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. 2021 Mar 5;12:1483. doi: 10.1038/s41467-021-21737-9

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — A Representative examples of CD4⁺ and CD8⁺ T cell staining in the heart of C57BL/6J mice following coronary ligation (MI) or sham surgery. B Kinetic of CD8⁺ T cell infiltration in the myocardium, performed at days 0, 1, 3, 7, and 14 after surgery in MI (white circle, n = 9/12/12/5, respectively, at day 1/3/7/14) and sham-operated mice (black box, n = 6/time point); data are presented as mean ± SEM. C Immunostaining in the ischemic myocardium at day 3 after MI or sham showing CD8⁺ T infiltration (green) in the ischemic heart tissue, scale bar 40 μm. D Kinetic of CD4⁺ T cell infiltration in the myocardium, performed at days 0, 1, 3, 7, and 14 after surgery in MI (white circle, n = 10/9/12/5, respectively, at day 1/3/7/14) and sham-operated mice (black box, n = 6/time point); data are presented as mean ± SEM. E Flow cytometry characterization of CD8⁺ T cell subsets in the ischemic heart at days 1, 3, and 7 after MI including naïve (CD3⁺CD8⁺CCR7^HighCD44⁻), effector memory (CD3⁺CD8⁺CCR7^LowCD44⁺), and central memory (CD3⁺CD8⁺CCR7^HighCD44⁺) subsets. F Representative examples and quantitative analysis of CD8⁺ T cells expressing CD69 or CD107a within the ischemic heart tissue in MI (white circle, n = 6/9/12, respectively, at day 1/3/7) and sham-operated mice (black box, n = 6/time point); data are presented as mean ± SEM. G Immunostaining in the ischemic myocardium at day 3 after MI showing CD8⁺ T cells (green), and Granzyme B (Red) and merged area (Yellow), Inf for infarct area; scale bar 40 μm. H mRNA levels of Granzyme B within the injured myocardium on days 1, 3, and 7 after coronary ligation (gray, n = 5/4/5 at day 1/3/7) or sham (white, n = 4/8/6 at day 1/3/7). I C57Bl6 WT mice received intraperitoneal injection of isotype control (CTR, blue borderline, n = 4/time point) or anti-CD4 depleting monoclonal antibody (blue filled, n = 5/time point) (150 μg/mice) 1 day before coronary occlusion. CD4⁺ T cell depletion was confirmed in the heart by flow cytometry at day 1 and day 3 following MI. J, K Quantification and representative example of CD8⁺ T cell count in the heart of control (n = 4/time point) or CD4-depleted (n = 5/time point) mice at day 1 and day 3 after MI. P values were calculated using two-tailed Mann-Whitney test (B, D, F, H, I, J). Inf, infarct; MI, myocardial infarction; FMO, fluorescent minus one.