Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Trimethylamine- N -Oxide Affects Cell Type–Specific Pathways and Networks in Mouse Aorta to Promote Atherosclerotic Plaque Vulnerability

doi: 10.1161/ATVBAHA.125.323047

Figure Lengend Snippet: Identification of major aortic cell types and cell type–specific gene markers. A , Schematic diagram of the study design. Athero-prone areas of the aorta (ascending aorta, aortic arch, and thoracic aorta) were collected from female Ldlr −/− mice on normal laboratory (NL), high-cholesterol (HC), or HC+ trimethylamine- N -oxide (TMAO) diets (n=6/group), and 2 mouse samples from the same diet group were pooled together, with n=3 pools/group. B , Plasma TMAO levels (µM) in the NL-, HC-, and HC+TMAO-fed mice as measured by mass spectrometry (n=6/group). Statistical significance was determined by unpaired t test. C through E , Uniform Manifold Approximation and Projection (UMAP) representation of cell clusters in NL, HC, and HC+TMAO conditions, respectively. F , Cluster-specific expression of previously known cell type markers. G , Normalized expression values of top markers of vascular smooth muscle cells (vSMC) subtype clusters: vSMC 1: Acta2, vSMC 2: Atf3 , Rgs5 + vSMC: Rgs5, modulated vSMC: Spp1 . H , Proportions of identified cell types within total cells recovered for each diet condition in order of abundance. Statistical significance was determined by unpaired t test. False discovery rate (FDR) was calculated with Benjamini-Hochberg. DEG indicates differentially expressed genes; and Mod. vSMC, modulated vSMC.

Article Snippet: Human immortalized coronary artery vSMCs (gift from Dr Clint Miller, University of Virginia, Charlottesville, VA) and human primary aortic vSMCs (PCS-100-012; ATCC) were cultured in vSMC growth medium with premixed supplements (catalog no. C-22062; PromoCell).

Techniques: Clinical Proteomics, Mass Spectrometry, Expressing

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Trimethylamine- N -Oxide Affects Cell Type–Specific Pathways and Networks in Mouse Aorta to Promote Atherosclerotic Plaque Vulnerability

doi: 10.1161/ATVBAHA.125.323047

Figure Lengend Snippet: Identification of macrophage subtypes and differences in vascular smooth muscle cells (vSMC)–derived macrophages with trimethylamine- N -oxide (TMAO). A through C , Uniform Manifold Approximation and Projection (UMAP) of macrophage subclusters in normal laboratory (NL), high-cholesterol (HC), and HC+TMAO conditions, respectively. D , Subcluster-specific expression of previously known macrophage subtype markers. E , Proportions of macrophage subtypes within total macrophage number recovered for each diet condition in order of abundance. Statistical significance was determined by unpaired t test. false discovery rate (FDR) was calculated with Benjamini-Hochberg. F , Shared and cell type–specific differentially expressed genes (DEGs; FDR<0.05) induced by TMAO feeding in macrophage subtypes. DEGs unique to a cell type are highlighted in red in the upset plot and histogram, and shared DEGs are indicated in black. The histogram above each plot indicates the DEG counts for each category. DEG direction is indicated by the color of the gene name: red: upregulated, blue: downregulated. G , Top representative pathways from Reactome enriched for vSMC-derived macrophage DEGs (FDR<0.05). ECM indicates extracellular matrix; Max, maximum expression; Min, minimum expression; and Trem2 + , foamy Trem2 + macrophages.

Article Snippet: Human immortalized coronary artery vSMCs (gift from Dr Clint Miller, University of Virginia, Charlottesville, VA) and human primary aortic vSMCs (PCS-100-012; ATCC) were cultured in vSMC growth medium with premixed supplements (catalog no. C-22062; PromoCell).

Techniques: Derivative Assay, Expressing

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Trimethylamine- N -Oxide Affects Cell Type–Specific Pathways and Networks in Mouse Aorta to Promote Atherosclerotic Plaque Vulnerability

doi: 10.1161/ATVBAHA.125.323047

Figure Lengend Snippet: Trimethylamine-N-oxide (TMAO) alters cell-cell communications between major cell types involved in atherosclerosis progression. A , Predicted differential number of interactions between sender (rows) and receiver (columns) cell types comparing high-cholesterol (HC)+TMAO and HC by CellChat. In the heatmap, red represents increased signaling with TMAO feeding, and blue represents decreased signaling. B , Predicted differential number of interactions in the cell-cell communication network between HC+TMAO and HC by CellChat. The color of the network edges represents increased (red) or decreased (blue) signaling with TMAO feeding. C , Specific incoming and outgoing signaling pathways predicted to change in modulated vascular smooth muscle cells (vSMCs) between HC+TMAO and HC by CellChat. Signaling pathways increased and decreased as both incoming and outgoing signals are highlighted in red and blue, respectively. D , Top ligands predicted by NicheNet to target the modulated vSMC differentially expressed gene (DEGs) involved in apoptosis or ECM (extracellular matrix) organization. E , Top ligands predicted by NicheNet to target the vSMC-derived macrophage and Trem2 + macrophage DEGs. In D and E , DEG direction is indicated by red (upregulated with TMAO) or blue (downregulated with TMAO). Mac indicates macrophage; and Mod, vSMC, modulated vSMC.

Article Snippet: Human immortalized coronary artery vSMCs (gift from Dr Clint Miller, University of Virginia, Charlottesville, VA) and human primary aortic vSMCs (PCS-100-012; ATCC) were cultured in vSMC growth medium with premixed supplements (catalog no. C-22062; PromoCell).

Techniques: Protein-Protein interactions, Derivative Assay