Collagen volume fraction and microvascular density in human atrial wall during atrial fibrillation

Introduction: Atrial fibrillation (AF) is one of the most common arrhythmic diseases in the clinical practice and it is associated with an increase in mortality risk that is strongly related with old age. Its pathogenesis is still not sufficiently explored. One of the generally recognized factors contributing to the initiation and maintenance of atrial fibrillation is structural remodeling of the myocardium. Structural remodeling is reflected by changes that affect both atrial cardiomyocytes as well as endomysium.

Aim: In this project we focused on morphological and functional changes in endomysium of atrial myocardium with special focus on changes in the collagen volume fraction and immunohistochemical detection of pericytes surrounding myocardial capillaries.

Methods: We studied the morphological changes of atrial biopsies performed at 46 patients (19 patients with AF, and 27 with sinus rhythm (SR)) undergoing bypass or mitral valve surgery. The atrial samples were fixed with 4% paraformaldehyde and embedded into paraffin. Sections from atrium were histologically examined using routine hematoxylineosin staining. For quantification of collagen volume fraction (CVF), the sections were stained with the picrosirius staining. All morphometrical parameters were obtained using interactive image analysis software (LeicaQWin, Leica Microsystems). CVF was quantified as an area fraction of myocardial tissue section containing collagen fibers labeled with picrosirius staining. Only endomysial collagen fibers were quantified, while perimysial connective tissue was omitted.

Results: We found variable amount of endomysial collagen in myocardial samples from both groups of patients. Morphometrical calculation of CVF revealed the following results. The CVF in the left auricle was 23.78±5.1% in patients with SR and 28.44±6.7% in patients with AF. The CVF in the left atrium was 24.20±3.1% in patients with SR and 29.31±8.8% in patients with AF. The CVF in the right auricle was 27.40±6.1% in patients with SR and 27.16±4.45% in patients with AF. In all atrial samples the average CVF was 27.94±6.15% in patients with AF and 25.96±5.66% in patients with SR. We assessed the use of two pericyte markers – alpha-SMA and desmin for detection of pericytes surrounding endomysial capillaries. We found that desmin was strongly expressed in cardiomyocytes and we detected this intermediate filament also in vascular smooth muscle cells. No immunoreactivity was found in the capillaries. In contrast, alpha-SMA was constantly expressed by perivascular cells surrounding endomysial capillaries. These pericytes surrounded variable portion of the capillaries.

Conclusion: Our results document the variable level of fibrosis in atrial myocardium from patients undergoing open heart surgery. In the left atrial samples from patients with AF we observed a tendency for increased CVF. We verified the applicability of alpha-SMA as a pericyte marker in the atrial myocardium.

This work was supported by grants from Ministry of Health (MZO-00023001), by the EU Operational Program Prague - Competitiveness; project “CEVKOON” (CZ.2.16/3.1.00/22126), by the Grant Agency of the Academy of Sciences of the Czech Republic (305/09/1390), and by the Research program of Charles University.