Senescent cardiomyocytes contribute to cardiac dysfunction following myocardial infarction

Abstract

Myocardial infarction is a leading cause of morbidity and mortality. While reperfusion is now standard therapy, pathological remodelling leading to heart failure remains a clinical problem. Cellular senescence has been shown to contribute to disease pathophysiology and treatment with the senolytic navitoclax attenuates inflammation, reduces adverse myocardial remodelling and results in improved functional recovery. However, it remains unclear which senescent cell populations contribute to these processes. To identify whether senescent cardiomyocytes contribute to disease pathophysiology post-myocardial infarction, we established a transgenic model in which p16 (CDKN2A) expression was specifically knocked-out in the cardiomyocyte population. Following myocardial infarction, mice lacking cardiomyocyte p16 expression demonstrated no difference in cardiomyocyte hypertrophy but exhibited improved cardiac function and significantly reduced scar size in comparison to control animals. This data demonstrates that senescent cardiomyocytes participate in pathological myocardial remodelling. Importantly, inhibition of cardiomyocyte senescence led to reduced senescence-associated inflammation and decreased senescence-associated markers within other myocardial lineages, consistent with the hypothesis that cardiomyocytes promote pathological remodelling by spreading senescence to other cell-types. Collectively this study presents the demonstration that senescent cardiomyocytes are major contributors to myocardial remodelling and dysfunction following a myocardial infarction. Therefore, to maximise the potential for clinical translation, it is important to further understand the mechanisms underlying cardiomyocyte senescence and how to optimise senolytic strategies to target this cell lineage.

Fig. 1. Cardiomyocyte specific knock out of p16 reduces cardiomyocyte senescence and senescence associated secretory phenotype.

a Schematic of transgenic cardiomyocyte p16 knock-out mouse. b Experimental design. c Percentage p16⁺ CMs in the peri-infarct region at 5 weeks post IR. d Representative images of p16 staining. Yellow arrow, p16⁺ CM and white arrow p16⁺ interstitial cell (p16 red, Troponin-C green, DAPI blue), n = 4/group. Scale bars 20 µm. e Percentage p21⁺ CMs in the peri-infarct region at 5 weeks post IR. f Representative images of p21 staining. Yellow arrows, p21⁺ CM (p21 red, Troponin-C green, DAPI blue), n = 4 per group. Scale bars 20 µm. g Clustered heatmap of cytokine protein levels in LV myocardium. h Expression of individual protein levels in the LV myocardium of heart in the indicated experimental groups. i Total number of cardiomyocytes with TAF and mean of TAF per cardiomyocyte at 5 weeks post IR. n = 3/group. j Representative images of TAF, γH2AX co-localised with Telomere immuno-FISH (telo-FISH red, γH2AX green, WGA white). Images are obtained from the z-stacks of 10 μm sections. Yellow arrow indicates a TAF. Scale bars top panel 20 µm and bottom panel 2.5 µm. Data are mean ± SEM, *P < 0.05 ***P < 0.05 using Student’s T test or Mann–Whitney test.

Fig. 2. Cardiomyocyte specific inhibition of senescence improves functional outcome, reduces scar size and attenuates senescence in interstitial cells following IR.

a Examples of individual short axis cine-MR images. b Ejection fraction and LV mass at 5 weeks post-IR, n > 6/group. c Mean CM cross-sectional area μm². N = 7/group. d Representative images of WGA staining for quantification of CM area. Scale bars 20 µm. e Quantification of infarct size relative to total LV area. N = 7/group. Scale bars 1 mm. f Representative images of Masson’s trichrome staining. g Percentage p16⁺ interstitial cells in the peri-infarct region at 5 weeks post IR. h Representative images of p16 staining. Yellow arrows, p16⁺ CMs and white arrows, p16⁺ interstitial cells (p16 red, Troponin-C green, DAPI blue), n = 4/group. Scale bars 20 µm. i Representative image of SA-β-gal staining at 5 weeks post IR and quantification of SA-β-Gal⁺ interstitial cells per field of view (FOV) in the peri-infarct region at 5 weeks post IR, n = 3. Scale bars 50 µm. Data are mean ± SEM. *P < 0.05, **P < 0.01 using Student’s T test or Mann–Whitney Test.