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Evidence for a role of Smad6 in chick cardiac development.

Bone morphogenetic proteins (BMPs), members of the transforming growth factor-beta (TGF-beta) superfamily, are obligatory growth factors for early embryogenesis and heart formation. SMAD proteins transduce signals of the TGF-beta superfamily. We isolated chicken Smad6 (cSmad6), a member of inhibitory SMADs, and found its expression to be remarkably restricted to the developing heart, eyes, and limbs. cSmad6 expression was detected in the cardiogenic region of stage 5 embryos and overlapped Nkx2-5 and bmp-2, -4, and -7 expression. Throughout development, cSmad6 was expressed strongly in the heart, primarily in the myocardium, endocardium, and endocardial cushion tissue. Myocardial expression of cSmad6 was stronger in the forming septum, where highly localized expression of bmp-2 and -4 was also observed. Ectopically applied BMP-2 protein induced the expression of cSmad6, a putative negative regulator of BMP-signaling pathway, in anterior medial mesoendoderm of stage 4-5 embryos. In addition, blocking of BMP signaling using Noggin downregulated cSmad6 in cardiogenic tissue. cSmad1, one of the positive mediators of BMP signaling, was also expressed in cardiogenic region, but was not BMP-2 inducible. Our data suggest that cSmad6 has a role in orchestrating BMP-mediated cardiac development. We propose the possible mechanism of action of cSmad6 as modulating BMP signal by keeping a balance between constitutively expressed pathway-specific cSmad1 and ligand-induced inhibitory cSmad6 in the developing heart.

Amino Acid Sequence↗

Angiotensin II binding sites in the heart of Scyliorhinus canicula: an autoradiographic study.

Dogfish (125)I [Asn(1), Pro(3), Ile(5)] angiotensin II ((125)I dfANG II) was used to establish the specific binding patterns of the different cardiac regions of the elasmobranch Scyliorhinus canicula by in vitro autoradiography. In the ventricular myocardium Scatchard analysis of saturation and displacement binding data revealed two classes of high- and low-affinity dfANG II binding sites (K(d) = 53 +/- 10 and 1300 +/- 900 pM). Two classes of dfANG II binding sites were also detected in the atrium (K(d) = 47 +/- 13 and 4690 +/- 930 pM) and in the outer layer of the conus arteriosus (K(d) = 16 +/- 9 and 398 +/- 83 pM). Conversely, the ventricular endocardium and the inner conal layer were characterized by a single class of dfANG II binding sites with affinity values of 48 +/- 11 and 106 +/- 3.3 pM, respectively. Competition experiments with either cold dfANG II or CV11974 or CGP42112 (specific ligands for mammalian AT(1) and AT(2) receptors, respectively) demonstrated a prevalence of CGP42112-selective dfANG II binding sites in both the inner and the outer conal layers. In the atrium, the ventricular myocardium, and the outer conal layer, dfANG II high-affinity binding sites poorly discriminated among the cold ligands. These results suggest that the dogfish heart may be a target organ of ANG II with distinct ANG II receptor subtype distributions.

Angiotensin II↗

Different onset of nucleolar activation in endocardial endothelial cells and cardiomyocytes following pressure overload in rat heart.

We evaluated the time course of activation of endocardial endothelial cells and cardiomyocytes in adult rats with pressure overload induced by the abdominal aortic constriction. The silver staining technique for nucleolar organizer region was used to mark an increased transcriptional activity of the cells. An increased number of nucleoli was already detected in the endocardial endothelium of the left ventricle 5 h post-operatively, while the response of cardiomyocytes was still absent. Twenty-four h after constriction, the activation of the endocardial endothelium and cardiomyocytes was evident in both ventricles. A similar delay was observed between the nucleolar activation in vascular endothelium and smooth muscle cells of the abdominal aorta. In contrast, these cells of the thoracic aorta did not exhibit any significant increase of the transcriptional activity. The sequential stimulation of the transcriptional activity of the left ventricular endocardial endothelial cells and the subjacent cardiac myocytes due to pressure overload is in accord with the view that the endocardium may serve as a mechanotransducer which converts the hemodynamic changes into the signals that influence the growth of the cardiac myocytes.

Animals↗

Isolation and culture of capillary endothelial cells from the eel, Anguilla rostrata.

There has been little development of endothelial cell cultures from vertebrates other than mammals. In this report the isolation of capillary endothelial cells from the rete mirabile of the eel, Anguilla rostrata, is described. The cells are isolated with collagenase H and dispase II. The cells are plated into fibronectin-hyaluronic acid coated flasks. The culture medium is M199 with Earle's salts supplemented with NaCl, HEPES, NaHCO(3), glutamine, pyruvate, heparin, antibiotics, endothelial cell growth supplement, and 20% serum. Cultures are incubated at 25 degrees C in humidified air. The rete mirabile contains pericytes in addition to endothelial cells. Variations in plating time, serum concentrations, and growth matrices were tried to separate the two cell types. The total number of endothelial cells and the ratio of endothelial cells to pericytes are the most important factors in obtaining pure cultures of capillary endothelial cells. Endothelial cells are isolated also from the endocardium, bulbus arteriosus, and large vessels. The initial isolates usually take 3-6 weeks to grow to confluence with subcultures taking about 2 weeks to confluence.

Air Sacs↗

Changes in diastolic function and collagen content in normotensive and hypertensive rats with long-term streptozotocin-induced diabetes.

We investigated the relationship between left ventricular diastolic function and interstitial collagen content in the endocardium, mesocardium and epicardium of transverse sections of the heart, using an image analysis system in normotensive and hypertensive long-term streptozotocin (STZ) diabetic rats. STZ-induced diabetes was characterised by elevated blood glucose, polyuria, polydypsia and loss of body weight. In vivo systolic blood pressure was 165 +/- 4, 136 +/- 3 and 129 +/- 7 mmHg in hypertensive and normotensive diabetic rats and age-matched controls, respectively. Heart rate was significantly lower (P < 0.01) in diabetic rats (283 +/- 8 and 280 +/- 10 beats min-1 in normotensive and hypertensive rats, respectively) than controls (393 +/- 18 beats min-1). Pressure-volume (P-V) curves were studied in isolated Langendorff perfused hearts at rest and after 20 min global ischaemia and 30 min reperfusion 6 months after induction of diabetes. Left ventricular volumes were significantly smaller in diabetic rats than age-matched controls, but volumes normalised for heart weight were higher in normotensive (by 28%) and hypertensive (by 10%) diabetic rats. Slopes of end-diastolic P-V curves were similar between groups in basal conditions, but left ventricular systolic P-V curves were steeper in normotensive and flatter in hypertensive diabetic hearts. Post-ischaemic left ventricular end-diastolic pressure was significantly higher than the pre-ischaemic value at comparable increments of volume in each group. Collagen content significantly increased in the heart of rats with STZ-diabetes both in the free left ventricular wall and septum, and suggested this may play a role in the cardiac defects in contractility and relaxation in our experimental conditions. These results indicate that diabetes, irrespective of associated hypertension, can cause major changes in cardiac performance and susceptibility to ischaemia and reperfusion.

Animals↗

Electrophysiologic and hemodynamic effects of apomorphine in dogs.

Apomorphine is a dopamine receptor agonist used as an emetic, for Parkinson's disease, and for treating erectile dysfunction. This study was conducted to monitor cardiovascular function in dogs given the standard emetic dose (0.05 mg/kg) or 10 times that. Measurements were made during baseline and at 1, 5, 15, 30, 45, and 60 min after iv administration. There were no changes produced by the 0.05 mg/kg dose of apomorphine except for a decrease in mean systemic arterial pressure (AoPm) at the 1 through 15 min recordings. For the 0.5 mg/kg dose, there were reductions in systemic vascular resistance at the 1 and 5 min recordings and in AoPm at the 1 through 60 min recordings. Although not significant, when AoPm fell, heart rate, stroke volume, and cardiac output tended to increase. Action potentials were recorded from superfused Purkinje and endocardial ventricular fibers while exposed to 10(-9) to 10(-5) M apomorphine (10(-10) M is considered therapeutic and 10(-7) M is considered lethal). There were no changes in action potential characteristics of Purkinje fibers, but action potential duration at 90% repolarization prolonged approximately 10-12% in endocardium at concentrations of 10(-6) M and greater. At the usual emetic dose (0.05 mg/kg) apomorphine resulted in no signs of cardiovascular toxicity and, at 0.5 mg/kg, cardiovascular changes were minimal. The emetic dose is higher than that for Parkinson's disease or erectile dysfunction; thus apomorphine appears to be a safe compound for clinical use in dogs and by extrapolation to man.

Action Potentials↗

Tunnelling descent: a new algorithm for active contour segmentation of ultrasound images.

The presence of speckle in ultrasound images makes it hard to segment them using active contours. Speckle causes the energy function of the active contours to have many local minima, and the gradient descent procedure used for evolving the contour gets trapped in these minima. A new algorithm, called tunnelling descent, is proposed in this paper for evolving active contours. Tunnelling descent can jump out of many of the local minima that gradient descent gets trapped in. Experimental results with 70 short axis cardiac ultrasound images show that tunnelling descent has no trouble finding the blood-tissue boundary (the endocardium). This holds irrespective of whether tunnelling descent is initialized in blood or tissue.

Algorithms↗

[Unusual intimal and endocardial damage following suicidal infusion of thiobutabarbital].

A suicide is reported that was carried out by i.v. infusion of thiobutabarbital. The most impressive findings were macroscopical alterations in the venous endothelium, the endocardium of the right atrium and ventricle, and the endothelium of the pulmonary artery. The same local cytotoxic side effects could be reproduced in animal experiments. The lesions are most striking after circulatory stagnation in the agonal and supravital period when highly concentrated thiobarbiturates come into contact with the endothelium.

Adult↗

The ultrastructure of the heart of Oniscus asellus L. and Asellus aquaticus L (Crustacea, Isopoda).

The endocardium of Oniscus asellus L. and Asellus aquaticus L. consists of lipid cells. The epicardium consists of a layer of cells with a vesiculated cytoplasm covered by a thick extracellular fibrous sheet. The myocardium is a single layer of cells, the sarcolemma invaginates at Z disc level forming transverse tubules, and longitudinal tubules branch off from these. At the A-I level longitudinal tubules from transverse systems, which form couplings with the sarcoplasmic reticulum. The sarcoplasmic reticulum appears as perforated sheets enveloping the myofibrils. Two types of nerve terminal are found: one is embedded in a myocardial cell process, the other lies in a myocardial cell depression. They contain clear and dense-cored synaptic vesicles.

Animals↗

Autoradiographic localization of [125I]-C-ANP compared to [125I]-ANP in rat tissue.

Whole-body autoradiography demonstrated the different distribution of [125I]-C-ANP and [125I]-ANP to rat tissues. Highest enrichment of radioactivity of both labelled peptides was found in the kidney. In some organs we found remarkable differences between [125I]-ANP and [125I]-C-ANP. In the kidney cortex, especially in the glomeruli, as well as in the endocardium, the zona glomerulosa and the medulla of the adrenal gland, where high levels of radioactivity after [125I]-ANP administration were detected, no or just few radioactivity was found after administration of [125I]-C-ANP. On the other hand in the kidney papilla and the outer subcortical medulla, characteristic blackening was found after [125I]-C-ANP administration. Those differences might be important for the understanding of pharmacological actions of ANP analogues.

Adrenal Glands↗

Endocardial endothelium in the rat: junctional organization and permeability.

Selective permeability of endocardial endothelium has been suggested as a mechanism underlying the modulation of the performance of subjacent myocardium. In this study, we characterized the organization and permeability of junctional complexes in ventricular endocardial endothelium in rat heart. The length of intercellular clefts viewed en face per unit endothelial cell surface area was lower, and intercellular clefts were deeper in endocardial endothelium than in myocardial vascular endothelium, whereas tight junctions had a similar structure in both endothelia. On this basis, endocardial endothelium might be less permeable than capillary endothelium. However, confocal scanning laser microscopy showed that intravenously injected dextran 10,000 coupled to Lucifer Yellow penetrated first the endocardial endothelium and later the myocardial capillary endothelium. Penetration of dextran 10,000 in myocardium occurred earlier through subepicardial capillary endothelium than through subendocardial capillary endothelium. Penetration of tracer might thus be influenced by hydrostatic pressure. Dextran of MW 40,000 did not diffuse through either endocardial endothelium or capillary endothelium. The ultrastructure of endocardial endothelium may constitute an adaptation to limit diffusion driven by high hydrostatic pressure in the heart. Differences in paracellular diffusion of dextran 10,000, between endocardial endothelium and myocardial vessels, may result from differing permeability properties of the endocardium and underlying myocardium.

Animals↗

The early development of the heart of Tupaia belangeri, with reference to other mammals.

The development of the heart of Tupaia belangeri from the first endothelial-lined lumina to the cardiac loop is described in 20 embryos with 2 to 14 somites, from ontogenetic days 11 and 12. Bilateral endocardial tubes transporting blood are found in the 8-somite embryo; in the middle cardiac plate, angioblasts and angiocysts are located between them. In the 9-somite embryo, formation of the cardiac loop has started, the endocardial tubes approach each other closely, most of the angiocysts have been incorporated by the expanding endocardial tubes, and fusion of the endocardial lumina has started in the cono-truncal area. Apparently, much of the endocardial cardiac loop found in the 9-somite embryo has been produced by the disproportionate lengthening of a segment of the endocardial tubes, which is very short in the 8-somite embryo. In the 13-somite embryo the endocardial tubes have largely fused, but tube-like strands of endothelia, remnants of the original endothelial walls separating them, form a "palisade" and mark the original boundary between them. Myoepicardial differentiations of the splanchnopleure begin separately on both sides of the embryo and gradually spread craniad until they coalesce in the midline, in front of the anterior intestinal portal. The caudal portions of the endocardial tubes with initial myoepicardial and cardiac jelly differentiations do not contribute to the definitive heart. The anterior intestinal portal is very broad in Tupaia. Contradictions in the literature as to the bilaterality of cardiac primordia of eutherian mammals are discussed. The hypothesis is developed that bilateral endocardial tubes and bilateral myoepicardial differentiations of the splanchnopleure develop in species with a large yolk-sac, relatively late closure of the foregut, and a broad anterior intestinal portal (e.g., Tupaia, ferret, and cat, etc.). This is probably the primitive condition in eutherian mammals. In species with a small yolk-sac and/or reversal of germ layers (man, rodents), the foregut and anterior intestinal portal are formed earlier, the heart primordium reaches its median position ventral to the foregut in the angiocyst-stage, and the first endocardial lumina appear close to the midline. In these species, the primordium of the endocardium seems to be plexiform and without clear evidence for bilaterality.

Animals↗

Ultrahistochemical studies on the moderately electron dense bodies in teleostean endocardial cells.

The staining properties of phosphotungstic acid (PTA) on moderately electron dense bodies (MDB), are studied in the endocardium of Gadus morrhua. In MDB fixed in aldehydes only, and stained with PTA at a low pH (0-1), Intensly electron dense material occurs on and beneath the limiting membrane. This latter area displays a declining electron density when stained with PTA solutions in which the pH is raised from 1 to 4. At pH greater than 5 the peripheral matrix appears nearly unstained. Collagen fibres fixed as above, and then stained with PTA at a low pH, appear electron dense. These results suggest that the peripheral matrix of the MDB consists mainly of basic proteins.

Animals↗

The transmural progression of the no-reflow phenomenon in globally ischemic hearts.

A no-reflow phenomenon (NRP) develops in hearts subjected to global ischemia and prevents reperfusion of the subendocardial myocardium upon restoration of arterial supply. In the present study the transmural progression of the NRP across the left ventricular wall in globally ischemic rat hearts was quantitatively defined by using autoradiographic nuclear track emulsion (NTE) as an indicator of microvascular competence. Rat hearts were isolated and perfused for 10 min with oxygenated Krebs-Henseleit buffer, then were made completely globally ischemic for from 0 to 60 min and were maintained at 37 degrees C. They were then fixed by perfusion with glutaraldehyde after which NTE was injected into the coronary arteries. Transverse sections through the left ventricles were examined by scanning electron microscopy using back-scattered electron imaging and the vessels in a standard transmural contiguous series of photomicrographs were classified according to whether they did or did not permit the flow of NTE. Non-ischemic control myocardium showed a mean proportion of filled vessels of 99.4 +/- 0.5% SD, and those subjected to 15 min of ischemia showed only a slight overall reduction. After 30 min of ischemia 96 +/- 3% of vessels in the subepicardial third could be reperfused, but the proportion progressively diminished across the myocardium to total no-reflow near the endocardium. From 45-60 min of ischemia the totally non-reperfusible region remained confined to the subendocardial third but there was a significant reduction in the proportion of reperfusible vessels in the subepicardial third to 40% +/- 27%. Ischemia thus progressively reduces the capacity of myocardium to be reperfused.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Fiber bundle direction in the mammalian heart. An extension of the "nested shells" model.

The orientation of the muscle fiber bundles within the mammalian left ventricle was examined in a variety of mammals. The hearts were arrested in situ in animals with an intact thorax by means of an isotonic K+ solution perfused via the aorta and coronaries. The hearts were then fixed by formalin perfusion through the same vessels and the hearts embedded in gelatin. Serial sections were prepared perpendicular to the Apex-Valve axis. On close examination, the muscle fibers show the change in orientation from endocardium to epicardium previously described by others. In addition, the clefts and voids of the inner one-third to inner one-half of the left ventricular wall add another dimension to the fiber direction: the fiber bundles appear to take a curving course from the middle of the wall to the endocardial surface. This pattern was visible in all studied hearts. Speculations are made on the significance of this anatomic arrangement.

Animals↗

Pharmacotoxic myocardial disease: an endomyocardial study.

Drug-induced toxic changes in the myocardium have become an increasing problem. The effect of drugs on heart morphology may be acute or cumulative. In general, adverse drug reactions manifest themselves as myocarditis (toxic or hypersensitivity), cardiomyopathy with chamber dilatation, or restrictive disease. Drugs affecting embryologic development of the heart will not be discussed. Drugs causing myocarditis can be divided into: toxic myocarditis, e.g., cyclophosphamide. The morphologic changes are dose-related and have lesions of different ages, which include myocyte necrosis with hemorrhage and vasculitis. Fibrous endocarditis, e.g., methysergide. These reactions include thickening of the endocardium and sometimes the cardiac valves with fibrosis. Drugs causing hypersensitivity myocarditis, e.g., thiazide diuretics. In this case, the lesions are not dose-related, are the same age, and there is an eosinophilic infiltrate. Drugs causing cardiomyopathic-like changes of ventricular dilatation and failure, e.g., anthracyclines, particularly adriamycin. This group of drugs cause a gradual myofibrillar loss within cardiac myocytes and a sarcotubular dilatation which is characteristic. The damaged cells are replaced by fibrosis and ventricular failure ensues. With the rapid synthesis of new drugs, the problem of drug cardiotoxicity may be an ever-increasing problem. With the more widespread use of the endomyocardial biopsy, drug-induced heart disease can be documented and the effects of different methods of drug delivery and pharmacologic antagonists studied.

Antibiotics, Antineoplastic↗

The effect of blood oxygen content and the no-reflow phenomenon on the subendocardial ultrastructural reversibility following anoxic arrest using hemodilution variables in the experimental model.

Hemodilution is an accepted modality for the conduct of cardiopulmonary bypass. The degree of hemodilution clinically employed has been in the range of 20 per cent of the initial hematocrit levels. This study was designed to evaluate the effects of hemoglobin levels and oxygen content on the reversibility of the damaged or altered ultrastructure of the endocardial layer of the ventricle and the possible role of the no-reflow phenomenon. Two series of mongrel dogs were subjected to cardiopulmonary bypass at normothermic levels and a standard period of ischemic arrest. One series was with hemoglobin levels between 5 and 10 grams (mean 7.25 grams) designated as moderate hemodilution (MH). The second series was of hemodilution with less than 5 grams of hemoglobin (mean 4.8 grams) designated as severe hemodilution (SH). The ultrastructural alteration gradient (UAG) between the epicardium and endocardium was studied with its relationship to reversibility of myocardial function and animal survival. The study demonstrated a definitive relationship between oxygen content and the reversibility of myocardial damage under normothermic conditions and anoxic arrest. The poor survival results in the SH group; two of 12 are attributed to the low oxygen availability possibly triggering the no-reflow phenomenon which prevents reversibility of the myocardial ultrastructural damage and ultimately the functional integrity of the ventricle.

Animals↗

Effect of autonomic nervous system on the transmural dispersion of ventricular repolarization in intact canine.

The effect of the autonomic nerves on the transmural dispersion of ventricular repolarization in intact canine was investigated. By using the monophasic action potential (MAP) recording technique, monophasic action potentials (MAPs) of the epicardium (Epi), midmyocardium (Mid) and endocardium (Endo) were recorded simultaneously by specially designed plunge-needle electrodes at the left ventricular free wall in 12 open-chest dogs. MAPD90 and transmural dispersion of repolarization among three myocardial layers as well as the incidence of the EAD before autonomic nervous stimulation and during autonomic nervous stimulation were compared. The results showed that the MAPD90 of Epi, Mid and Endo before autonomic nervous stimulation were 278 +/- 11 ms, 316 +/- 16 ms and 270 +/- 12 ms respectively, the MAPD90 of Mid was significantly longer than that of Epi or Endo (P<0.01). MAPD90 of Epi, Mid and Endo were shortened by 19 +/- 4 ms, 45 +/- 6 ms, 18 +/- 3 ms respectively during sympathetic stimulation. Compared with that of the control, the transmural dispersion of repolarization during sympathetic stimulation was shortened from 44 +/- 4 ms to 15 +/- 3 ms (P<0.01), but early afterdepolarizations were elicited in the Mid of 5 dogs (41%) during sympathetic stimulation. Parasympathetic stimulation did not significantly affect the MAPD90 in the three layers. It is concluded that there is the transmural dispersion of ventricular repolarization in intact canine. Sympathetic stimulation can reduce transmural dispersion of repolarization, but it can produce early afterdepolarizations in the Mid. Parasympathetic stimulation does not significantly affect the transmural dispersion of ventricular repolarization.

Action Potentials↗