24 tumors received intratumoral injections of vehicle or LPC14:0. Growth curves demonstrated that LPC14:0 significantly enhanced tumor expansion. (I) Gross morphology of tumors harvested from the vehicle and LPC14:0 treated groups at the study endpoint. (J) Statistical comparison of final tumor weights and mouse body weights between the vehicle and LPC14:0 groups (N=7/group). (K-L) IHC analysis of Ki67 expression in tumor tissues from mice treated with LPC14:0. Scale bars represent 100 microns. The percentage of Ki67 positive cells was significantly higher in the LPC14:0 group, confirming the pro proliferative effect of the metabolite. (M) Biochemical analysis of FAO activity in mice tumor under the LPC14:0 treatment conditions (N=4/group). Two-tailed Student’s t -test: *, P < 0.05; **, P < 0.01; ***, P < 0.001; See also Figure S5 and Table S6.

Article Snippet: FAO capacity was assessed using a Fatty Acid Oxidation (FAO) Colorimetric Assay Kit (Elabscience, Cat# E-BC-K784-M) in T24 cells (treated similarly to those in the fatty acid uptake assay) and in subcutaneous tumors harvested from the in vivo models.

Techniques: Injection, Expressing, Immunohistochemistry, Staining, Activity Assay, Comparison, Two Tailed Test

Journal: bioRxiv

Article Title: Pan-cancer circular genomics identifies intratumoral Staphylococcus lugdunensis as a metabolic driver in bladder cancer

doi: 10.64898/2026.04.14.717102

Figure Lengend Snippet: (A) Schematic representation of the chemical proteomics workflow utilizing a bifunctional photo-crosslinking probe (*LPC14:0) for target protein capture, visualization, and identification in T24 cells. (B) In situ fluorescence imaging of T24 cells following incubation with *LPC14:0 and bioorthogonal conjugation with Azide 594 (red). Specific binding is demonstrated by competitive binding of the probe signal with 10-fold excess native LPC14:0 (100 µM). Nuclei were counterstained with DAPI (blue). Scale bars, 20 µm. (C) Venn diagram showing the intersection of MS-identified LPC14:0-interacting proteins with the PPAR signaling pathway, pinpointing PPARδ as a primary candidate. (D, E) GSEA pathway enrichment analysis in tumor tissues from mice treated with S. lugdunensis (D) or LPC14:0 (E). Significant enrichment of pathways is ranked by Normalized Enrichment Scores (NES). (F, G) Volcano plots representing differentially expressed genes in S. lugdunensis -treated (F) and LPC14:0-treated (G) tumors. (H) Molecular docking model illustrating the binding configuration of LPC14:0 (green sticks) within the active pocket of PPARδ (gray ribbons) (Vina score=-6.5; Cavity volume= 408 Å). The inset provides a detailed view of stable hydrogen bonds formed with amino acid residues Thr292 and Thr289 (yellow dashed lines; distances <3.5 Å). Atoms are color coded as follows: red, oxygen; orange, phosphorus; blue, nitrogen. (I, J) Kaplan-Meier curves of overall survival in bladder cancer patients (TCGA cohort, via GEPIA2) stratified by CD36 (I) or ACOX2 (J) expression levels. Statistical significance was determined by the log-rank test. (K) A proposed working model: S. lugdunensis secretes LPC14:0 into the tumor tissue, where it directly engages and activates host PPARδ. This activation triggers the transcriptional upregulation of fatty acid transporters and oxidases, fueling malignant progression through enhanced lipid uptake and bioenergetic demand. *, P < 0.05; **, P < 0.01; ***, P < 0.001; See also Figure S5 and Table S7.

Article Snippet: FAO capacity was assessed using a Fatty Acid Oxidation (FAO) Colorimetric Assay Kit (Elabscience, Cat# E-BC-K784-M) in T24 cells (treated similarly to those in the fatty acid uptake assay) and in subcutaneous tumors harvested from the in vivo models.

Techniques: In Situ, Fluorescence, Imaging, Incubation, Conjugation Assay, Binding Assay, Expressing, Activation Assay