Figure 2.

Acute mental stress exposure aggravates inflammatory blood leucocyte migration into atherosclerotic plaques. (A, B) Gating and quantification of GFP^high myeloid cells in atherosclerotic aortas after adoptive transfer of GFP^high monocytes and neutrophils into non-stressed vs. stressed ApoE^−/− mice (21 h after a single 3 h episode of restraint stress, n = 5 per group, Student’s t-test). (C, D) Gating and quantification of aortic myeloid cells from non-stressed vs. stressed ApoE^−/− mice (n = 6 per group, Mann–Whitney U-test for Ly6C^high monocytes, Student’s t-test for neutrophils and macrophages). (E, F) Representative images (myeloid cells, CD11b^high) and quantification of adherent myeloid cells in non-stressed vs. stressed ApoE^−/− mice using intravital microscopy on carotid arteries (immediately after a single 1 h episode of restraint stress, n = 7–10 per group, Student’s t-test, Mann–Whitney U-test for neutrophils). Scale bars represent 100 µm. (G, H) Representative immunohistochemical staining of aortic roots from non-stressed vs. stressed ApoE^−/− mice (after three episodes of 3 h restraint stress once daily) for myeloid cells (CD11b). Scale bars represent 200 µm. Bar graphs show the percentage of positive area per plaque area (n = 8 per group, Student’s t-test). (I) Quantification of GFP^high myeloid cells in aortas after adoptive transfer of GFP^high monocytes and neutrophils into non-stressed vs. stressed non-atherosclerotic wild-type mice (21 h after a single 3 h episode of restraint stress, n = 12 per group, Mann–Whitney U-test). (J) Quantification of aortic myeloid cells from non-stressed vs. stressed non-atherosclerotic wild-type mice (n = 10–12 per group, Student’s t-test). (K, L) Representative images and quantification of plaque rupture in carotid arteries from non-stressed vs. stressed ApoE^−/− mice (after three episodes of 3 h restraint stress once daily; N, necrotic core; F, fibrous cap; L, arterial lumen; T, thrombus). Thrombus formation is considered an indicator for plaque rupture. Scale bars represent 50 µm. Bar graphs show the incidence of plaque rupture (n = 18–20 per group, Fisher’s exact test). Numbers next to gates indicate frequencies (%). Data are presented as mean ± SD.