Figure 3: EndoMT and MMT are markedly upregulated in lesions lacking SMC investment.

Loss of Pdgfrb signaling significantly increased the proportion of non-SMC derived ACTA2⁺ fibrous cap following 18 weeks of WD feeding. ***p-value < 0.0001.(A) SMC account for only about 60% of the ACTA2⁺ fibrous cap cells in Pdgfrb^SMC-WT/WT mice. Representative images of MMT (B) and EndoMT. (C). (D) Marker protein staining and single-cell counting indicates that up to 40% of ACTA2⁺ cells were derived from EndoMT, ***p-value = 0.0006, or MMT, **p-value = 0.0059, in Pdgfrb^SMC-WT/WT lesions, with these contributions increasing dramatically in Pdgfrb^SMC-Δ/Δ mice. (E) EC-lineage-tracing mice (Cdh5-eYFP) were fed a WD for 18 weeks and analyzed for EC-derived cells within the fibrous cap. (F) Representative image of a lesion from the Cdh5-eYFP mice. (G) Quantification shows about 20% of ACTA2⁺ cells within the fibrous cap were derived from EC. (H) Cdh5-eYFP mice underwent lethal irradiation and BMT with tdTomato⁺ bone marrow prior to WD feeding to ablate SMC accumulation within BCA lesions. (I) Representative image showing loss of SMC within Myh11-eYFP lesions (left) and accumulation of EC within Cdh5-eYFP lesions (right) after lethal radiation. (J) Cdh5-eYFP⁺ cells account for nearly 70% of the ACTA2⁺ fibrous cap cells following radiation-induced loss of SMC accumulation. BM-tracing shows nearly 16% of ACTA2⁺ fibrous cap cells are tdTomato⁺ and thus of bone marrow origin. Scale bar: 100μm (B, C, F, I) or 20μm (F, I; zoom). Graphs were analyzed using two-tailed Mann-Whitney U test (D) or two-way ANOVA with Sidak correction and multiple comparisons (A) biologically independent animals are indicated as individual dots in (D) and n = 8, 9, 6 (WT: 300, 450, 750 μm) or n = 8, 8, 7 (KO) in (A), n= 12, 13 for each location (G), n= 12 (J), error bars represent mean ± SEM, p-values displayed refer to two-way ANOVA between genotype unless otherwise indicated.