Elsevier

Heart Rhythm

Volume 5, Issue 7, July 2008, Pages 1063-1067
Heart Rhythm

Original-experimental
Identification and distribution of interstitial Cajal cells in human pulmonary veins

https://doi.org/10.1016/j.hrthm.2008.03.057Get rights and content

Background

The major determinant of atrial fibrillation (AF) initiation is focal firing within the muscular portion of the pulmonary veins. We hypothesized that interstitial Cajal cells (ICCs), a known type of pacemaker cells, could underlie the pacemaking activity of isolated pulmonary veins.

Objective

The aim of the study was to characterize the presence and the distribution of ICCs in human pulmonary veins.

Methods

Immunohistochemistry was performed on a transversal section of each pulmonary vein of eight adult human hearts obtained at autopsy from January 2005 to December 2005. A history of AF was documented in two of these eight patients. Two immunostainings were performed on successive sections to differentiate ICCs from mast cells (antibody c-kit and antibody AA1). Morphological and distribution analyses were performed manually and automatically. Electron microscopy and immunostaining with HCN4 and smooth muscle alpha-actin antibodies were also used to further characterize Cajal cells.

Results

ICCs were found in the pulmonary vein sections of three of the eight patients and were mainly identified in sections with a thick muscular sleeve. Two of these three patients had a history of AF. The mean distribution density of these cells was 0.6 ICCs/3 mm2, with the highest density reaching 14.6 ICCs/3 mm2 in a pulmonary vein of a patient with a history of AF. A positive immunostaining of Cajal cells with HCN4 was also demonstrated.

Conclusions

ICCs may be detected in human pulmonary veins, particularly in patients with AF. Given the electrophysiological attributes of these cells, their role as AF triggers deserve to be more documented.

Introduction

Because the pathophysiology of atrial fibrillation (AF) is not fully understood, this common arrhythmia remains difficult to treat. Pulmonary veins (PVs) are considered to be the main trigger of AF initiation, but the precise identification of the substrate underlying ectopic firing within the muscular layers is unknown.1, 2 Independent pacemaker activity of the isolated PVs has been demonstrated in many mammalian species.3, 4 Numerous studies have shown that interstitial Cajal cells (ICC) are responsible for the pacemaker activity of the gastrointestinal (GI) muscles along the alimentary tract.5, 6 These cells have the unique property that they generate and propagate slow waves in GI muscles. They form a three-dimensional network between smooth muscle cells and nerves and are responsible for peristaltic activity in concert with the enteric nervous system.7 A decrease of ICC density has also been observed in many GI motility disorders.8, 9, 10, 11, 12 For these reasons, we hypothesized that ICCs are present in human PVs and that they may be the substrate of normal or abnormal electrical activity in the PVs of patients with AF.

The goal of this study was to identify ICCs and to analyze their distribution in human PVs.

Section snippets

Methods

Eight adult human hearts were obtained at autopsy from January 2005 to December 2005. Two of the patients had histories of AF. Samples of PVs and atrium were preserved in alcohol-formalin-acetic acid fixative solution. For seven patients, one transversal section of each PV was performed, perpendicular to the direction of blood flow. The segments were embedded in paraffin blocks and cut into 7-μm slides. These 28 samples (7 × 4PV) were used to analyze the locations and distributions of ICCs in

Immunohistochemistry

For each paraffin block, three serial sections were cut. One slide was stained with hematoxyline phloxine saffron and underwent immunostainings twice successively. To investigate the presence and distribution of ICCs, one section was stained for c-kit (polyclonal rabbit anti-human CD117/c-kit antibody, 1/200 dilution, A4502, Dako Cytomation). To differentiate mast cells from ICCs, a second immunostaining for human mast cell tryptase AA1 (primary mouse monoclonal anti-mast cell tryptase

Results

ICCs were found in PV sections in three of the eight patients. Two of these three patients had documented AF. Sections without muscular sleeves were generally poorer in c-kit-positive cells compared with sections of PV with a thick muscular sleeve. In the various sections of PVs, the proportion of mast cells was found to be higher than that of ICCs. Not all of the PV sections in the same patient contained ICCs. The mean density distribution of ICCs for these three patients was 2.6 ± 4.3 ICCs/3

Discussion

To our knowledge, this study is the first to demonstrate the presence of Cajal cells in human PVs. Cells that resemble ICCs have been described in ventricular and atrial myocardium of the heart.13, 14 The morphology of these cells with long processes is unique, and the demonstration that they react positively to HCN4 suggests that they may govern the ryhthmicity of the PVs.

ICCs had been identified in the rabbit portal vein by Povstyan et al.4 In that study, ICCs were located in the

Limitations

First, because there are c-kit-negative ICC-like cells that display spontaneous electrical activity,29 we have probably underestimated the number of ICCs in PVs. Second, we recognize that the presence of Cajal cells in two patients with AF does not prove that these cells are implicated in AF initiation.

Conclusion

This study has demonstrated, for the first time, the presence of interstitial cells of Cajal in human PVs. According to the known function of these types of cells, we could expect that they act as pacemaker cells in PVs. The abnormal density or function of these cells could be at the origin of abnormal firing focus, trigger AF, and fit into the “venous wave” hypothesis described by M. Haissaguerre et al.30 Additional studies are required to characterize the electrical properties of PV ICCs and

Acknowledgments

Tissues samples were provided by Cardiobiotec, biological resource center from Hospices Civils de Lyon.

The authors thank Simone Peyrol and Elisabeth Errazuriz from the Centre commun d'imagerie Laennec (CeCIL, Université Claude Bernard Lyon 1) and Yves Tourneur and Anne Beghin from the “Centre Commun de quantimétie” (Université Claude Bernard Lyon 1).

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    This work was supported by a grant from Biosense Webster, Inc.

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