MAST CELLS IN DAMAGED LYMPH-HEART STRIATED MUSCLE OF ADULT FROG: AN ULTRASTRUCTURAL, IMMUNOCYTOCHEMICAL AND AUTORADIOGRAPHIC STUDY.

Ultrastructural features of mast cell activation were studied in degenerative/reparative experimental model of mechanically damaged lymph-heart striated muscle during the first postoperative week. 24 h after damage, the intracytoplasmic empty degranulation channels were revealed in a certain part of resident and circulating mast cells (MCs) located near the site of injury. These findings are evidence that previously ther was activation and secretory response of MCs by a process known as compound exocytosis which involves not only granule- to-plasma membrane fusions and formation of degranulation channels. We have found rare MCs showing release of single. altered, membrane-free granules into the space restricted by the plasma membrane of mature resident MCs and the basal lamina surrounding them. During the fist postoperative week we have more commonly observed activated lymph-heart MCs with prominent intragranular changes characterized by diminishing of electron-dense component of granules and loss of granule matrix. We have also revealed close association of some unaltered and altered secretory granules with completely or partially empty degranulation channels that permeate the mast cell cytoplasm. Some of these channels remained open directly to the exterior of the cells. It is not improbable that these findings provide the ultrastructural basis for potential discharge of small amounts of granule mediators into the open channels (incomplete exocytosis) duding partial degranulation of activated in vivo frog MCs. Immunocytochemical staining for histamine of activated frog MCs showed the presence of the immunoreactive material both in unaltered and altered cytoplasmic granules. As a rule, gold particles were located over residual electron-dense component of altered granule, whereas little or no gold particles were revealed over the swollen matrix. Ultrastructural features of secretory granule formation in the cytoplasm of some MCs were observed toward the end of the first postoperative week. Electron-microscopic autoradiography has shown that replicative and transcriptional activity of MCs is not observed in the studied postoperative period. No mitoses in MCs have been revealed in this period. A few mast cells were found in necrosis. Taken together, our results indicate that MCs are one the first cell types that respond to injury of lymph-heart striated muscle and release a variety of preformed mediators, including histamine. These findings suggest that activated MCs play an important role in the initiation of an inflammatory process during repair of damaged lymph-heart striated muscle.

Comparison of clinical and laboratory parameters in patients with end-stage renal failure in the outcome of chronic glomerulonephritis and patients with end-stage renal failure in the outcome of other diseases.

BACKGROUND: frequent complications of hemodialysis treatments are coagulation disorders. This is due to activation of the coagulation of blood flow in the interaction with a dialysis membrane material vascular prostheses and extracorporeal circuit trunks. In addition, in hemodialysis patients receiving heparin for years, there is depletion of stocks in endothelial cells in tissue factor inhibitor, inhibits the activity of an external blood clotting mechanism. AIM: the aim of our study was to evaluate the hemostatic system parameters in patients with end-stage renal failure, depending on the cause of renal failure. MATERIAL AND METHODS: to evaluate the hemostatic system parameters in patients with end-stage renal failure, depending on the cause of renal failure and hemodialysis treatment duration conducted a study that included 100 patients observed in the department of chronic hemodialysis and nephrology hospital №1 Republican National Medical Center in the period of 2013-2016. RESULTS: in patients with end-stage renal failure in the outcome of chronic glomerulonephritis, a great expression of activation of blood coagulation confirm increased the mean concentration of fibrinogen, whereas in the group, which included patients with end-stage renal failure in the outcome of other diseases, such is not different from the norm, and a higher rate of hyperfibrinogenemia, identified in 2/3 patients in this group. CONCLUSIONS: it was revealed that the state of homeostasis in patients with end-stage renal failure in increasingly characterizes the level of fibrinogen and the activation of the hemostatic markers: soluble fibrin monomer complexes, D-dimers.

[Mast cells of lymph hearts during ontogenesis of frogs Rana temporaria].

Electron microscopic observations of the lymph hearts of tadpoles and yearling frogs of Rana temporaria showed that mast cells (MCs) were present not only between muscle fibers (population of resident MCs), but in the cavities of lymph heart (population of circulating MCs), too. There were some differences in the ultrastructure of the resident MCs at each studied stage of larval development. The first recognizable MCs were revealed in the lymph hearts at premetamorphosis (stages 39-41). MCs presented as mononuclear relatively small and slightly elongated cells with a few immature secretory granules and numerous free ribosomes, polysomes and short cisternae of rough endoplasmic reticulum (RER) in the cytoplasm. Chromatin of their nuclei was poorly condensed; the Golgi apparatus was moderately developed. At pro-metamorphosis (stages 44-45), we revealed MCs at different levels of their differentiation. Some MCs demonstrated an active process of granulogenesis in their cytoplasm. Among densely packed cytoplasmic organelles, immature secretory granules were closely associated with cisternae of RER and free ribosomes. Other MCs appeared as more differentiated cells. They were characterized by a predominantly heterochromatic nuclei and cytoplasm filled with polymorphic and heterogeneous granules. MCs also showed a reduction in the number of free ribosomes and cisternae of RER in the cytoplasm. On the contrary, the Golgi apparatus was well developed. Stacks of Golgi cisternae, detaching vacuoles, and progranules occupied the perinuclear region. The majority of the outlines above ultrastructural features of differentiated MCs were typical for MCs of yearling frogs. At metamorphic climax (stages 52-53), MCs often tightly contacted with macrophages. We did not reveal apoptotic MCs. However, some MCs exhibited morphological features typical for programmed necrosis-like death, which was characterized by mitochondria swelling, dilatation of cisternae of RER and nuclear envelope, plasma membrane rupture and subsequent loss of intracellular contents. Electron microscopical immunocytochemistry revealed the localization of atrial natriuretic peptide (ANP), substance S (SP) and heat shock protein (Hsp70) in the secretory granules of the resident and circulating MCs at different stages of tadpole development and in yearling frogs.

[Resident and circulating mast cells in propulsative organs of the frog Rana temporaria].

Mast cells (MCs) of the "blood" and lymph hearts of the adult frog Rana temporaria were investigated at histochemical and ultrastructural levels. Two populations of MCs were revealed in these propulsative organs: population of resident MCs and population of circulating MCs. It has been shown that the resident cardiac MCs have an oval or elongated form and are located between atrial or ventricular myocytes and under endocardial endothelium. The resident cardiac MCs are situated in connective tissue of epicardium, too. Avascular myocardium of the frog ventricle consists of a spongy network of muscle trabeculae. We revealed circulating MCs in intertrabecular spaces and clefts of the spongy myocardium and in the blood of the main central cavity. Circulating MCs are round in shape and contain a large central nucleus enriched with condensed chromatin. They resemble the lymphocytes, but show cytoplasm filled with granules. These granules ultrastructure is much like that of the granules of the cardiac resident MCs. In the lymph heart, oval and somewhat elongated resident MCs are located in the interstitial space among cross-striated muscle fibers and among smooth muscle cells of tubular (afferent and efferent) valves. Sometimes lymphocyte-like circulating MCs are revealed in the cavity of lymph heart. Circulating MCs are also present in the lymphatics located adjacent to the lymph hearts. In certain parts of the lymphatic walls MCs are in close adhesion to the mesothelial cells lining the lymphatic cavity. Our histochemical investigation revealed that both the resident and circulating MCs of the propulsative organs give a strongly positive reaction with alcian blue, but weakly red with safranin and weakly metachromatic with toluidine blue. The presence of population of circulating MCs in the frog suggests that there are differences in biology of MCs between lower and higher vertebrates.

[Chromogranin A: immunocytochemical localization in secretory granules of frog atrial cardiomyocytes].

Chromogranin A (CgA) is a member of the granin family of acidic proteins that present in the secretory granules (SGs) of many endocrine, neuroendocrine and neuronal cells. Atrial natriuretic peptide (ANP)-storing SGs in atrial cardiomyocytes of rat heart also contain CgA. Cardiosuppressive effect of CgA-derived peptides (vasostatins) on in vitro isolated and perfused working frog and rat hearts has been shown under both basal conditions and beta-adrenergic stimulation. More recently it has been revealed that rat heart produces and processes CgA-derived vasostatin-containing peptides. Until now nothing has been known about the presence of CgA in an amphibian heart. We have investigated the subcellular localization of CgA in atrial myocytes of adult frog Rana temporaria heart using ultraimmunocytochemical method. Immunocytochemical staining of the frog atrial tissue for CgA and ANP has shown that out of three morphologically different types (A, B and D) of specific cytoplasmic granules (SCGs) present in myocytes only two (A and B)--large (120-200 nm in diameter) granules with more and with less electron dense core--exhibit immunoreactivity (IR) to these two antigens. The third type (D) of granules (80-100 nm in diameter) are small membrane bound granules characterized by highly electron dense core surrounded with a thin halo. These granules revealed negative reaction on immunostaining for both CgA and ANP. The presence of CgA- and ANP-IR in the same SCGs in frog atrial myocytes is consistent with the endocrine nature of these granules. Taking into account our and literature data we propose that CgA present in frog atrial cardiomyocite SCGs might be a precursor of vasostatin-containing peptides, as it takes place in rat heart. It is possible that these CgA-derived peptides together with ANP exert their regulatory function through the autocrine and/or paracrine mechanisms and play important cardioprotective role in frog heart under stress condition.

[Immunocytochemical localization of atrial natriuretic peptide in endothelial and granular cells of the heart of lophotrochozoa]. / Immunotsitokhimicheskaia lokalizatsiia predserdnogo natriiureticheskogo peptida v éndotelial'nykh i granuliarnykh kletkakh serdtsa lofotrokhoforovykh.

In parallel with contraction, vertebrate cardiomyocytes perform endocrine function and produce natriuretic peptides (NP)--ANP and BNP--involved in cardiovascular homeostasis maintenance. ANP-like peptides have been reported also in hearts of some invertebrates, however, their cellular localization was not determined. By electron microscopical immunocytochemistry with polyclonal monospecific antibodies raised against ANP and protein A-gold technique, we have localized ANP-like immunoreactivity in granules within endothelial cells in the heart of the brachiopod Rhynchonella psittacea, the polychaete Arenicola marina, and the gastropod mollusc Achatina fulica--all being representatives of the major phylogenetic group Lophotrochozoa. ANP-like immunoreactivity was also revealed in one of 3 morphologically distinguishable types of granules in the snail heart granular cells. By electron microscopical autoradiography with the use of [3H]-thymidine, the ability for DNA synthesis was demonstrated in heart endothelial cells of the investigated animals. Forms of NP-system organization in hearts of Lophotrochozoa and Vertebrates, and close histogenetic relationships of endothelial and granular cells in the snail heart are discussed.

[Heart ventricular cells of neonatal rats in a culture: DNA synthesis, ultrastructure and localization of atrial natriuretic peptide]. / Kletki zheludochkov serdtsa neonatal'nykh krys v kul'ture: sintez DNK, ul'trastruktura i lokalizatsiia predserdnogo natriiureticheskogo peptida.

We determined the optimal conditions suitable for expanding cardiac cells in vitro for their future use in experimental transplantation into injured myocardium of adult animals. Ventricular cardiac cells were isolated enzymatically from 2-3 day-old rats and cultured at different cell densities within 5-7 days to 4 weeks. Mixed cultures of muscle and non-muscle cells were examined by light autoradiography, electron microscopy, and immunogold method. The best results were obtained at a density of 3 x 10(5) cells/ml in the medium, consisting of 90% DMEM and 10% fetal calf serum, during 5-7 days of cultivation. In such cultures myocytes made 62.5 +/- 7.9%. After a 24 h incubation with 3H-thymidine, 22.0 +/- 2.2% of myocytes were labeled. Muscle cells contact with each other and with non-muscle cells, contain myofibrils, contract and display atrial natriuretic peptide (ANP)-like immunoreactivity.

[Reaction of lymphatic heart muscle in the adult grass frog to autotransplantation. Electron microscopic and electron microscopic autoradiographic studies]. / Reaktsiia myshechnoi tkani limfaticheskogo serdtsa vzrosloi travianoi liagushki na autotransplantatsiiu. Issledovanie metodami élektronnoi mikroskopii i élektronno-mikroskopicheskoi avtoradiografii.

Cellular and subcellular responses, related to destruction, survival and regeneration of striated muscle fibers in the autografted intact lymph heart of the adult frog, were examined by electron microscopy and electron microscopical autoradiography. It has been shown that a small number of myofibers in the grafted lymph heart wall undergo an irreversible degeneration. Numerous myofibers retain their almost normal ultrastructure up to the end of the 1st posttransplantation week and later. However, a considerable part of surviving muscle fibers exhibited various degrees of reversible degenerative changes. Prominent morphological features of activation, related to the nuclear and cytoplasmic ultrastructural reorganization, were revealed in these reversibly degenerated myofibers since the middle of the second week after autotransplantation. It is shown that some myonuclei of these reactive fibers are able to synthesize DNA. No mitoses were observed in myonuclei. The number of satellite cells (normally approximately 11%) doubled by the end of the 3rd week of posttransplantation. Evidence was provided for the formation of some myotubes in the interstitium outside the basal membrane of the original muscle fibers. Cellular sources of regeneration are discussed in addition to peculiar reaction of lymph heart muscle fibers to free grafting.