Journal: Cancer Research

Article Title: Single-Cell Analysis Identifies NOTCH3-Mediated Interactions between Stromal Cells That Promote Microenvironment Remodeling and Invasion in Lung Adenocarcinoma

doi: 10.1158/0008-5472.CAN-23-1183

Figure Lengend Snippet: A, Left, UMAP highlighting lymphatic ECs and vascular ECs. Middle, vascular ECs colored by cell subclusters. Top right, cells in ANT tissues; bottom right, cells in T tissues. B, Boxplot visualizing the relative fractions of each endothelial subcluster to all vascular EC populations in T (red) and ANT (blue). The statistical significance was calculated using a Wilcoxon rank-sum test. C, The average expression of marker genes used for EC subcluster labeling. D, Bar chart visualizing significant cell–cell contact signaling pathways between mesenchymal cells and ECs generated using CellChat, where the relative strength in T was colored in red and ANT was in green, and labels were colored in red if the signal was significantly enriched in T comparing to ANT, or green if significantly enriched in ANT. The pathways are ranked on the basis of their differences of relative information flow between T and ANT. E, Heat map visualizing the relative signaling strengths of significant T-enriched pathways in D among endothelial and mesenchymal cells generated using CellChat. The top bar plot represents the total signaling strength of all displayed signaling pathway in each cell group. The right gray bar plot shows the total signaling strength of all displayed cell groups in each signaling pathway. F, Chord diagram visualizing the information flow strength of NOTCH signaling pathway from ECs to mesenchymal cells. The chords were color-coded by the signal senders or receivers of represented ligand–receptor pairs. NOTCH3 expressed by pericytes is shown to be the dominant receptor. G, Forest plot visualizing the significances of NOTCH pathway receptor expression enrichment in mesenchymal cells among leave-one-donor-out iterations. The interquartile range of Benjamini–Hochberg P adj values is shown. If there was no significant enrichment between T and NAT before removing any donors, the data points would be omitted. The red dash lines represent P = 0.05. H, Forest plots visualizing the significances of NOTCH pathway ligands expression enrichment in ECs among leave-one-donor-out iterations. I, Immunofluorescent imaging of a tumor sample from the scRNA-seq cohort. NOTCH3 staining, green; MCAM staining, orange; DAPI nuclei staining, blue. Scale bar, 100 μm. White arrow, MCAM + NOTCH3 + cells. J, Boxplots showing the log 10 expression of NOTCH receptor genes between T and ANT LUAD samples in TCGA. For genes with significantly different expression between T and ANT (Wilcoxon rank sum test, Benjamini–Hochberg P adj < 0.05), the receiver operating characteristic – area under the curve (ROC-AUC) values were calculated and are in red (T-enriched) or green (ANT-enriched). *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: The Gene set pathway enrichment analysis was performed using the combination of Kyoto Encyclopedia of Genes and Genomes (KEGG; ref. ) and GeneGo MetaCore Pathways.

Techniques: Expressing, Marker, Labeling, Protein-Protein interactions, Generated, Imaging, Staining

Journal: Cancer Research

Article Title: Single-Cell Analysis Identifies NOTCH3-Mediated Interactions between Stromal Cells That Promote Microenvironment Remodeling and Invasion in Lung Adenocarcinoma

doi: 10.1158/0008-5472.CAN-23-1183

Figure Lengend Snippet: A, Spearman correlation between global genes and NOTCH3 expression in the TCGA LUAD tumor dataset ( x -axis) and collaboration LUAD tumor dataset ( y -axis). NOTCH3 and COL1A1 are highlighted in red. The legend presents the correlation value of gene COL1A1 with NOTCH3 expression: 0.50 for TCGA dataset and 0.46 for Collaboration dataset. B, Dot plots of top 10 pathways enriched in KEGG or GeneGo pathways using genes with NOTCH3 correlations over 0.4(d). Colors indicate P values; sizes of the dots indicate overlap gene counts in the pathways. C – E, Cox proportional hazards analysis showing the predictivity of NOTCH genes in LUAD samples under different stromal level conditions. Genes with significant predictivities ( P < 0.05) were color-coded by their HR value. Red, poor prognosis; blue, better prognosis. Black dots, nonsignificant genes. F and G, The Kaplan–Meier survival analysis depicting the prognostic value of GEP expression levels in NOTCH3 - high ( F ) and NOTCH3 -low ( G ) LUAD samples. A total of 1,434 LUAD samples with corresponding overall survival (OS) data in the Collaboration dataset were evenly divided into two groups by NOTCH3 expression level. HRs were derived from a Cox proportional model fit; no multiple testing. The predictivity of GEP was only sufficient in NOTCH3 -low samples. H, Spearman correlation between NOTCH3 and consensus gene signatures, added to the global gene correlation, in the TCGA LUAD tumor dataset ( x -axis) and Collaboration LUAD tumor dataset ( y -axis). The legend shows the correlation values, with the first value representing the TCGA dataset and the second value representing the Collaboration dataset. I, Graph illustration of the interaction between mesenchymal and ECs via the NOTCH pathway in the TME. The interaction activates mesenchymal cells and leads to collagen deposition and cell invasion. ( I, Created with BioRender.com.)

Article Snippet: The Gene set pathway enrichment analysis was performed using the combination of Kyoto Encyclopedia of Genes and Genomes (KEGG; ref. ) and GeneGo MetaCore Pathways.

Techniques: Expressing, Derivative Assay