Extended Data Figure 8. Increased cell proliferation in conditions of p53-deficient CHIP.

a) 20% KO-BMT female mice and 20% WT-BMT controls were fed a high-fat/high-cholesterol (HF/HC) diet for 9 weeks, starting 4 weeks after BMT. Plaque cell proliferation was estimated based on immunohistochemical staining of the Ki-67 proliferation marker (mean±SEM, n=17 20% WT-BMT mice, n=14 20% KO-BMT mice). A two-tailed unpaired t-test was used for statistical analysis. Representative images of Ki-67-stained sections are shown; color deconvolution was applied to show separately the staining of hematoxylin (nuclei) and Ki-67. Atherosclerotic plaques are delineated by dashed lines. Scale bars, 50 μm. b) Cell cycle phase distribution of cultured Trp53−/− and +/+ bone marrow-derived macrophages proliferating asynchronously in the presence of 100 ng/ml MCSF, evaluated by propidium iodide staining of cellular DNA content and flow cytometry (mean±SEM, n=6 Trp53+/+ mice, n=6 Trp53−/− mice). A two-way ANOVA with Sidak’s multiple comparison test was used for statistical analysis. c) Analysis of the proliferation of cultured Trp53−/− and +/+ bone marrow-derived macrophages through immunostaining of BrdU incorporation into the DNA. Quiescent G0-synchronized macrophages were treated with MCSF to induce proliferation (mean±SEM, n=3 Trp53+/+ mice, n=3 Trp53−/− mice). A two-tailed unpaired t-test was used for statistical analysis. A representative experiment is shown; three separate experiments were conducted. Scale bars, 100 μm.