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. Author manuscript; available in PMC: 2015 Dec 1.
Published in final edited form as: Nat Med. 2015 May 18;21(6):628–637. doi: 10.1038/nm.3866

Figure 3. SMCs within human coronary artery lesions express the Mϕ marker CD68.

Figure 3

(a) SMCs within advanced atherosclerotic lesion specimens were identified based on PLA detection of the SMC specific stable epigenetic signature H3K4dime on the MYH1127. MYH11 H3K4dime PLA+ cells exhibit a punctate red dot within the nucleus while the non-nuclear amorphous red staining is autofluorescence or non-specific background. (a) Samples were also immuno-stained for CD68 (green), and DAPI (blue). Results showed three distinct cell populations highlighted in enlarged panels to the right and indicated with white arrows: (i) MYH11 H3K4dime PLA+ SMCs that are CD68−, (ii) MYH11 H3K4dime PLA−CD68+ (HSC-derived Mϕs), and (iii) H3K4dime MYH11 H3K4dime PLA+CD68+ SMC-derived Mϕ-like cells. Scale bar = 100 μm. (b) Shoulder regions within plaques [stained with DAPI (blue), ACTA2 (green), PLA (red), and CD68 (cyan)] exhibited a high incidence of SMC-derived Mϕ-like cells (MYH11 H3K4dime PLA+CD68+) (yellow arrows) and several phenotypically modulated SMCs negative for CD68 (MYH11 H3K4dime PLA+ACTA2− CD68−) (white arrows). Scale bar = 50 μm. (c) Quantitative analysis of SMC-derived Mϕ-like cells within human coronary lesions based on MYH11 H3K4dime ISH-PLA +/− adjustment for the efficiency of PLA27. Error bars = S.E.M. for 12 independent samples of human atherosclerosis in the right coronary artery. (d) Combined epigenetic SMC and genetic HSC lineage tracing analyses of cross gender human heart transplant samples. Coronary artery specimens from a male patient who received a female heart were processed for MYH11 H3K4dime PLA (red), Y-chromosome FISH (green), and CD68 staining (yellow). Results show cells that were MYH11 H3K4dime PLA+ Y-chromosome− and CD68+ (yellow arrows) reflecting a SMC-derived Mϕ-like cell not of hematopoietic origin (top). In contrast, Mϕs of hematopoietic origin are MYH11 H3K4dime PLA−Y-chromosome+CD68+ (red arrows) (bottom). Scale bar = 50 μm.