Major gaps in human evidence for structure and function of the vasa vasora limit our understanding of the link with atherosclerosis.

Atherosclerosis is the major pathology causing death in the developed world and, although risk factor modification has improved outcomes over the last decade, there is no cure. The role of the vasa vasora (VV) in the pathogenesis of atherosclerotic plaque is unclear but must relate to the predictability of diseased sites in the arterial tree. VV are small vessels found on major arteries and veins which supply nutrients and oxygen to the vessel wall itself while removing waste. Numerous studies have been carried out to investigate the anatomy and function of the VV as well as their significance in vascular disease. There is convincing evidence that VV are related to atherosclerotic plaque progression and vessel thrombosis, however, their link to the pathology of plaque initiation remains an interesting but neglected topic. We aim to present the evidence on the anatomy and functional behaviour of VV as well as their relationship to the initiation of atherosclerosis. At the same time, we wish to highlight inconsistencies in, and limitations of, the evidence available.

Arterial smooth muscle dynamics in development and repair.

Arterial vasculature distributes blood from early embryonic development and provides a nutrient highway to maintain tissue viability. Atherosclerosis, peripheral artery diseases, stroke and aortic aneurysm represent the most frequent causes of death and are all directly related to abnormalities in the function of arteries. Vascular intervention techniques have been established for the treatment of all of these pathologies, yet arterial surgery can itself lead to biological changes in which uncontrolled arterial wall cell proliferation leads to restricted blood flow. In this review we describe the intricate cellular composition of arteries, demonstrating how a variety of distinct cell types in the vascular walls regulate the function of arteries. We provide an overview of the developmental origin of arteries and perivascular cells and focus on cellular dynamics in arterial repair. We summarize the current knowledge of the molecular signaling pathways that regulate vascular smooth muscle differentiation in the embryo and in arterial injury response. Our review aims to highlight the similarities as well as differences between cellular and molecular mechanisms that control arterial development and repair.

Glycolysis and oxidative phosphorylation are essential for purinergic receptor-mediated angiogenic responses in vasa vasorum endothelial cells.

Angiogenesis is an energy-demanding process; however, the role of cellular energy pathways and their regulation by extracellular stimuli, especially extracellular nucleotides, remain largely unexplored. Using metabolic inhibitors of glycolysis (2-deoxyglucose) and oxidative phosphorylation (OXPHOS) (oligomycin, rotenone, and FCCP), we demonstrate that glycolysis and OXPHOS are both essential for angiogenic responses of vasa vasorum endothelial cell (VVEC). Treatment with P2R agonists, ATP, and 2-methylthioadenosine diphosphate trisodium salt (MeSADP), but not P1 receptor agonist, adenosine, increased glycolytic activity in VVEC (measured by extracellular acidification rate and lactate production). Stimulation of glycolysis was accompanied by increased levels of phospho-phosphofructokinase B3, hexokinase (HK), and GLUT-1, but not lactate dehydrogenase. Moreover, extracellular ATP and MeSADP, and to a lesser extent adenosine, increased basal and maximal oxygen consumption rates in VVEC. These effects were potentiated when the cells were cultured in 20 mM galactose and 5 mM glucose compared with 25 mM glucose. Treatment with P2R agonists decreased phosphorylation of pyruvate dehydrogenase (PDH)-E1α and increased succinate dehydrogenase (SDH), cytochrome oxidase IV, and ß-subunit of F1F0 ATP synthase expression. In addition, P2R stimulation transiently elevated mitochondrial Ca2+ concentration, implying involvement of mitochondria in VVEC angiogenic activation. We also demonstrated a critical role of phosphatidylinositol 3-kinase and Akt pathways in lactate production, PDH-E1α phosphorylation, and the expression of HK, SDH, and GLUT-1 in ATP-stimulated VVEC. Together, our findings suggest that purinergic and metabolic regulation of VVEC energy pathways is essential for VV angiogenesis and may contribute to pathologic vascular remodeling in pulmonary hypertension.

Ascending aorta of hooded seals with particular emphasis on its vasa vasorum.

The pressure-volume relationship in the ascending aorta ("windkessel") of the hooded seal was determined and the morphology of its vasa vasorum described in some detail. We found that the ascending aorta has a high compliance and can easily accommodate the entire stroke volume when the peripheral vascular resistance becomes much increased and maintain perfusion pressure during the much extended diastole and thereby reduce cardiac stroke work during diving. We also found that the 3- to 5-mm thick wall of the ascending aorta had a very elaborate vasa vasorum interna with a hitherto undescribed vascular structure that penetrates the entire vascular wall. If similar structures with similar importance for the nutrition of the wall of the vessel are found in humans, important implications for the understanding of pathological conditions, such as aneurisms, may be indicated.

De la tortuosidad vascular fisiológica a la tortuositas vasorum retinae / From physiological vascular tortuosity to the tortuositas vasorum retinae

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Quantification of Adventitial Vasa Vasorum Vascularization in Double-injury Restenotic Arteries.

BACKGROUND: Accumulating evidence indicates a potential role of adventitial vasa vasorum (VV) dysfunction in the pathophysiology of restenosis. However, characterization of VV vascularization in restenotic arteries with primary lesions is still missing. In this study, we quantitatively evaluated the response of adventitial VV to vascular injury resulting from balloon angioplasty in diseased arteries. METHODS: Primary atherosclerotic-like lesions were induced by the placement of an absorbable thread surrounding the carotid artery of New Zealand rabbits. Four weeks following double-injury induced that was induced by secondary balloon dilation, three-dimensional patterns of adventitial VV were reconstructed; the number, density, and endothelial surface of VV were quantified using micro-computed tomography. Histology and immunohistochemistry were performed in order to examine the development of intimal hyperplasia. RESULTS: Results from our study suggest that double injured arteries have a greater number of VV, increased luminal surface, and an elevation in the intima/media ratio (I/M), along with an accumulation of macrophages and smooth muscle cells in the intima, as compared to sham or single injury arteries. I/M and the number of VV were positively correlated (R2 = 0.82, P < 0.001). CONCLUSIONS: Extensive adventitial VV neovascularization occurs in injured arteries after balloon angioplasty, which is associated with intimal hyperplasia. Quantitative assessment of adventitial VV response may provide insight into the basic biological process of postangioplasty restenosis.

Nerve growth factor stimulates regeneration of perivascular nerve, and induces the maturation of microvessels around the injured artery.

An immature vasa vasorum in the adventitia of arteries has been implicated in induction of the formation of unstable atherosclerotic plaques. Normalization/maturation of the vasa vasorum may be an attractive therapeutic approach for arteriosclerotic diseases. Nerve growth factor (NGF) is a pleotropic molecule with angiogenic activity in addition to neural growth effects. However, whether NGF affects the formation of microvessels in addition to innervation during pathological angiogenesis is unclear. In the present study, we show a new role for NGF in neovessels around injured arterial walls using a novel in vivo angiogenesis assay. The vasa vasorum around arterial walls was induced to grow using wire-mediated mouse femoral arterial injury. When collagen-coated tube (CCT) was placed beside the injured artery for 7-14 days, microvessels grew two-dimensionally in a thin layer on the CCT (CCT-membrane) in accordance with the development of the vasa vasorum. The perivascular nerve was found at not only arterioles but also capillaries in the CCT-membrane. Biodegradable hydrogels containing VEGF and NGF were applied around the injured artery/CCT. VEGF significantly increased the total length and instability of microvessels within the CCT-membrane. In contrast, NGF induced regeneration of the peripheral nerve around the microvessels and induced the maturation and stabilization of microvessels. In an ex vivo nerve-free angiogenesis assay, although NGF potentially stimulated vascular sprouting from aorta tissues, no effects of NGF on vascular maturation were observed. These data demonstrated that NGF had potent angiogenic effects on the microvessels around the injured artery, and especially induced the maturation/stabilization of microvessels in accordance with the regeneration of perivascular nerves.

The adventitia: essential regulator of vascular wall structure and function.

The vascular adventitia acts as a biological processing center for the retrieval, integration, storage, and release of key regulators of vessel wall function. It is the most complex compartment of the vessel wall and is composed of a variety of cells, including fibroblasts, immunomodulatory cells (dendritic cells and macrophages), progenitor cells, vasa vasorum endothelial cells and pericytes, and adrenergic nerves. In response to vascular stress or injury, resident adventitial cells are often the first to be activated and reprogrammed to influence the tone and structure of the vessel wall; to initiate and perpetuate chronic vascular inflammation; and to stimulate expansion of the vasa vasorum, which can act as a conduit for continued inflammatory and progenitor cell delivery to the vessel wall. This review presents the current evidence demonstrating that the adventitia acts as a key regulator of vascular wall function and structure from the outside in.

Recanalisation of the internal carotid artery via the vasa vasorum after coil occlusion.

Antegrade recanalisation of a completely occluded internal carotid artery (ICA) via the vasa vasorum is extremely rare. Here, we report such a case after proximal endovascular coiling in a case of dissected (i.e. non-atherosclerotic) ICA. A 42-year-old man presented with thromboembolic stroke of the left frontal lobe owing to pseudo-occlusion of the left ICA manifesting as motor aphasia and right hemiparesis. There were abundant floating thrombi in the petrous portion of the left ICA. Because of good collateral flow in the left middle cerebral artery territory through the anterior communicating artery and external carotid artery system, endovascular coil embolisation of the left ICA was performed for prevention of further thromboembolic stroke. The patient showed progressive recovery following endovascular treatment, and was discharged with mild right hemiparesis 1 month later. He maintained a regimen of aspirin and physical rehabilitation. At follow-up, 38 months later, the patient was asymptomatic. Angiography demonstrated occlusion of the left ICA and multiple serpiginous vessels originating from the proximal internal and external carotid arteries and which filled the ICA distal to the occlusion. This case suggests that an ICA occluded by proximal coil embolisation-even in a non-atherosclerotic case-might be recanalised via the vasa vasorum.

P2Y1 and P2Y13 purinergic receptors mediate Ca2+ signaling and proliferative responses in pulmonary artery vasa vasorum endothelial cells.

Extracellular ATP and ADP have been shown to exhibit potent angiogenic effects on pulmonary artery adventitial vasa vasorum endothelial cells (VVEC). However, the molecular signaling mechanisms of extracellular nucleotide-mediated angiogenesis remain not fully elucidated. Since elevation of intracellular Ca(2+) concentration ([Ca(2+)](i)) is required for cell proliferation and occurs in response to extracellular nucleotides, this study was undertaken to delineate the purinergic receptor subtypes involved in Ca(2+) signaling and extracellular nucleotide-mediated mitogenic responses in VVEC. Our data indicate that stimulation of VVEC with extracellular ATP resulted in the elevation of [Ca(2+)](i) via Ca(2+) influx through plasma membrane channels as well as Ca(2+) mobilization from intracellular stores. Moreover, extracellular ATP induced simultaneous Ca(2+) responses in both cytosolic and nuclear compartments. An increase in [Ca(2+)](i) was observed in response to a wide range of purinergic receptor agonists, including ATP, ADP, ATPγS, ADPßS, UTP, UDP, 2-methylthio-ATP (MeSATP), 2-methylthio-ADP (MeSADP), and BzATP, but not adenosine, AMP, diadenosine tetraphosphate, αßMeATP, and ßγMeATP. Using RT-PCR, we identified mRNA for the P2Y1, P2Y2, P2Y4, P2Y13, P2Y14, P2X2, P2X5, P2X7, A1, A2b, and A3 purinergic receptors in VVEC. Preincubation of VVEC with the P2Y1 selective antagonist MRS2179 and the P2Y13 selective antagonist MRS2211, as well as with pertussis toxin, attenuated at varying degrees agonist-induced intracellular Ca(2+) responses and activation of ERK1/2, Akt, and S6 ribosomal protein, indicating that P2Y1 and P2Y13 receptors play a major role in VVEC growth responses. Considering the broad physiological implications of purinergic signaling in the regulation of angiogenesis and vascular homeostasis, our findings suggest that P2Y1 and P2Y13 receptors may represent novel and specific targets for treatment of pathological vascular remodeling involving vasa vasorum expansion.

Molecular imaging of inflammation and intraplaque vasa vasorum: a step forward to identification of vulnerable plaques?

Current developments in cardiovascular biology and imaging enable the noninvasive molecular evaluation of atherosclerotic vascular disease. Intraplaque neovascularization sprouting from the adventitial vasa vasorum has been identified as an independent predictor of intraplaque hemorrhage and plaque rupture. These intraplaque vasa vasorum result from angiogenesis, most likely under influence of hypoxic and inflammatory stimuli. Several molecular imaging techniques are currently available. Most experience has been obtained with molecular imaging using positron emission tomography and single photon emission computed tomography. Recently, the development of targeted contrast agents has allowed molecular imaging with magnetic resonance imaging, ultrasound and computed tomography. The present review discusses the use of these molecular imaging techniques to identify inflammation and intraplaque vasa vasorum to identify vulnerable atherosclerotic plaques at risk of rupture and thrombosis. The available literature on molecular imaging techniques and molecular targets associated with inflammation and angiogenesis is discussed, and the clinical applications of molecular cardiovascular imaging and the use of molecular techniques for local drug delivery are addressed.

Niacin in the prevention of atherosclerosis: Significance of vasodilatation.

There is a rising interest towards the old drug, nicotinic acid (niacin, vitamin B(3)), because at pharmacological concentrations it has a beneficial effect on HDL cholesterol. Its use, however, was limited due to its adverse effect, flushing. When the mechanism of flushing was solved, a combination of niacin and DP1 receptor antagonist or prostaglandin inhibitor is used, there has been a comeback of niacin with extensive clinical trials. This paper argues that the new strategy with niacin for the prevention of atherosclerosis should be re-evaluated, because vasodilatation of the peripheral vessels might be crucially important in the early primary prevention according to our "vasa vasorum hypoxia" hypothesis.

Endothelial cell-selective adhesion molecule modulates atherosclerosis through plaque angiogenesis and monocyte-endothelial interaction.

Endothelial cell-selective adhesion molecule (ESAM) is a new member of the immunoglobulin superfamily, which is expressed in vascular endothelial cells. Previous studies have demonstrated that ESAM regulates angiogenesis, endothelial permeability, and leukocyte transmigration. However, little is known concerning the role of ESAM in atherosclerosis. In this study, we assessed the effects of ESAM inactivation on atherosclerosis in mice. ESAM-/- mice were bred with apoE-/- mice to generate double knockout mice, and the aortic lesion size of apoE-/- and ESAM-/-apoE-/- mice was compared histologically. Although plasma cholesterol levels were higher in ESAM-/-apoE-/- mice, the lesion size was markedly smaller than in apoE-/- mice. ESAM-/-apoE-/- mice exhibited a decrease in the number of vasa vasorum and macrophages in the vessel wall. In vitro adhesion assays showed that THP-1 cells, which did not express ESAM, bound to the ESAM-coated culture plates, suggesting that ESAM may interact with heterophilic ligand(s) on monocytes. Moreover, downregulation of ESAM by siRNA in the endothelial monolayer diminished transendothelial migration of THP-1 cells. In conclusion, ESAM inactivation can reduce susceptibility to atherosclerosis by inhibiting plaque neovascularization and macrophage infiltration into the atheroma.

The absence of lymphatics in normal and atherosclerotic coronary arteries in man: a morphologic study.

It has been suggested by various investigators that the impairment of lymphatic drainage from the coronary arteries may play a role in predisposition to coronary atherosclerosis, the pathogenesis of which is certainly multifactorial. In our study, no lymphatic vessels were found in the walls of the coronary arteries (adventitia, media and intima) in 51 human hearts from patients ranging in ages from 3 months to 83 years with normal coronary arteries, coronary atherosclerosis, and cardiomyopathy. Visualized lymphatics were located solely in the periadventitial area, and these lymphatics were more irregular in hearts from older persons. With injection, histology, and electronmicroscopy methods we could not detect penetration of lymphatics into the wall of coronary trunks in normal as well atherosclerotic arteries. In all coronary arteries studied, and particularly in the atherosclerotic lesions, blood vasa vasorum could be visualized. In the atherosclerotic areas, vasa vasorum (angiogenesis) could be seen penetrating into the media and intima. Many of the thin-walled vasa vasorum could easily be mistaken for lymphatics. The absence of lymphatics draining the epicardial coronary arteries may be a predisposing factor to coronary atherosclerosis.

Descending vasa recta endothelium is an electrical syncytium.

We examined gap junction coupling of descending vasa recta (DVR). DVR endothelial cells or pericytes were depolarized to record the associated capacitance transients. Virtually all endothelia and some pericytes exhibited prolonged transients lasting 10-30 ms. Carbenoxolone (100 microM) and 18beta-glycyrrhetinic acid (18betaGRA; 100 microM) markedly shortened the endothelial transients. Carbenoxolone and heptanol (2 mM) reduced the pericyte capacitance transients when they were prolonged. Lucifer yellow (LY; 2 mM) was dialyzed into the cytoplasm of endothelial cells and pericytes. LY spread diffusely along the endothelial monolayer, whereas in most pericytes, it was confined to a single cell. In some pericytes, complex patterns of LY spreading were observed. DVR cells were depolarized by voltage clamp as fluorescence of bis(1,3-dibarbituric acid)-trimethine oxanol [DiBAC(4)(3)] was monitored approximately 200 microm away. A 40-mV endothelial depolarization was accompanied by a 26.1 +/- 5.5-mV change in DiBAC(4)(3) fluorescence. DiBAC(4)(3) fluorescence did not change after 18betaGRA or when pericytes were depolarized. Similarly, propagated cytoplasmic Ca(2+) responses arising from mechanical perturbation of the DVR wall were attenuated by 18betaGRA or heptanol. Connexin (Cx) immunostaining showed predominant linear Cx40 and Cx43 in endothelia, whereas Cx37 stained smooth muscle actin-positive pericytes. We conclude that the DVR endothelium is an electrical syncytium and that gap junction coupling in DVR pericytes exists but is less pronounced.

Impact of coronary vasa vasorum functional structure on coronary vessel wall perfusion distribution.

Noncoronary vasa vasorum have been described as networks of microvessels in the wall of arteries and veins. However, we have shown, using microcomputerized tomography (micro-CT) imaging methods, that porcine coronary vasa vasorum have a tree-like branching structure similar to the vasculature in general. In this study, we elucidate functional aspects of coronary vasa vasorum perfusion territories. Three pig hearts were injected with radiopaque Microfil via the coronary sinus to fill the left anterior descending coronary arteries (LADs) retrogradely at atmospheric pressure. In three other hearts, LADs were injected antegradely at 100-mmHg pressure via the left main carotid artery. Additionally, six LADs were injected in vivo with a suspension of 100- or 300-microm-diameter microspheres before harvesting of the hearts and injection of the LADs with Microfil. All harvested LADs were scanned intact with micro-CT (20 microm cubic voxels). The spatial density of vasa vasorum (no. of vasa/mm2) was measured in 20-microm-thick cross sections (at 0.4-mm intervals). Retrogradely injected LADs showed high and uniformly distributed vasa vasorum densities in the adventitia (means +/- SE; 5.38 +/- 0.09 vs. 3.58 +/- 0.1 vasa/mm2 in antegradely prepared LADs; P < 0.001). Antegradely prepared LADs showed patchy distributed, low-vasa-vasorum-density territories especially on the myocardial side of the coronary artery wall (epicardial density: 4.29 +/- 0.13 vasa/mm2 vs. myocardial density: 2.80 +/- 0.1 vasa/mm2, P < 0.001). Microembolization reduced vasa vasorum densities significantly (100-mum-diameter microspheres: 3.26 +/- 0.07 vasa/mm2, P < 0.05; 300-microm-diameter microspheres: 2.66 +/- 0.07 vasa/mm2, P < 0.001 vs. antegrade controls) and increased the size of low-vasa-vasorum-density territories. We conclude that coronary vasa vasorum are functional endarteries not connected via a plexus. This characteristic may have a significant impact on the spatial distribution of perfusion and drainage of the coronary vessel wall.