Figure 3. Prevention of the aortic challenge–induced expression of genes involved in smooth muscle cell (SMC) death and inflammation in Sting^gt/gt mice.

A-C, Bulk RNA-sequencing (RNA-seq) analysis was performed in ascending aortas from wild-type (WT) mice and Sting^gt/gt mice that were unchallenged or challenged with a high-fat diet (HFD) for 5 weeks and angiotensin II (AngII; 2000 ng/min/kg) infusion during the last week. For each group, 5 aortas were pooled as 1 sample, and duplicate samples were tested. A, Heatmap showing different gene expression patterns among groups. B, Gene ontology (GO) analysis showing that aortic challenge induced the expression of genes involved in several biologic processes (left) and that this induction was prevented in Sting^gt/gt mice (right). C, Heatmap showing that Sting deficiency prevented the challenge-induced upregulation of genes involved in the STING pathway, cell death pathways (apoptosis, necroptosis, and pyroptosis), and the inflammatory response D-I, Single-cell transcriptome analysis was performed in ascending aortas from WT mice and Sting^gt/gt mice that were unchallenged or challenged with a HFD for 5 weeks AngII (2000 ng/min/kg) infusion during the last week. For each group, single cell suspensions from 3 aortas were pooled as one sample. VSMC, vascular smooth muscle cell. D, t-Stochastic neighbor embedding (t-SNE) plots from the ascending aortas of WT mice showing 12 different cell clusters, including 6 clusters of SMCs and 2 clusters of macrophages. E, Sting expression was projected onto t-SNE plots of challenged WT mice showing the high expression of Sting in SMCs, macrophages, and fibroblasts (left). Quantification showing that aortic challenge increased Sting gene expression in SMC cluster 5 and SMC cluster 6 (right). F, GO enrichment analysis of Sting-positive SMCs (cluster 5) showing that aortic challenge induced the expression of several genes with roles in different biologic processes (up) but that this was prevented in challenged Sting- deficient mice (down). G, Heatmap of Sting-positive SMCs (cluster 5) showing that Sting deficiency prevented the challenge-induced expression of genes in inflammation and cell death (apoptosis, necroptosis, and pyroptosis). H, GO enrichment analysis of Sting-positive macrophages (cluster 11) showing that Sting deficiency prevented the aortic challenge-induced expression of several genes. I, Heatmap of Sting-positive macrophages (cluster 11) showing that Sting deficiency prevented the challenge-induced upregulation of genes involved in inflammation.