Role of atrial lymphatics in atrial myocardial fibrosis and atrial fibrillation: a human and mouse study

Abstract

Abstract Background/Introduction Cardiac lymphatics maintain myocardial fluid and immune cell homeostasis, but its effects on atrial fibrosis and atrial fibrillation (AF) have not been clarified. Purpose To address this issue, we first evaluated the left atrial appendage (LAA) in humans histologically and biologically. We then studied mice to elucidate the mechanisms. Methods Human LAA samples were collected from 34 consecutive AF patients during cardiovascular surgery. They were assigned to the paroxysmal AF (PAF) group (n=17, 70.9±11.6 years) and the persistent AF (PeAF) group (n=17, 72.1±7.2 years). In the mouse experiments, 8-week-old male C57BL/6 mice were administrated subcutaneously with vehicle or Angiotensin II (AngII) (2.0 mg/kg/day) for 4 weeks. In some mice, VEGFC, a pro-lymphangiogenesis inducer, was administrated subcutaneously at a dose of 50 μg/kg/day simultaneously with AngII. AF was induced by transesophageal burst pacing in vivo, and by burst pacing in isolated perfused hearts using a Langendorff apparatus. Results In human LAA samples, mRNA expression of lymphangiogenesis-related genes (LYVE1, PROX1, and VEGFR3) was significantly lower in the PeAF group compared to the PAF group (p=0.037, 0.029, and 0.041). The number of LYVE1-positive atrial lymphatic endothelial cells (LECs) was also significantly lower in the PeAF group compared to the PAF group (p=0.026). There was a negative correlation between the number of LYVE1-positive atrial LECs and the area percentage of atrial myocardial fibrosis (r=-0.609, p<0.001). In mouse experiments, continuous infusion of AngII suppressed the mRNA expression of lymphangiogenesis-related genes, including Lyve1, Prox1, and Vegfr3 (n=6 in each group, p<0.001, p=0.003, and p<0.001, respectively). AngII infusion reduced the number of LYVE1-positive atrial LECs (p<0.001) and increased the mRNA expression of fibrosis-related genes (Tgfb, Col1a1, and Col3a1). The area percentage of fibrosis in atrial myocardium was also increased by AngII. Treatment with VEGFC for 4 weeks effectively reversed the effects of AngII, i.e., suppression of lymphangiogenesis-related genes expression, reduction of LYVE1-positive atrial LECs, increase in fibrosis-related genes expression, and increase in atrial fibrosis. Additionally, VEGFC treatment attenuated AngII-induced enhancement of vulnerability to AF in in vivo experiments (n=8 in each group, p=0.007) and in isolated perfused hearts (n=8 in each group, p=0.007). Conclusions Our study with human LAA demonstrated a strong association between atrial lymphatics and atrial myocardial fibrosis/progression of AF. Our mouse study showed that VEGFC reversed AngII -induced increase in atrial myocardial fibrosis and vulnerability to AF, possibly via promotion of lymphangiogenesis. Defects of lymphangiogenesis may be involved in atrial myocardial fibrosis and AF, and may be a therapeutic target for atrial cardiomyopathy.