Increased Lymphangiogenesis and Lymphangiogenic Growth Factor Expression in Perivascular Adipose Tissue of Patients with Coronary Artery Disease.

Drosos Drosos,Carrillo Carrillo,Vahl Vahl,Tziakas Tziakas,Schäfer Schäfer,Gogiraju Gogiraju,Chalikias Chalikias,Konstantinou Konstantinou,Pavlaki Pavlaki,Tortopidis Tortopidis,Bochenek Bochenek,Buschmann Buschmann,Kourkouli Kourkouli,Konstantinides Konstantinides,Mikroulis Mikroulis,Münzel Münzel

doi:10.3390/jcm8071000

Abstract

Experimental and human autopsy studies have associated adventitial lymphangiogenesis with atherosclerosis. An analysis of perivascular lymphangiogenesis in patients with coronary artery disease is lacking. Here, we examined lymphangiogenesis and its potential regulators in perivascular adipose tissue (PVAT) surrounding the heart (C-PVAT) and compared it with PVAT of the internal mammary artery (IMA-PVAT). Forty-six patients undergoing coronary artery bypass graft surgery were included. Perioperatively collected C-PVAT and IMA-PVAT were analyzed using histology, immunohistochemistry, real time PCR, and PVAT-conditioned medium using cytokine arrays. C-PVAT exhibited increased PECAM-1 (platelet endothelial cell adhesion molecule 1)-positive vessel density. The number of lymphatic vessels expressing lymphatic vessel endothelial hyaluronan receptor-1 or podoplanin was also elevated in C-PVAT and associated with higher inflammatory cell numbers, increased intercellular adhesion molecule 1 (ICAM1) expression, and fibrosis. Significantly higher expression of regulators of lymphangiogenesis such as vascular endothelial growth factor (VEGF)-C, VEGF-D, and VEGF receptor-3 was observed in C-PVAT compared to IMA-PVAT. Cytokine arrays identified angiopoietin-2 as more highly expressed in C-PVAT vs. IMA-PVAT. Findings were confirmed histologically and at the mRNA level. Stimulation of human lymphatic endothelial cells with recombinant angiopoietin-2 in combination with VEGF-C enhanced sprout formation. Our study shows that PVAT surrounding atherosclerotic arteries exhibits more extensive lymphangiogenesis, inflammation, and fibrosis compared to PVAT surrounding a non-diseased vessel, possibly due to local angiopoietin-2, VEGF-C, and VEGF-D overexpression.

Highlights

Accumulating clinical and experimental data suggest an active contribution of the adventitia to remodeling processes in the vascular wall, including neointima formation and atherosclerosis.For example, preclinical studies in hypercholesterolemic animal models have shown that adventitial angiogenesis promotes atherosclerotic plaque progression, whereas inhibition of plaque neovascularization reduced lesion growth [1,2,3]
These analyses revealed that both the number of lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1) (24.7 (0.61–32.3) vs. 2.97 (0–6.58) per mm2, p = 0.027; Figure 1B,E; Figure S1B) and PDPN (5.45 ± 1.54 vs. 0.74 ± 0.34 per mm2, p = 0.023; Figure 1C,F; Figure S1C) -immunopositive vessels was significantly increased in C-perivascular adipose tissue (PVAT) compared to internal mammary artery (IMA)-PVAT
To examine whether the more extensive lymphatic network in C-PVAT is accompanied by a higher expression of growth factors known to mediate angiogenesis, quantitative real-time PCR (qPCR) analysis was employed to determine gene expression levels of vascular endothelial growth factor (VEGF)-A, VEGF-B, VEGF-C, and VEGF-D in C-PVAT and IMA-PVAT. These analyses revealed no significant differences in the mRNA levels of VEGF-A (0.85 (0.68–1.21) vs. 0.72 (0.41–1.02), p = 0.421; Figure 2A), whereas the expression of VEGF-C (0.83 (0.55–1.25) vs. 0.36 (0.22–0.72), p = 0.022; Figure 2C) and VEGF-D (0.58 (0.42–0.87) vs. 0.25 (0.21–0.31), p = 0.007; Figure 2D) was significantly increased in C-PVAT compared to IMA-PVAT

Summary

Introduction

Accumulating clinical and experimental data suggest an active contribution of the adventitia to remodeling processes in the vascular wall, including neointima formation and atherosclerosis.For example, preclinical studies in hypercholesterolemic animal models have shown that adventitial angiogenesis promotes atherosclerotic plaque progression, whereas inhibition of plaque neovascularization reduced lesion growth [1,2,3]. The expansion of thin-walled, leaky vasa vasorum and microvessels within atherosclerotic lesions has been pathophysiologically linked to plaque instability by increasing inflammatory cell infiltration and intraplaque hemorrhage [4,5,6], among others. Lymph vessels control the drainage of interstitial fluids and macromolecules (proteins, lipids) leaking from capillaries back to the venous circulation and are critical for the maintenance of tissue homeostasis [8]. They have an important function in immune surveillance and may accelerate the resolution of inflammation by removing inflammatory mediators or trafficking immune cells to draining lymph nodes. Lymph vessel dysfunction results in lymphedema, and may play a role during inflammation, cancer, obesity, or hypertension (reviewed in [9])

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Conclusion