Cardiac Lymphatics Research Articles

Cardiac lymphatic system

The lymphatic system is a network of vessels and lymphoid tissues that maintain tissue fluid homeostasis, transport intestinal fat, and regulate immune surveillance. Despite a large body of evidence showing the importance of lymphatic vessels in cardiovascular diseases, the role of cardiac lymphatics has not been extensively investigated. This review highlights the chronology of key discoveries in cardiac lymphatic development and function. In physiology, the cardiac lymphatic system dynamically regulates interstitial fluid drainage to the mediastinal lymph nodes to maintain homeostasis and prevent edema. After myocardial infarction, lymphatic vessels in the ischemic heart become dysfunctional and contribute to the development of chronic myocardial edema that aggravates cardiac fibrosis and dysfunction. Stimulation of cardiac lymphangiogenesis, based on the delivery of lymphangiogenic growth factors, such as VEGF-C, may represent a novel therapeutic strategy to improve cardiac function.

Normal Tissue Complications following Hypofractionated Chest Wall Radiotherapy in Breast Cancer Patients and Their Correlation with Patient, Tumor, and Treatment Characteristics.

Introduction:Normal tissue complications following chest wall radiotherapy (RT) are inevitable, and the long-term data on hypofractionation are still limited. To quantify the late effects of hypofractionated RT on cardiac, pulmonary, brachial plexus, and regional lymphatics and their correlation with patient, tumor, and treatment characteristics is the main objective of this study.Materials and Methods:Two hundred and sixteen breast cancer patients following mastectomy were treated with hypofractionated schedules either 40 Gy in 15 fractions or 42.5 Gy in 16 fractions. Common Toxicity Criteria version 3.0 was utilized to quantify the late effects of hypofractionation on cardiac, pulmonary, brachial plexus, and lymphedema at a maximum follow-up of 5 years.Results:Median follow-up was 42 months. Median age was 49 years. 14.8% developed ≥Grade (Gr) 2 late cardiac toxicity. 10.2% developed ≥Gr2 late pulmonary toxicity. There were 28.7% patients who developed ≥Gr2 lymphedema. Sixty-seven out of 216 patients had symptomatic brachial plexopathy at 5-year follow-up. Variables found to increase the incidence of these adverse events included smoking, hypertension, diabetes mellitus, body mass index ≥25, extent of axillary dissection, and use of supraclavicular field.Conclusion:Hypofractionation leads to increased risk of normal tissue complications partly influenced by some patient- and treatment-related factors, but these were manageable and minimally disabling.

Anatomy and development of the cardiac lymphatic vasculature: Its role in injury and disease.

Lymphatic vessels are present throughout the entire body in all mammals and function to regulate tissue fluid balance, lipid transport and survey the immune system. Despite the presence of an extensive lymphatic plexus within the heart, until recently the importance of the cardiac lymphatic vasculature and its origins were unknown. Several studies have described the basic anatomy of the developing cardiac lymphatic vasculature and more recently the detailed development of the murine cardiac lymphatics has been documented, with important insight into their cellular sources during embryogenesis. In this review we initially describe the development of systemic lymphatic vasculature, to provide the background for a comparative description of the spatiotemporal development of the cardiac lymphatic vessels, including detail of both canonical, typically venous, and noncanonical (hemogenic endothelium) cellular sources. Subsequently, we address the response of the cardiac lymphatic network to myocardial infarction (heart attack) and the therapeutic potential of targeting cardiac lymphangiogenesis.

Abstract 12624: Therapeutic Lymphangiogenesis Improves Diastolic Function by Limiting Cardiac Edema and Fibrosis After Myocardial Infarction

Little knowledge exists on how cardiac lymphatics respond to myocardial infarction (MI). We investigated the impact of MI on cardiac lymphatic structure and function during the development of chronic heart failure (CHF). We hypothesized that 1) insufficient cardiac lymph drainage may contribute to the deleterious cardiac remodeling post-MI; and 2) therapeutic stimulation of lymphatic growth, or lymphangiogenesis, may reduce chronic myocardial edema and inflammation in this setting. MI was modeled in Wistar rats by coronary occlusion (45 min) followed by reperfusion. Lymphangiogenesis-selective VEGF-C was delivered intramyocardially using slow-release albumin-alginate microparticles. Animals were divided into: sham (n=19), control MI (n=64), and treated MI, receiving low (1.5 μg; n= 54) or high (5 μg; n=26) dose of VEGF-C upon reperfusion. Cardiac function, lymphatics, edema, and levels of infiltrating immune cells were evaluated at 3 and 8 weeks post-MI. We found that control rats displayed lymphatic remodeling with increased lymphatic capillary density but rarefaction of epicardial pre-collector and collector vessels, leading to cardiac lymphatic dysfunction. In accordance with insufficient lymph drainage, chronic myocardial edema and low grade inflammation were found at both 3 and 8 weeks post-MI as determined by gravimetry, MRI T2 imaging, and immunohistochemistry, respectively. VEGF-C treatment led to a dose-dependent increase in lymphatic capillary density (sham, 255±16; con, 267±25; VEGF-C, 362±22 lymphatics/mm 2 ; p<0.05) and reduced rarefaction of pre-collectors and collector vessels. As a consequence, myocardial edema (sham, 75.5±0.1; con, 77.9±0.2; VEGF-C, 77.3±0.1% water content; p<0.05) and macrophage infiltration levels were both significantly reduced. Further, cardiac interstitial fibrosis was completely prevented by 8 weeks, even by low dose VEGF-C (con, 6.0±0.6; VEGF-C, 3.3±0.1% fibrosis; p<0.001), leading to reduced diastolic dysfunction (LVDPVR, LV end-diastolic pressure-volume relation: sham, 0.8±0.1; con, 2.0±0.2; VEGF-C, 1.1±0.1; p<0.001). In conclusion, our data suggest that therapeutic lymphangiogenesis may represent a new therapeutic target in CHF.

Lymphatic Vessels at the Heart of the Matter

Emergent research in the past decades has brought to light the importance of lymphatic vessels in tissue homeostasis, immunity, metabolism, and inflammation. In Nature, Klotz etal. (2015) demonstrate that cardiac lymphatics have a unique ontology compared to visceral lymphatics and that promoting their growth can improve cardiac function following injury.

Cardiac lymphatics are heterogeneous in origin and respond to injury.

The lymphatic vasculature is a blind-ended network crucial for tissue fluid homeostasis, immune surveillance and lipid absorption from the gut. Recent evidence has proposed an entirely venous-derived mammalian lymphatic system. In contrast, we reveal here that cardiac lymphatic vessels have a heterogeneous cellular origin, whereby formation of at least part of the cardiac lymphatic network is independent of sprouting from veins. Multiple cre-lox based lineage tracing revealed a potential contribution from the hemogenic endothelium during development and discrete lymphatic endothelial progenitor populations were confirmed by conditional knockout of Prox1 in Tie2+ and Vav1+ compartments. In the adult heart, myocardial infarction (MI) promoted a significant lymphangiogenic response, which was augmented by treatment with VEGF-C resulting in improved cardiac function. These data prompt the re-evaluation of a century-long debate on the origin of lymphatic vessels and suggest that lymphangiogenesis may represent a therapeutic target to promote cardiac repair following injury.

Cardiac Mouse Lymphatics: Developmental and Anatomical Update

The adult mouse heart possesses an extensive lymphatic plexus draining predominantly the subepicardium and the outer layer of the myocardial wall. However, the development of this plexus has not been entirely explored, partially because of the lack of suitable methods for its visualization as well as prolonged lymphatic vessel formation that starts prenatally and proceeds during postnatal stages. Also, neither the course nor location of collecting vessels draining lymph from the mouse heart have been precisely characterized. In this article, we report that murine cardiac lymphatic plexus development that is limited prenatally only to the subepicardial area, postnatally proceeds from the subepicardium toward the myocardial wall with the base-to-apex gradient; this plexus eventually reaches the outer half of the myocardium with a predominant location around branches of coronary arteries and veins. Based on multiple marker immunostaining, the molecular marker-phenotype of cardiac lymphatic endothelial cells can be characterized as: Prox-1(+), Lyve-1(+), VEGFR3(+), Podoplanin(+), VEGFR2(+), CD144(+), Tie2(+), CD31(+), vWF(-), CD34(-), CD133(-). There are two major collecting vessels: one draining the right and left ventricles along the left conal vein and running upwards to the left side of the pulmonary trunk and further to the nearest lymph nodes (under the aortic arch and near the trachea), and the other one with its major branch running along the left cardiac vein and further on the surface of the coronary sinus and the left atrium to paratracheal lymph nodes. The extracardiac collectors gain the smooth muscle cell layer during late postnatal stages.

Comparative and Developmental Anatomy of Cardiac Lymphatics

The role of the cardiac lymphatic system has been recently appreciated since lymphatic disturbances take part in various heart pathologies. This review presents the current knowledge about normal anatomy and structure of lymphatics and their prenatal development for a better understanding of the proper functioning of this system in relation to coronary circulation. Lymphatics of the heart consist of terminal capillaries of various diameters, capillary plexuses that drain continuously subendocardial, myocardial, and subepicardial areas, and draining (collecting) vessels that lead the lymph out of the heart. There are interspecies differences in the distribution of lymphatic capillaries, especially near the valves, as well as differences in the routes and number of draining vessels. In some species, subendocardial areas contain fewer lymphatic capillaries as compared to subepicardial parts of the heart. In all species there is at least one collector vessel draining lymph from the subepicardial plexuses and running along the anterior interventricular septum under the left auricle and further along the pulmonary trunk outside the heart and terminating in the right venous angle. The second collector assumes a different route in various species. In most mammalian species the collectors run along major branches of coronary arteries, have valves and a discontinuous layer of smooth muscle cells.

Cellular phenotypes and spatio-temporal patterns of lymphatic vessel development in embryonic mouse hearts

The origin of cardiac lymphatics from venous endothelial cells or from scattered lymphangioblasts has been discussed in the literature. We aimed to establish the stage when lymphatic vessels appear in the developing mouse heart, the location of the first lymphatics, and to define cellular phenotypes of growing lymphatics. We found that scattered Lyve-1-positive cells located in the subepicardial area of developing heart expressed CD45, CD68, F4/80, or CD11b but not CD31. Prox-1(+)/Lyve-1(+) cellular cords or vessels were found to invade 12.5-13.5-dpc hearts via two routes: from the venous pole, i.e., dorsal atrioventricular sulcus, or on the dorsal atrial surface from mediastinum and from the arterial pole, i.e., along the great arteries. The Prox-1(+)/Lyve-1(+) vessels were located among the Prox-1(+)/Lyve-1(-) cords and among the scattered Prox-1(-)/Lyve-1(+) cells. The Prox-1(+)/Lyve-1(-) cellular cords/tubules dominate initially at the arterial pole whereas Lyve-1(+)/Prox-1(-) cellular cords/tubules dominate initially on the venous pole, i.e., dorsal atrioventricular sulcus. The Lyve-1(+)/CD45(+), Lyve-1(+)/CD11b(+), Lyve-1(+)/F4/80(+) and Lyve-1(+)/CD68(+) cells were subsequently found to be co-opted to the wall of the developing lymphatic vessels while gaining Flk-1. Lymphatic primordia exhibit different cellular phenotypes and different spatiotemporal pattern on the venous pole as compared with the arterial pole of the heart.

The regulation of lymphangiogenic signaling in the epicardium

We have demonstrated that the cardiac lymphatics are derived in part from mesothelial cells in the embryonic epicardium, and that their development is impacted by VEGFR-3 inhibition. In this study, we explored the mechanisms that may affect lymphatic differentiation in pluripotent epicardial cells. Adult and embryonic epicardial cells were treated with the lymphangiogenic growth factor VEGF-C, resulting in a dramatic increase in nuclear expression for Prox-1, a master lymphatic regulatory gene, within 5 min of VEGF-C administration. The nuclear accumulation of Prox-1 was not observed after FGF or VEGF-A treatment. The ERK inhibitor UO126 induced a decrease in Prox-1 expression in nuclei in both untreated and VEGF-C treated cells, suggesting that activated ERK may regulate Prox-1 nuclear accumulation. Inhibiting ERK function in the epicardium also inhibited cell migration and differentiation. Our studies suggest that the lymphangiogenic growth factor VEGF-C binds to VEGFR-3 and acts via ERK to stimulate nuclear accumulation of Prox-1, in adult and embryonic epicardial cells. Preliminary findings suggest that tissue oxygen levels, mediated by the hypoxia inducible factor HIF-1, may also regulate lymphangiogenic signaling in the epicardium. Source of Research Support: R01 HL091171(ARRA), ES013507, AHA 0715153B. Grant Funding Source: NIH

The cardiac lymphatic system

The lymphatic system, a network of vessels carrying clear interstitial fluid called lymph, is found throughout the human body. The system maintains homeostasis, receiving proteins and excess fluid from the interstitial tissues, and returning them to the venous system. Understanding of lymphatic drainage remains important in the diagnosis, prognosis, and treatment of diseases, including the metastasis of malignant diseases. Information specific to the cardiac lymphatics is scarce. Indeed, quite often the topic is not even mentioned in many medical textbooks. The goal of our review is to compile and analyze the information currently available concerning the cardiac lymphatics, hoping further to demonstrate the clinical importance of this neglected system.

Ultrastructure Changes of Cardiac Lymphatics During Cardiac Fibrosis in Hypertensive Rats

Hypertension is one of the most common diseases that induce a series of pathological changes in different organs of the human body, especially in the heart. There is a wealth of evidence about blood vessels in hypertensive myocardium, but little is known about structural changes in the cardiac lymphatic system. To clarify the changes in structure of the cardiac lymphatic system during hypertension, we developed a hypertension animal model with Dahl S rats and we used Dahl R rats as the control group. We examined the expression of collagen fibers, atrial natriuretic peptide, connexin43, and LYVE-1 in the rat heart by immunohistochemistry. The ultrastructure of the cardiac lymphatics was detected by transmission electron microscopy and scanning electron microscopy. We demonstrated extensive lymphatic fibrosis in the hearts of the Dahl S hypertension group, characterized by increased thin collagen fibrils that connected with the lymphatics directly. Ultrastructural changes in the cardiac lymphatic endothelium such as an increase of vesicles and occurrence of vacuoles, active exocytosis, and cytoplasmic processes, restored the draining of tissue fluid. Our study suggests that during hypertension, the changes in structure of the cardiac lymphatics may be one important factor involved in cardiac fibrosis. Therefore, the lymphatics may be a possible target for reducing fibrosis in the treatment of hypertension.

Lymphangiogenesis in the heart

Our goal was to follow the development of the lymphatic system in the developing heart using markers that identify the lymphatics in adult tissues. The transcription factor Prox‐1 identified a branching network of vessels in the avian embryonic heart that first appeared on the outflow tract at Embryonic Day 5 (Hamburger and Hamilton Stage 25‐26). Prox‐1 was also expressed in cardiac myocytes and cardiac valves throughout development. In the mouse embryo heart, the lymphatic marker LYVE‐1 was expressed in epicardial cells and PECAM‐positive/VEGFR‐3‐positive myocardial vessels that were Prox‐1‐negative. LYVE‐1 co‐localization with Prox‐1 was not observed in the epicardial lymphatics until after ED 13.5. Lymphatic markers were also present in adult rat, embryonic mouse, and embryonic avian epicardial cell lines. According to Western Blot analysis, Prox‐1 has two isoforms in the embryonic heart, including one that is sensitive to phosphatases. Our data are consistent with the hypothesis that the cardiac lymphatics arise from at least two sources: they migrate into the embryonic heart and may also originate from epicardial precursors. Source of Research Support: AHA 0715153B and grants from the National Institutes of Health R01 ES013507, HL65314, HL0775436, HL083048, and NASA GRC IR&D 04‐54.

Aortic fat pad and atrial fibrillation: cardiac lymphatics revisited

The lymphatics of the heart have not generated any broad or sustained interest among clinicians. Few publications on cardiac lymphatics are available, the anatomy is not routinely known and the true role of cardiac lymphatics remains doubtful. One important anatomical concept needing clarification is that of the lymphatic drainage of conduction tissue. The sinoatrial node lymphatic collector and right principal lymphatic trunk are both incorporated into the aortic fat pad of the ascending aorta and are the most frequently damaged lymphatic vessels during cardiac surgery. Thus, preservation of the aortic fat pad and its lymphatic collectors should reduce the incidence of new atrial fibrillation observed in patients after cardiac surgery. This review assesses current knowledge of cardiac lymphatics and shows their possible role in triggering arrhythmias in the postoperative period.

CT06 AORTIC FAT PAD AND NEW ATRIAL FIBRILLATION POST CARDIAC SURGERY. CARDIAC LYMPHATICS REVISITED

Lymphatics of the heart have not generated any broad or sustained interests among clinicians. Few publications on cardiac lymphatics are available, anatomy not routinely known and the true role of cardiac lymphatics remains doubtful. One important anatomical concept needing clarification is that of the lymphatic nature of conduction tissue. Sino‐atrial node lymphatic collector is incorporated into Aortic Fat Pad and as such is the most frequently damaged lymphatic vessel during cardiac surgery. Thus, preservation of Aortic Fat Pad and its lymphatic collector should reduce the incidence of new atrial fibrillation in patients post cardiac surgery, an observation reported in the literature recently.

The proepicardium delivers hemangioblasts but not lymphangioblasts to the developing heart

The mass of the myocardium and endocardium of the vertebrate heart derive from the heart-forming fields of the lateral plate mesoderm. Further components of the mature heart such as the epicardium, cardiac interstitium and coronary blood vessels originate from a primarily extracardiac progenitor cell population: the proepicardium (PE). The coronary blood vessels are accompanied by lymph vessels, suggesting a common origin of the two vessel types. However, the origin of cardiac lymphatics has not been studied yet. We have grafted PE of HH-stage 17 (day 3) quail embryos hetero- and homotopically into chick embryos, which were re-incubated until day 15. Double staining with the quail endothelial cell (EC) marker QH1 and the lymphendothelial marker Prox1 shows that the PE of avian embryos delivers hemangioblasts but not lymphangioblasts. We have never observed quail ECs in lymphatics of the chick host. However, one exception was a large lymphatic trunk at the base of the chick heart, indicating a lympho-venous anastomosis and a ‘homing’ mechanism of venous ECs into the lymphatic trunk. Cardiac lymphatics grow from the base toward the apex of the heart. In murine embryos, we observed a basal to apical gradient of scattered Lyve-1 +/CD31 +/CD45 + cells in the subepicardium at embryonic day 12.5, indicating a contribution of immigrating lymphangioblasts to the cardiac lymphatic system. Our studies show that coronary blood and lymph vessels are derived from different sources, but grow in close association with each other.

Importance of the cardiac lymphatic system in open heart surgery

We read with great interest the article (August 2000) by Vasquez-Jimenez et al. [1]. They emphasized the cardiac lymph uid as the most direct medium for analyzing metabolical changes in the myocardial cell. The lymphatic drainage of the heart ows from subendocardial vessels to an extensive capillary plexus lying throughout the subepicardium [2]. The lymphatics begin in the tissues with blind-end lymphatic capillaries. These capillaries are very porous and easily collect large particles accompanied by interstitial uid [3]. Endothelial cells of the venous microcirculation and lymphatic vessels provide the exit sites and also work as conduits for some metabolic end products of myocytes and some hormonal substances. Endothelial cells of the lymphatic capillaries are anchored within the interstitium and to neighboring myocytes by a ®brillar collagen network. When the interstitial uid volume rises, it serves to exert a tension on these collagen ®bers and dilates lymphatic capillaries to foster the clearance of tissue uid. The extensive lymphatic circulation of the myocardium is responsible for returning tissue uid and plasma protein that gained access to the interstitium to the systemic and osmotic pressures and the uid volume of the extracellular space, while protecting against interstitial edema. Cardiac lymph also contains hormones that were released into the interstitium from secretory granules (atrial natriuretic peptide) and adrenergic neurons (norepinephrine) [4]. These substances can have important systemic effects. Another function of the cardiac lymph relates to the removal of cellular elements and tissue debris from the interstitium. This makes the lymphatic circulation an essential element in the myocardium's response to injury and wound healing. Because of these functions, the lymphatic system plays an important role for myocardial functions and cardiac lymph uid can be used as markers for the myocardial edema, reperfusion injury and myocardial damage. We have several concerns regarding the issue. We would like to ask the authors about the importance of lymphatic circulation on myocardial protection. As they know, we have to create external pressure on superior vena cava (SVC) (by using snare or vena cava clamp) for some open heart surgeries that require total cardiopulmonary bypass and total circulatory arrest (valve surgeries, congenital heart disease, arcus and ascending aorta surgery, etc.). We wonder, if the connection system and contractility might be affected with myocardial edema that caused by impaired lymphatic circulation, especially in the `right drainage types'. Do they advise using cuffed venous cannules to prevent this problem? We understand that the researchers were snared SVC, while preparing the swine heart. Did they ®nd any difference in ow rate on the `right drainage type' compare with the other types? Also, we want to learn if the impaired lymphatic circulation has any role on early graft failure of orthotopic and heterotopic heart transplantation and how the lymphatic system regenerates itself on these cases. The knowledge on cardiac lymphatics is very limited and their importance is almost always neglected in cardiac surgery practice. So, we congratulate the authors for such a good paper related the subject. We think, Dr VasquezJimenez and his colleagues deserve credits for their contributions on this unique subject.

Pharmacokinetics of calcium dobesilate in beagle dogs after a single administration.

A pharmacokinetic study was carried out in beagle dogs after a single intravenous infusion of 100 mg/kg of calcium dobesilate, a dose claimed to produce a cardiac lymphagogue effect. This effect on cardiac lymphatics is known to contribute to the reduction of myocardial infarct size after coronary artery occlusion. At the end of the intravenous infusion, which lasted about 30 minutes, the plasma level was 263 +/- 68 micrograms/ml, falling to 56 +/- 23 micrograms/ml by the third hour. This high plasma level of calcium dobesilate between the end of the infusion and the 3rd hour might explain the pharmacological effect of the drug on the cardiac lymphatic system.

Direct cannulation and injection lymphangiography of the canine cardiac and pulmonary efferent mediastinal lymphatics in experimental congestive heart failure.

Lymphangiograms were made in dogs with experimental congestive heart failure by cannulation of the left cardiac efferent (LCE), left pulmonary efferent (LPE), or cardiopulmonary lymphatic and injection of radiopaque medium. The lymphoangiograms showed cardiac enlargement and dilated mediastinal lymphatic channels consistent with an increase in lymph flow. By consecutive injections in the LCE and LPE, wer demonstrated that a common channel, the cardiopulmonary lymphatic, is opacified by injection in either the LPE or LCE. This common channel has been called the "cardiac lymphatic" and used for the collection of "cardiac" lymph. The authors' experiments suggest that this cardiopulmonary lymphatic carries both cardiac and pulmonary lymph.

Cardiac lymph and lymphatics in normal and infarcted myocardium

Cardiac lymphangiography was applied to 29 normal dogs (ND) and 14 dogs following acute coronary occlusions. In five of the ND, lymph collected from infarcted dogs was selectively injected into the coronary artery prior to the lymphangiogram. In ND, the lymphatic density (LD) was 6.6% ventricular area (VA). Acute coronary occlusion induced a change in the LD. The infarcted zone registered a decrease in LD (3.6% of VA p <0.001 vs ND). Hyaluronidase increased LD in normal dogs (13.8% of VA p < 0.01 vs ND). Hyaluronidase pretreatment increased the LD in infarcted myocardium (20.9% of VA, p < 0.001 vs non-treated infarcts). No difference was seen between uninfarcted areas of treated and nontreated animals. (10.6% vs 11.2% of VA). Lymph was collected from cardiac lymphatics in five dogs before and after coronary occlusion. When selectively injected into coronary arteries of ND, post-infarct lymph induced ECG changes of ischemia, arrhythmias, and increased the LD (14.6% VA, p < 0.001 vs ND). The results suggest (1) Cardiac lymphatics are altered in the uninfarcted and infarcted zones following coronary occlusion. (2) Hyaluronidase preserves the lymphatic density in infarction. (3) Lymph collected from zones of infarction is toxic.