Activated inflammatory cells in human atrial myocardium from patients with atrial fibrillation

Introduction:  The processes underlying the initiation and development of atrial fibrillation (AF) are still not sufficiently explored. This most common arrhythmia is associated with an increase in mortality risk that is strongly related with old age. One of the generally recognized factors contributing to persistency of AF is structural remodeling of the myocardium. There is alteration in both cardiomyocyte morphology as well as changes affecting endomysium.

Aim: In this study we focused on morphological and functional changes in endomysium of atrial myocardium with special focus on immunohistochemical detection of CD45+- cells, representing the whole leukocyte population. In addition, we began to characterize different types of infiltrating leukocytes.

Methods:   We analyzed atrial biopsies obtained from patients undergoing bypass or mitral valve surgery. The patients had a regular sinus rhythm or were suffering from AF. The atrial samples were fixed with 4% paraformaldehyde and embedded into paraffin. Sections from atrium were histologically examined using routine hematoxylin-eosin staining. For detection of different types of leukocytes the following markers were detected immunohistochemically: CD45 as a pan-leukocyte marker, CD3 for T-lymphocytes, CD68 for monocyte/ macrophages and mast cell tryptase for mast cells. Threestep immunoperoxidase detection was performed on paraffin sections after antigen retrieval and images were taken using Leica DMLB microscope equipped with DC300 camera. For double labeling we used immunofluorescence approach and images were takén using a confocal laser scanning microscope (FV1000, Olympus)

Results: In all atrial samples from both groups examined in the pilot study we detected CD45+-cells in atrial myocardium as well as in endocardium and epicardium. The frequency of these cells was variable. Cells were either isolated or grouped in small clusters. Apparently, CD45+ cells formed a heterogeneous group of cells. We also detected CD68+cells, CD3+cells and mast cells. A dual immunostaining showed CD45+/CD3+ and CD45+/CD68+ double positive cells. There were also CD45+ and CD68+ cells with elongated shape sending out long cellular processes, which may correspond to activated macrophages and/or dendritic cells. We performed a semiquantitative analysis of these cells and found that they are more frequent in auricular samples form patients with atrial fibrillation. The difference reached statistical significance in the right atrial samples.

Conclusion: Our results document that CD45+ cells are a heterogeneous cell population in atrial myocardium from patients undergoing open heart surgery and these cells can be detected regardless of the heart rhythm. The finding of higher frequency of CD45+ leukocytes with elongated processes in the AF samples suggests the activation of inflammatory cells in this arrhythmia.

This work was supported by the Research Program of Charles University – PRVOUK -P25/LF1/2.