Fig. 5

Integrin β3 modulates SMC transdifferentiation. a–c Mice were fed a HFD for 6 or 16 weeks as indicated, and then transverse aortic root sections were stained. In a, b sections from ApoE^(−/−), SMMHC-CreER^T2, ROSA26R^(mTmG/+) mice were stained for SMA, GFP (fate marker), nuclei (DAPI), and either integrin β3 (a) or CD36 (b). Dashed yellow lines separate cap from core (a, b) and core from media (b). n = 5. In c sections from ApoE^(−/−) mice that were also wild type or null for Itgb3 were stained for SMMHC, CD68, and nuclei (DAPI). n = 3. Boxed regions (a, c) are shown as close-ups on right; in c CD68⁺SMMHC⁺ cells in the media (arrowheads) and plaque (arrows) of the Itgb3 null atherosclerotic aorta are indicated. Med, tunica media; Lu, lumen; Pl, plaque. Scale bars, 25 μm. d–h Aortic SMCs were isolated from ApoE^(−/−) mice and then subjected to siRNA-mediated knockdown with si-Itgb3 vs. scrambled (Scr; d–f) or with si-Itgb3, si-Tlr4 vs. si-Itgb3 (g, h). Levels of indicated transcripts from qRT-PCR are relative to Gapdh and normalized to either Scr in d, f or to si-Itgb3 in g, h. For d, g, n = 4–5 in duplicate. In e silenced SMCs were cultured with DiI-conjugated ox-LDL for 10 h and stained with DAPI; n = 5. In f, h silenced SMCs were exposed to soluble cholesterol:methyl-β-cyclodextrin complexes for 3 days, and then mRNA levels were assessed; n = 4–7 in duplicate. *^, **^, ***^, ^^, **** vs. control (Scr in d, f and si-Itgb3 in g, h), p < 0.05, p < 0.01, p < 0.005, p < 0.001, and p < 0.0005, respectively. NS, not significant. Student’s t-test was used, and error bars represent standard deviations. i Schematic of the effect of β3 reduction via TLR4 and CD36 in SMCs on transdifferentiation