Objectives: Ischemic cardiovascular diseases, particularly myocardial infarction (MI), are associated with an increased mortality and morbidity worldwide. The recruitment of myeloid cells after MI has a decisive impact on the inflammatory response and the healing process. Besides, macrophage polarization towards an anti-inflammatory M2 phenotype was found to improve cardiac healing after MI. In contrast, overwhelming inflammation may favour adverse cardiac remodeling and subsequent heart failure. The enzyme Peptidylarginine deiminase 4 (PAD4) was recently found to be critically involved in cardiac regeneration after MI. PAD4-mediated histone citrullination and subsequent chromatin decondensation are essential for gene regulation and also for the formation of neutrophil extracellular traps. In this study, we investigated the effects of PAD4 deficiency on cardiac inflammation and regeneration after MI in mice.

 

Methods: MI was induced by permanent ligation of the left coronary artery (LAD) in 9-12 weeks old, male C57BL/6J (WT) and PAD4-/- mice. On days 1, 3 and 7 after MI, cardiac recruitment of myeloid cells and reactive oxygen species (ROS) were assessed by FACS analysis and malondialdehyde quantification, respectively. The infarct sizes were quantified by 2,3,5-triphenyltetrazolium chloride (TTC) staining and cardiac function was analysed by echocardiography at days 1, 7 and 28 post-surgery. Kaplan-Meier survival analysis was performed to compare survival of the different genotypes after MI.

 

Results: PAD4-/- mice displayed significantly diminished cardiac ROS levels and infarct sizes at days 1 and 7 after MI. Additionally, these mice showed reduced leukocyte recruitment and increased number of cardiac lymphocytes at day 1 after MI. Of note, the number of leukocytes and monocytes/macrophages increased significantly over time and more anti-inflammatory M2-like macrophages were detected in the infarcted hearts of PAD4-/- mice at day 7. On the contrary, leukocyte counts strongly decreased in WT mice at the same time. PAD4-/- mice displayed improved cardiac function and reduced remodeling, as demonstrated by increased ejection fraction, fractional shortening and LV wall thickness. Accordingly, PAD4-/- mice exhibited increased survival rates compared with WT mice.

 

Conclusions: PAD4 deficiency markedly influences cardiac recruitment of immune cells, ROS generation and the extent of cardiac damage after MI. Therefore, inhibition of PAD4 may reduce adverse remodeling and mortality after MI.