Journal: Nature Communications

Article Title: Spatially organized tumor-stroma boundary determines the efficacy of immunotherapy in colorectal cancer patients

doi: 10.1038/s41467-024-54710-3

Figure Lengend Snippet: a Bubble plots represent the expression profiling of PDCD1 and CD274 in single cell clusters from scRNA-seq dataset. The plots are sized by the fraction of cells with positive gene expression, while the color represents the gene expression level . b The interactions among indicated cell clusters through PD1-PD-L1 axis from scRNA-seq dataset. c The box and whisker plot shows show the nearest distance from DC_ LAMP3 to CD4_ CXCL13 (N number of the counted cell-cell distances: distant stroma =918, boundary region=1867; tumor center=251) or ( d ) to CD8_ CXCL13 (N number of the counted cell-cell distances: distant stroma =368, boundary region=567; tumor center=45) in distance stroma, tumor-stroma boundary and tumor center from treatment naïve dMMR patients. Data are represented as mean±IQR and analyzed by unpaired 2-tailed Student t test. ns, not significant; *, p < 0.05; **, p < 0.01. Pie charts represent the fractions of CD4_ CXCL13 or CD8_ CXCL13 in the indicated regions stratified by colors in accordance with ( c ) d . Data are analyzed by unpaired Student-t test. ns, not significant; ****, p < 0.0001. e Representative mIF images of panCK, COL1A1, PD-L1 and PD1 in treatment naïve dMMR patients (5 samples were analyzed). DAPI was used as a positive control for cell nuclei staining. Scale bars were set as 5 mm for global image (left) and 50μm for FOVs (middle and right). f The box and whisker plot shows the nearest distances from PD1 + cells to PDL1 + cells in distance stroma, tumor-stroma boundary and tumor center from 7 treatment naïve dMMR patients. Each dot represents a PD1-expressing cell. N number of the counted cell-cell distances: stroma = 750; boundary = 935; tumor = 892. Data are represented as mean ± IQR and analyzed by unpaired 2-tailed Student t test with Bonferroni correction. Ns, not significant; *, p < 0.05; **, p < 0.01; ***, p < 0.001; ***, p < 0.0001. g The box and whisker plot shows the T cell exhaustion scores in the distant stroma, tumor-stroma boundary and tumor center from treatment naïve dMMR patients. Data are represented as mean±IQR and analyzed by unpaired 2-tailed Student t test. Ns not significant; *, p < 0.05; **, p < 0.01; ***, p < 0.001; ***, p < 0.0001. N number of the spots: distant stroma =86, boundary region=12; tumor center=287. h Representative images of H&E staining, FOV, and spatial map of T cell exhaustion scores in the tumor-stroma boundary of the treatment naïve dMMR patient #56. Scale bars were set as 1.5 mm for global image (left) and 500 μm for FOVs . i Heatmap of PDCD1 and CD274 expressions in the distant stroma, tumor-stroma boundary and tumor center of treatment naïve dMMR patients.

Article Snippet: Anti-COL1A1 rabbit mAb , 1:500 , Cell Signaling , CST72026S.

Techniques: Expressing, Gene Expression, Whisker Assay, Positive Control, Staining

Journal: Nature Communications

Article Title: Spatially organized tumor-stroma boundary determines the efficacy of immunotherapy in colorectal cancer patients

doi: 10.1038/s41467-024-54710-3

Figure Lengend Snippet: a The top 5 GO terms of DEGs (p value < 0.001; fold change >2) identified from CAF_ CXCL14 and CAF_ CXCL8 clusters are shown. Comparison is made by two-tailed t test. b GSEA plot of the upregulated genes related to extracellular matrix organization in CAF_ CXCL14 (n cells=268) compared to CAF_ CXCL8 ( n cells=118) is shown. Comparison is made by two-tailed t test. c RCTD frequencies of CAF_ CXCL14 (up) and ECM organization signature scores (below) in the tumor-stroma boundary of dPR/dCR (n spots=1436) and dSD (n spots=2244) patients are shown as violim plots. Data are analyzed by unpaired 2-tailed Student t test. Ns, not significant; *, p < 0.05; **, p < 0.01; ***, p < 0.001; ***, p < 0.0001. d Pearson correlation of RCTD frequencies of CAF_ CXCL14 and ECM organization signature scores in dPR/dCR and dSD patients. e Loess smoothed curves and ( f ) violin plots of RCTD frequencies of CAF_ CXCL14 (up) and ECM organization signature scores (below) in tumor ( + 300 μm, n spots = 9313) to the tumor-stroma boundary (0 μm, n spots = 56,357) from treatment naïve pMMR and dSD patients are shown. The tumor edge region ( + 150 μm to 0 μm) is highlighted in the yellow frame. g Representative images of FOV, CAF_ CXCL14 and ECM organization scores in the tumor-stroma boundary of treatment naïve pMMR patient #59 and dSD patient #95 are shown. h Representative images of Masson’s trichrome staining from treatment naïve dMMR and pMMR patients (3 samples were analyzed in each group). Scale bar = 50 μm. i Representative mIF images of panCK, COL1A1, CD3 and CXCL14 in indicated patient groups (3 samples were analyzed in each group). DAPI was used as a positive control for cell nuclei staining. Scale bars, 50 μm. j CXCL14 expression level and ECM organization scores in fibroblasts from the three public scRNAseq datasets , , .

Article Snippet: Anti-COL1A1 rabbit mAb , 1:500 , Cell Signaling , CST72026S.

Techniques: Comparison, Two Tailed Test, Staining, Positive Control, Expressing

Journal: Nature Communications

Article Title: Spatially organized tumor-stroma boundary determines the efficacy of immunotherapy in colorectal cancer patients

doi: 10.1038/s41467-024-54710-3

Figure Lengend Snippet:

Article Snippet: Anti-COL1A1 rabbit mAb , 1:500 , Cell Signaling , CST72026S.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: Wanted: Dead or Alive Cells with Propidium Iodide Staining in Liver Tissue

doi: 10.3390/ijms252413521

Figure Lengend Snippet: Co-staining of PI with a fluorescence-labeled collagen antibody. Collagen antibody is conjugated to Alexa Fluor 488 (green) and was co-stained with PI (red) and DAPI (blue) in liver cryosections of mice treated with EtOH + CCl4. Collagen1a1 (Col1a1) stained the fibrotic tissue characteristic from these treated mice. Merge image and the augmentation showed the region in the liver tissue where the cells are positive for PI and Col1a1 (yellow). Images were taken at 20× magnification, scale bar = 500 μm.

Article Snippet: Anti-col1a1 pre-labeled (Alexa488) , Cell Signaling Technology , Danvers, MA, USA , 28368S.

Techniques: Staining, Fluorescence, Labeling

Journal: International Journal of Molecular Sciences

Article Title: Wanted: Dead or Alive Cells with Propidium Iodide Staining in Liver Tissue

doi: 10.3390/ijms252413521

Figure Lengend Snippet: Reagents and resources that were used for this study together with their source and identifier.

Article Snippet: Anti-col1a1 pre-labeled (Alexa488) , Cell Signaling Technology , Danvers, MA, USA , 28368S.

Techniques: Fluorescence, Microscopy, Saline, Staining

Journal: Biomolecules & therapeutics

Article Title: Dimethyl α-Ketoglutarate Promotes the Synthesis of Collagen and Inhibits Metalloproteinases in HaCaT Cells.

doi: 10.4062/biomolther.2023.131

Figure Lengend Snippet: Fig. 2. DMK increases the level of 4-hydroxyproline and promotes the synthesis of collagen in HaCaT cells. (A) Diagram depicting the con- version of proline to hydroxyproline by prolyl 4-hydroxylase (P4H). (B) HaCaT cells (2×10 6 cells) were exposed to DMK at various times, and the mRNA level of P4H was measured by real-time RT-PCR (n=4). (C) HaCaT cells (1×10 5 cells) were exposed to DMK for 24 h, and the level of 4-hydroxyproline was measured by an immunocytochemical assay. (D) HaCaT cells (1×10 6 cells) were exposed to DMK at vari- ous times, and the production of collagen type 1α1 (COL1A1) was measured by the collagen type 1α1 ELISA assay (n=3). (E) HaCaT cells (2×10 6 cells) were exposed to DMK at various times, and the mRNA level of COL1A1 was measured by real-time RT-PCR (n=4). *p<0.05, **p<0.01, and ***p<0.001 vs Control (DMSO).

Article Snippet: HaCaT cells were seeded at a density of 1×106 cells in 60 mm culture plates, and the amount of human collagen 1α1 was measured using a FastScanTM COL1A1 ELISA Kit (Cell Signaling Technology) according to the manufacturer’s protocol.

Techniques: Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Control