The myocardium and its vasculature: a histochemical comparison of the normal and chronically sympathectomized dog heart

Summary Using histochemical techniques, the reactivities of selected enzymes and other metabolic components were examined in the myocardium, coronary arteries, and coronary arterioles of normal, two-week-sympathectomized, and sham-operated canine hearts. There were no differences in the histochemistry of coronary arteries in any of the hearts, but important differences were noted in the myocardium and especially in the arterioles. The reactivities of the enzyme glucose-6-phosphate dehydrogenase and the nucleic acids were increased in arterioles of the sympathectomized heart, possibly indicating an increased protein synthesis. The reactivities of succinate dehydrogenase, NAD-isocitrate dehydrogenase, and cytochrome oxidase were reduced in myocardium and arterioles of sympathectomized hearts as well as in arterioles of sham-operated hearts; the changes were greater in the sympathectomized arterioles where there was also observed an increase in reactivity of lactate dehydrogenase. These findings suggest a depression in aerobic metabolic capacity and, in the case of the sympathectomized arteriole, imply a possible shift in adaptation from aerobic to anaerobic metabolism.     

Investigation of the structure of human coronary vasculature

Some results of a morphometric study of the parameters of coronary arteries are presented. The parameters that characterize the structure of the arterial vasculature as an optimal branching system have been calculated. Statistically reliable correlations between the diameter of the bigger of two daughter vessels in a bifurcation with the diameter of the parent vessel as well as between the diameter of the smaller daughter vessel and the asymmetry coefficient have been obtained. Differences in the structural parameters of the two types of coronary arteries that provide blood delivery and distribution have been revealed. The relationships between the lengths and diameters of the arteries of different subsystems have been obtained. It is shown that asymmetrical branching is characteristic of the coronary vasculature, and self-similar asymmetric tree-like systems may be used for its modeling.     

Studies on the structure of human coronary vasculature

Based on morphometric data, we calculate the structural parameters of the coronary vasculature as an optimal branching bed. We show (i) significant correlations between the diameters of the larger daughter and the parent vessel and between the diameter of the smaller daughter vessel and the asymmetry coefficient; (ii) differences in the structural parameters for two types of artery that deliver and distribute blood in the cardiac muscle; and (iii) the length-diameter relationships for different arteries. The coronary vasculature is characterized by asymmetrical branching and thus should be modeled with self-similar asymmetrical tree-like systems.     

Bifurcation asymmetry of the porcine coronary vasculature and its implications on coronary flow heterogeneity

The branching pattern of the coronary arteries and veins is asymmetric, i.e., many small vessels branch off of a large trunk such that the two daughter vessels at a bifurcation are of unequal diameters and lengths. One important implication of the geometric vascular asymmetry is the dispersion of blood flow at a bifurcation, which leads to large spatial heterogeneity of myocardial blood flow. To document the asymmetric branching pattern of the coronary vessels, we computed an asymmetry ratio for the diameters and lengths of all vessels, defined as the ratio of the daughter diameters and lengths, respectively. Previous data from silicone elastomer cast of the entire coronary vasculature including arteries, arterioles, venules, and veins were analyzed. Data on smaller vessels were obtained from histological specimens by optical sectioning, whereas data on larger vessels were obtained from vascular casts. Asymmetry ratios for vascular areas, volumes, resistances, and flows of the various daughter vessels were computed from the asymmetry ratios of diameters and lengths for every order of mother vessel. The results show that the largest orders of arterial and venous vessels are most asymmetric and the degree of asymmetry decreases toward the smaller vessels. Furthermore, the diameter asymmetry at a bifurcation is significantly larger for the coronary veins (1.7-6.8 for sinus veins) than the corresponding arteries (1.5-5.8 for left anterior descending coronary artery) for orders 2-10, respectively. The reported diameter asymmetry at a bifurcation leads to significant heterogeneity of blood flow at a bifurcation. Hence, the present data quantify the dispersion of blood flow at a bifurcation and are essential for understanding flow heterogeneity in the coronary circulation.     

Exercise limits the production of endothelin in the coronary vasculature

We previously demonstrated that endothelin (ET)-mediated coronary vasoconstriction wanes with increasing exercise intensity via a nitric oxide- and prostacyclin-dependent mechanism (Ref. 23). Therefore, we hypothesized that the waning of ET coronary vasoconstriction during exercise is the result of decreased production of ET and/or decreased ET receptor sensitivity. We investigated coronary ET receptor sensitivity using intravenous infusion of ET and coronary ET production using intravenous infusion of the ET precursor Big ET, at rest and during continuous treadmill exercise at 3 km/h in 16 chronically instrumented swine. In the systemic vasculature, Big ET and ET induced similar changes in hemodynamic parameters at rest and during continuous exercise at 3 km/h, indicating that exercise does not alter ET production or receptor sensitivity in the systemic vasculature. In the coronary vasculature, infusion of ET resulted in similar dose-dependent decreases in coronary blood flow and coronary venous oxygen tension and saturation at rest and during exercise. In contrast, administration of Big ET resulted in dose-dependent decreases in coronary blood flow, as well as coronary venous oxygen tension and saturation at rest. These effects of Big ET were significantly reduced during exercise. Altogether, our data indicate that continuous exercise at 3 km/h attenuates ET-mediated coronary vasoconstriction through reduced production of ET from Big ET rather than through reduced ET sensitivity of the coronary vasculature. The decreased ET production during exercise likely contributes to metabolic coronary vasodilation.     

Mechanical properties of the coronary vasculature: indirect evaluation.

Information on the mechanical properties of the coronary vascular bed can be obtained indirectly by modelling the vascular system. This indirect approach, unlike 'in vitro' measurements, allows to take into account the vasomotor conditions of the circulatory district as well as the effect of the surrounding embedding tissue on the vascular performance. An experimental manoeuvre of sudden occlusion and subsequent release of the thoracic descendent aorta on 5 anaesthetized dogs with open pericardium induces a step-like variation in the coronary perfusion pressure and the occurrence of oscillations in the mean coronary flow. Such a behaviour can be described using a second-order model ('windkessel'+inductance, which takes into account blood inertia in the large vessels). The value of the coefficients entering the equations have been obtained with a 'best-fit' procedure (minimum of the chi-squared variable) on the haemodynamical data. Coefficient variations are in agreement with the direct estimation of the myocardial compliance and volume, measured by Ultrasound Echocardiographic imaging (4-chamber projection mode).     

The vasculature and limb development

The developing vascular pattern of the embryonic chick limb results from a combination of two properties: the intrinsic self-assembly and branching properties of the vascular cells and the extrinsic information associated with the expanding mitotic population of mesenchymal cells; and the inhibitory factors which restrict the entrance of vessels into particular domains and/or decrease the branching frequency of such vessels. It is hypothesized that an important component of limb pattern formation is the interplay between the dividing population of mesenchymal cells and the intrinsic properties of the vascular cells. It is further asserted that the presence of particular vascular elements may, indeed, be 'positional information'. Two examples are cited involving aspects of limb duplication to support this possibility; it is suggested that vascular vessel size of a host limb may dictate the polarity of duplication events. The presented hypothesis emphasizes that the interplay between the intrinsic properties of self-assembly into tissues and extrinsic factors which establish boundaries and morphologies is involved in both vascular and limb pattern formation.     

Muscle metaboreflex-induced increases in cardiac sympathetic activity vasoconstrict the coronary vasculature.

Ischemia of active skeletal muscle evokes a powerful blood pressure-raising reflex termed the muscle metaboreflex (MMR). MMR activation increases cardiac sympathetic nerve activity, which increases heart rate, ventricular contractility, and cardiac output (CO). However, despite the marked increase in ventricular work, no coronary vasodilation occurs. Using conscious, chronically instrumented dogs, we observed MMR-induced changes in arterial pressure, CO, left circumflex coronary blood flow (CBF), and coronary vascular conductance (CVC) before and after alpha1-receptor blockade (prazosin, 100 microg/kg iv). MMR was activated during mild treadmill exercise by partially reducing hindlimb blood flow. In control experiments, MMR activation caused a substantial pressor response-mediated via increases in CO. Although CBF increased (+28.1 +/- 3.7 ml/min; P < 0.05), CVC did not change (0.45 +/- 0.05 vs. 0.47 +/- 0.06 ml x min(-1) x mmHg(-1), exercise vs. exercise with MMR activation, respectively; P > 0.05). Thus all of the increase in CBF was due to the increase in arterial pressure. In contrast, after prazosin, MMR activation caused a greater increase in CBF (+55.9 +/- 17.1 ml/min; P < 0.05 vs. control) and CVC rose significantly (0.59 +/- 0.08 vs. 0.81 +/- 0.17 ml x min(-1) x mmHg(-1), exercise vs. exercise with MMR activation, respectively; P < 0.05). A greater increase in CO also occurred (+2.01 +/- 0.1 vs. +3.27 +/- 1.1 l/min, control vs. prazosin, respectively; P < 0.05). We conclude that the MMR-induced increases in sympathetic activity to the heart functionally restrain coronary vasodilation, which may limit increases in ventricular function.     

Ghrelin induces vasoconstriction in the rat coronary vasculature without altering cardiac peptide secretion

We administered ghrelin, a novel growth hormone-releasing hormone, to isolated perfused rat hearts, coronary arterioles, and cultured neonatal cardiomyocytes to determine its effects on coronary vascular tone, contractility, and natriuretic peptide secretion and gene expression. We also determined cardiac levels of ghrelin and whether the heart is a source of the circulating peptide. Ghrelin dose dependently increased coronary perfusion pressure (44 +/- 9%, P < 0.01), constricted isolated coronary arterioles (12 +/- 2%, P < 0.05), and significantly enhanced the pressure-induced myogenic tone of arterioles. These effects were blocked by diltiazem, an L-type Ca(2+) channel blocker, and bisindolylmaleimide (Bis), a protein kinase C (PKC) inhibitor. Interestingly, coinfusion of ghrelin with diltiazem completely restored myocardial contractile function that was decreased 30 +/- 3% (P < 0.01) by diltiazem alone. In contrast, combination of ghrelin with diltiazem or Bis did not significantly alter atrial natriuretic peptide (ANP) secretion, which was decreased 40% (P < 0.01) and 50% (P < 0.05) by these agents alone, respectively. Administration of ghrelin to cultured cardiomyocytes had no effect on ANP or B-type natriuretic peptide secretion or gene expression. Detectable amounts of low-molecular-weight ghrelin were present in cardiac tissue extracts but not in isolated heart perfusate. Thus we provide the first evidence that ghrelin has a coronary vasoconstrictor action that is dependent on Ca(2+) and PKC. Furthermore, the data obtained from diltiazem infusion suggest that ghrelin has a role in regulation of contractility when L-type Ca(2+) channels are blocked. Finally, the observation that immunoreactive ghrelin is found in cardiac tissue suggests the presence of a local cardiac ghrelin system.     

Atrial natriuretic factor in the Langendorff perfused coronary vasculature of the rabbit isolated heart

Removal of exogenously administered rat ANF (99-126) (rANF) from the rabbit coronary vasculature was investigated. Rabbit hearts were perfused using a modified Langendorff technique and ANF concentrations in the perfusate were measured by a radio-receptor assay. Under these conditions no major degradation of ANF was observed. On perfusion, however, the heart liberated large amounts of ANF. This release peaked 15 minutes after the initiation of perfusion, (685 + 220 pM) and then fell to a sustained basal level (305 + 80 pM) after 45 minutes. Although an increase in the perfusate flow rate reduced the ANF concentration, there was no significant difference in the rate of ANF release between the two flow rates used. After momentary cessation of flow ANF concentration fell to a significantly lower level, however, once again no significant change in rate of release occurred. These results suggest that the heart is not a major site of ANF degradation and that alterations in flow rate through the coronary vascular bed can cause changes in amounts of ANF released.     

Immunohistochemical localization of 6-keto-PGF1-alpha in canine coronary vasculature

The localization of the prostacyclin metabolite, 6-keto-PGF1-alpha, in canine coronary vasculature was accomplished using immunohistochemical techniques (avidin-biotin method of immunoperoxidase staining). Six-keto-PGF1-alpha was localized to the intimal endothelial cell layer of epicardial and intramyocardial arteries and veins. No specific staining was seen in the the media or adventitia of canine coronary vasculature, or in capillaries, or myocardial fibers. To our knowledge these studies represent the first immunohistochemical demonstration of the endothelial cell localization of the prostacyclin metabolite, 6-keto-PGF1-alpha. The described technique allows the cellular localization of prostaglandin metabolites in histologic sections.     

Protective effects of epoxyeicosatrienoic acids on human endothelial cells from the pulmonary and coronary vasculature

Epoxyeicosatrienoic acids (EETs) are cytochrome P-450 (CYP) metabolites synthesized from the essential fatty acid arachidonic acid to generate four regioisomers, 14,15-, 11,12-, 8,9-, and 5,6-EET. Cultured human coronary artery endothelial cells (HCAECs) contain endogenous EETs that are increased by stimulation with physiological agonists such as bradykinin. Because EETs are known to modulate a number of vascular functions, including angiogenesis, we tested each of the four regioisomers to characterize their effects on survival and apoptosis of HCAECs and cultured human lung microvascular endothelial cells (HLMVECs). A single application of physiologically relevant concentration of 14,15-, 11,12-, and 8,9-EET but not 5,6-EET (0.75-300 nM) promoted concentration-dependent increase in cell survival of HLMVECs and HCAECs after removal of serum. The lipids also protected the same cells from death via the intrinsic, as well as extrinsic, pathways of apoptosis. EETs did not increase intracellular calcium concentration ([Ca2+]i) or phosphorylate mitogen-activated protein kinase p44/42 when applied to these cells, and their protective action was attenuated by the phosphotidylinositol-3 kinase inhibitor wortmannin (10 microM) but not the cyclooxygenase inhibitor indomethacin (20 microM). Our results demonstrate for the first time the capacity of EETs to enhance human endothelial cell survival by inhibiting both the intrinsic, as well as extrinsic, pathways of apoptosis, an important underlying mechanism that may promote angiogenesis and endothelial survival during atherosclerosis and related cardiovascular ailments.     

MicroRNA-processing enzyme Dicer is required in epicardium for coronary vasculature development

The epicardium is a sheet of epithelial cells covering the heart during early cardiac development. In recent years, the epicardium has been identified as an important contributor to cardiovascular development, and epicardium-derived cells have the potential to differentiate into multiple cardiac cell lineages. Some epicardium-derived cells that undergo epithelial-to-mesenchymal transition and delaminate from the surface of the developing heart subsequently invade the myocardium and differentiate into vascular smooth muscle of the developing coronary vasculature. MicroRNAs (miRNAs) have been implicated broadly in tissue patterning and development, including in the heart, but a role in epicardium is unknown. To examine the role of miRNAs during epicardial development, we conditionally deleted the miRNA-processing enzyme Dicer in the proepicardium using Gata5-Cre mice. Epicardial Dicer mutant mice are born in expected Mendelian ratios but die immediately after birth with profound cardiac defects, including impaired coronary vessel development. We found that loss of Dicer leads to impaired epicardial epithelial-to-mesenchymal transition and a reduction in epicardial cell proliferation and differentiation into coronary smooth muscle cells. These results demonstrate a critical role for Dicer, and by implication miRNAs, in murine epicardial development.     

The vasculature of the peroneal tissue transfer

Peroneal vascularized composite-tissue transfer has many useful applications and advantages. An anatomic study of the peroneal artery and vein and their branches was carried out on 80 adult cadaver legs. The number of cutaneous branches averaged 4.8 +/- 1.4 per leg. The length of the cutaneous branches averaged 5.4 +/- 1.5 cm. The external diameters of cutaneous branches at the skin distribution site were 0.6 +/- 0.2 mm for the artery and 0.8 +/- 0.3 mm for the vein. The communicating branches were branched at anterior or posterior tibial vessels 6.1 +/- 2.4 cm proximal to the lateral malleolus. The range of rotation of the island flap when transposed proximally was 14.3 +/- 3.3 cm proximal from the head of the fibula, and when transposed distally, the range of rotation was 16.9 +/- 5.3 cm distally.     

Development of the adrenergic innervation in the myocardium and coronary arteries of the dog

The development of the adrenergic cardiac innervation was studied in premature dog fetuses, puppies and adult dogs by means of the formalin-induced fluorescence technique. A point-counting technique was used to evaluate the density of innervation. Two types of fluorescent profiles can be observed in the heart during development: (1) sprouting axons, and (2) beaded terminals. The axonal fluorescence disappears in adult neurons. A different morphology and a different time course of development enable to study separately the innervation of the myocardium (cardiomotor innervation) and that of the vessels (vasomotor innervation). The late prenatal innervation is very poor (0.1 hit). The first but very scant cardiomotor terminals appear in this period. A mature cardiomotor innervation is found in 4-month-old puppies [1.5 +/- 0.3 (SD) hits]. The vasomotor innervation is shifted to the right. The development of beaded vascular terminals begins and matures 1-2 weeks later. The growing fluorescent axons reveal that the myocardium is supplied by axons of the cardiac plexus and of the perivascular nerves; the vascular wall, on the other hand, is supplied by the perivascular nerves only. The developmental, spatial and morphological differences in innervation suggest that two different types of neurons exist in the sympathetic ganglia: (1) neurons innervating the vessels (coronaromotor neurons), and (2) neurons innervating the myocardium (cardiomotor neurons).     

Effect of passive myocardium on the compliance of porcine coronary arteries

The objective of this study was to determine the effect of passive myocardium on the coronary arteries under distension and compression. To simulate distension and compression, we placed a diastolic-arrested heart in a Lucite box, where both the intravascular pressure and external (box) pressure were varied independently and expressed as a pressure difference (DeltaP = intravascular pressure - box pressure). The DeltaP-cross-sectional area relationship of the first several generations of porcine coronary arteries and the DeltaP-volume relationship of the coronary arterial tree (vessels >0.5 mm in diameter) were determined using a video densitometric technique in the range of +150 to -150 mmHg. The vasodilated left anterior descending (LAD) coronary artery of six KCl-arrested hearts were perfused with iodine and 3% Cab-O-Sil. The intravascular pressure was varied in a triangular pattern, whereas the absolute cross-sectional area of each vessel and the total arterial volume were calculated using video densitometry under different box pressures (0, 50, 100, and 150 mmHg). In the range of positive DeltaP, we found that the compliance of the proximal LAD artery in situ (4.85 +/- 3.8 x 10-3 mm2/mmHg) is smaller than that of the same artery in vitro (16.5 +/- 6 x 10-3 mm2/mmHg; P = 0.009). Hence, the myocardium restricts the compliance of the epicardial artery under distension. In the negative DeltaP range, the LAD artery does not collapse, whereas the same vessel readily collapses when tested in vitro. Hence, we conclude that myocardial tethering prevents collapse of large blood vessel under compression.     

The lymphatic vasculature in disease

Blood vessels form a closed circulatory system, whereas lymphatic vessels form a one-way conduit for tissue fluid and leukocytes. In most vertebrates, the main function of lymphatic vessels is to collect excess protein-rich fluid that has extravasated from blood vessels and transport it back into the blood circulation. Lymphatic vessels have an important immune surveillance function, as they import various antigens and activated antigen-presenting cells into the lymph nodes and export immune effector cells and humoral response factors into the blood circulation. Defects in lymphatic function can lead to lymph accumulation in tissues, dampened immune responses, connective tissue and fat accumulation, and tissue swelling known as lymphedema. This review highlights the most recent developments in lymphatic biology and how the lymphatic system contributes to the pathogenesis of various diseases involving immune and inflammatory responses and its role in disseminating tumor cells.