Beneficial effects of mesenchymal stem cells on adult porcine cardiomyocytes in non-contact co-culture.

Mesenchymal stem cells (MSCs) have been reported to improve survival of cardiomyocytes (CMCs) and overall regeneration of cardiac tissue. Despite promising preclinical results, interactions of MSCs and CMCs, both direct and indirect, remain unclear. In this study, porcine bone marrow MSCs and freshly isolated porcine primary adult CMCs were used for non-contact co-culture experiments. Morphology, viability and functional parameters of CMCs were measured over time and compared between CMCs cultured alone and CMCs co-cultured with MSCs. In non-contact co-culture, MSCs improved survival of CMCs. CMCs co-cultured with MSCs maintained CMCs morphology and viability in significantly higher percentage than CMCs cultured alone. In viable CMCs, mitochondrial respiration was preserved in both CMCs cultured alone and in CMCs co-cultured with MSCs. Comparison of cellular contractility and calcium handling, measured in single CMCs, revealed no significant differences between viable CMCs from co-culture and CMCs cultured alone. In conclusion, non-contact co-culture of porcine MSCs and CMCs improved survival of CMCs with a sufficient preservation of functional and mitochondrial parameters.

Propofol-induced mitochondrial and contractile dysfunction of the rat ventricular myocardium.

Propofol is a short-acting hypnotic agent used in human medicine for sedation and general anesthesia. Its administration can be associated with serious cardiovascular side-effects that include decrease in arterial blood pressure and cardiac output. The aim of the present study was to evaluate propofol effects on mitochondrial respiration, myocardial contractility and electrophysiology in the same samples isolated from the heart ventricles of adult rats. Mitochondrial oxygen consumption was measured in permeabilized samples dissected from free walls of both ventricles using high-resolution respirometry. State LEAK was determined with malate and glutamate. Active respiration was induced by ADP (state PI) and further by succinate, a Complex II substrate (PI+II). Rotenone was injected to measure state PII. Antimycin A, a Complex III inhibitor was used to determine residual oxygen consumption (ROX). N,N,N',N'-tetramethyl-p-phenylenediamine dihydrochloride and ascorbate were injected simultaneously for respirometric assay of cytochrome c oxidase activity (CIV). Isometric contractions and membrane potentials were determined on multicellular preparations isolated from right and left ventricles. Propofol concentrations used ranged from 0.005 to 0.5 mmol/l. All respiratory parameters were significantly higher in the left control ventricles compared to the right ones. Propofol significantly decreased Complex I activity at concentration 0.025 mmol/l and papillary muscle contraction force at 0.1 mmol/l. Propofol did not affect action potential duration at any concentration studied. Our study suggests that mechanisms contributing to the impaired myocardial contraction during propofol anesthesia might include also mitochondrial dysfunction manifested by compromised activity of the respiratory Complex I.

Effect of prostaglandin E2 on the ductus arteriosus in the newborn rat. An ultrastructural study.

A patent ductus arteriosus (DA) was maintained in newborn rats (Wistar strain) by administering prostaglandin E2 (PG E2) in doses of 15 micrograms.kg-1 at 30 min intervals up to 300 min after birth. In the control animals, the DA was functionally closed 300 min after birth. The lumen was blocked by clustered endothelial cells at various stages of degeneration. Elastic membranes of the media had disintegrated into irregular fragments and the smooth muscle cells were contracted. Cytoplasm excrescences formed on their surface as a result of contraction protruded as hernias into adjacent muscle cells and into endothelial cells. The smooth muscle cells degenerated. The administration of PG E2 inhibited contraction of the smooth muscle cells and so also the development of degenerative changes; 300 min after birth the DA was fully patent, the elastic membranes were structurally intact, regularly organized and continuous. The smooth muscle cells had the character of synthesizing cells with richly developed granular endoplasmic reticulum. The intima and its endothelial lining were likewise free from structural changes. The ultrastructural image of the wall of the DA correspondent to the state 10 min after birth, when the DA was fully patent. The administration of PG E2 did not induce any ultrastructural changes indicative of injury to the wall of the DA.

Effect of the administration of prostaglandins (PGE2) in the early postnatal period on closure of the ductus arteriosus in the laboratory rat.

Maintenance of a patent ductus arteriosus by means of prostaglandins enables the surgical correction of a congenital heart defect in infants to be postponed until a phase of development when the operation hazards are smaller. We investigated the pathophysiological consequences of this therapeutic measure in an experimental model in which E2 prostaglandin was administered to newborn laboratory rats. It was found that, physiologically, the ductus arteriosus (DA) closed progressively within 180 min after birth. The repeated administration of PGE2 (subcutaneously, 15 micrograms.kg-1 every 30 min from the 5th min after birth) blocked closure of the DA, which was still fully patent 300 min after birth. Histological tests showed no significant differences in the structure of the tunica media of the physiologically patent and the PGE2-treated DA. The results show that PGE2 also inhibit physiological closure of the DA in newborn rats. Long-term study of this pathophysiological process is at present impeded by the need for the continuous administration of prostaglandins.

The physiological closure of ductus arteriosus in the rat. An ultrastructural study.

The evolution of morphological changes in the wall of the ductus arteriosus during its physiological closure in newborn rats was examined by electron microscopy. The contraction of smooth muscle cells in the tunica media seems to be the primary mechanism which leads to the physiological closure of the ductus arteriosus. For this reason our attention was centred mainly on the morphology of the tunica media. No important changes in the ultrastructure of smooth muscle cells can be observed in the early phases of the closure. Most of them exhibit ultrastructural features of cells with enhanced synthetic activity during all phases of the closure. The permanent contraction of smooth muscle cells results in their morphological changes. The most striking is the herniation of smooth muscle cell cytoplasm into the endothelial and later into adjoining muscle cells. These changes together with signs of degeneration of the smooth muscle cells are already clearly discernible 120 min after birth. The elastic component of the tunica media exhibits surprisingly fast changes. As soon as 60 min after birth, the fragmentation of elastic membranes and their structural changes provided evidence about the degradation of elastic material. The matrix vesicles, probably derived from the lysosomal apparatus of the muscle cells, may play an essential role in this process.

Intermittent high altitude hypoxia - induced structural changes in the pulmonary myocardium in young mice.

Seven-day-old mice, strain H, were exposed to intermittent high altitude hypoxia (IHA) in a barochamber (7,000 m, 4 h/day, 5 days a week); a total number of exposures was 10. It has been shown that the layer of cardiac musculature in the adventitia of the pulmonary veins, the so-called pulmonary myocardium, reacts to IHA hypoxia by enlargement even sooner than the right ventricular myocardium. The average thickness of the layer of pulmonary myocardium was significantly greater in animals exposed to IHA hypoxia as compared with the controls. Furthermore, IHA hypoxia induces the extension of the pulmonary myocardium to the periphery of the pulmonary venous bed. Ultrastructural investigation of the pulmonary myocardium in hypoxic animals revealed the presence of unoriented myofilaments in the peripheral myofibril-free sarcoplasma, increase in the number of ribosomes and appearance of profiles of granular endoplasmic reticulum. Our data provide quantitative support for the hypothesis that it is not only the contraction of pulmonary arteries, but also venoconstriction which contribute to the hypoxic pressure response in mice.

Ultrastructure of pulmonary macrophagic system.

A mature AMF, ready for phagocytosis, is a relatively large cell with an oval nucleus, with indentations of a nuclear envelope of varying depth. Evenly distributed chromatin forms beneath the nuclear envelope a rim of heterochromatin. There is a prominent nucleus with a distinct nucleolonemma. The cytoplasm is differentiated into a continuous ectoplasmic zone with numberous finger-like processes and pseudopodia. The organelles are formed by scattered round or oval mitochondria, a description is given of the Golgi apparatus in the juxtanuclear position, in certain sites multifocal in form, of the centriolar apparatus, scarce profiles of endoplasmic granular reticulum, dispersed free polyribosomes and microfilaments in varying amounts. The most outstanding feature is the rich lysosomal apparatus of varying structure depending on the functional state. Dynamics of the maturation of AMF was studied in intermediate phases, described here. The peribronchovascular connective tissue is the seat of free macrophages of a structure analogous with that of AMF. Fixed macrophages are anchored in the loose connective tissue by processes of different shape and length. The prevailing component of the cytoplasm are numerous vesicular structures and vacuoles as well as a marked lysosomal apparatus. Fixed macrophages phagocytize foreign material in situ. The septal cell exists in normal state as an element with numerous intricated processes pervading the fibrillar substrate. Numerous free polyribosomes and vacuoles are its most marked component. Activation of septal cells was demonstrated under experimental conditions. Their transformation into free macrophages is probable. In the pulmonary intersitium, in the perivascular loose connective tissue particularly free cells of a similar structure as blood monocytes were shown in normal state. Under experimental conditions an increased number of monocytes is present in pulmonary capillaries, e.g. 24 hours after an intratracheal instillation of India ink colloid solution. At the same interval a number of free cells of the monocyte structure was found in the perivascular connective tissue and also in the alveolar lumen, with phagocytized carbon. An increased number of monocytes transforming into macrophagic cells was visualized in this localization as late as 14 days after the instillation of India ink colloid solution. The experimental study with an intratracheal instillation of India ink colloid solution in the mouse gave evidence of a high readiness of AMF. The carbon particles were seen phagocytized at an interval of 5 minutes after instillation. In all intervals during which free carbon particles were present in the alveolar epithelium mature AMF were observed with no or very low phagocytic activity, their lysosomal apparatus being prominent. On the contrary, in the phagocytizing AMF, the lysosomal apparatus disappeared in the greatest part of the cytoplasm. The clearance of AMF occurs predominantly by the air route...