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Recent studies demonstrate an increasing interest to cardio-vascular biomarkers with assessing the risk in heart ischemic disease. Along with cardiac electrophysiological indices the molecular-mass distribution of peptides appears as one of novel markers of the risk of cardiovascular complications, since they indicate the effect on developing unfavorable cardiovascular events. Fluorescent probes are also applied for diagnosis and forecasting the developing diseases, revealing the risk factors and control of the treatment efficiency. Research aim was to analyze the changes in electrophysiological indices of heart, molecular-mass distribution of peptides in blood serum and albumin loading with ligands during developing necrosis and remodeling of heart with experimental infarction in rats. Research object and methods. Experiments were performed in 30 outbred male rats of 180-250 g weight under inhalation anestheticon spontaneous breath under the regulations approved by the Bioethics Committee of the IPC&Ñ of the National Academy of Sciences of Ukraine. Myocardial necrosis was simulated by left anterior descending coronary artery ligation. Electrocardiograms (ECG) were recorded with digital ECG Poly-Spectrum 8/B system (Neorsoft, Russia). To determine the molecular weight distribution of low molecular weight fractions of peptide nature, the method of highly effective gel permeation chromatography was used. Fluorescence probes K-35 and E-176, synthesized at State Scientific Institution Institute for Single Crystals of the National Academy of Sciences of Ukraine (Kharkiv) was used the studies. The fluorescence spectra were recorded with a Cary Eclipse spectrometry (Varian, Australia). Microcal Origin 6.0 was used to process and visualize the spectra. Experimental studies were performed at days 1, 7, 14 and 30. The results were statistically processed with Kruskal-Wallis criterion using the STATISTICA 6.0 software (StatSoft, USA).Results. During the entire observation period, the electrocardiographic indices noted a change in the phases of the development of an experimental myocardial infarction and the processes of remodeling of the cardiac tissue of anterior-lateral parts of the heart. According to the analysis of the chromatogram of rat serum peptides, it was found that in an acute phase of myocardial infarction, the changes in molecular weight distribution in an acute phase of myocardial infarction were observed, which was reflected by a decrease of peak in the region of macromolecular peptides with m.w.> 10,000 from 61.5% to 3.3%, and the appearance of peaks in the region of low molecular weight peptides within the range of m.w. 290-527. Reducing the fluorescence intensity of probes in one day after the simulation of myocardial infarction indicates an increase in the loading of albumin by ligands. Conclusions. Characteristic peptide molecular-mass distribution in blood serum and albumin loading with ligands for each stage of myocardium necrosis development and its remodeling in experimental myocardium infarction has been shown.

The NO-synthase activity (mtNOS) in mitochondria of uterine smooth muscle was studied. The mitochondrial localization of NO synthesis in myocytes was proved using laser confocal microscopy method and specific fluorescent probes MitoTracker Orange (specific to mitochondria) and DAFFM (NO-sensitive fluorescent probe). It was demonstrated using flow cytometry that nitric oxide biosynthesis in isolated mytochondria decreased in the presence of a constitutive NOsynthase blocker 2-aminopyridine (100 μmol/l, 50% inhibition) and monoclonal antibodies (2.5 μg anti-Letm1/50 μg protein) against the H+-Ca2+-exchanger (Letm1 protein), but was't sensitive to the mitochondrial permeability transition pore inhibitor cyclosporin A (5 μmol/l). A decrease of potassium ions concentration in the incubation medium and the presence of various types of potassium channel inhibitors significantly inhibited the NO-synthase reaction. We have concluded that potassium permeability of the inner mitochondrial membrane plays important role in the regulation of mtNOS activity. [ABSTRACT FROM AUTHOR]

The work aimed to investigate the use of new probes Е-176, К8-3000 and К8-1300 to research the specificity of peptides in the extracts of cryopreserved fragments of organs of pigs’ and piglets’ and their interaction with blood serum albumin. Extracts of cryopreserved fragments of skin of new-born piglets’, spleen of pigs’, heart of mature pigs’ and new-born piglets’ cleared of proteins were used in the research. Titration of extracts with investigated probes revealed differences in the peptides properties that are a part of them, to bind probes, reflecting tissue specificity of peptide drugs. Adding the extracts to albumin solution triggered quenching of intrinsic fluorescence of albumin and affected the state of tryptophan residues of the protein. Investigated probes show a high sensitivity to the interaction with blood serum albumin substances of peptide nature and could be recommended as new fluorescent probes for use in the biomedical researches. Since fluorescence of these probes is excited in the long wavelength of spectrum, it enables to reduce the artifacts by their use that are introduced by chromophore groups of the biomacromolecules absorbing in the visible region.

The work aimed to investigate the use of new probes E-176, K8-3000 and K8-1300 to research the specificity of peptides in the extracts of cryopreserved fragments of organs of pigs' and piglets' and their interaction with blood serum albumin. Extracts of cryopreserved fragments of skin of new-born piglets', spleen of pigs', heart of mature pigs' and new-born piglets' cleared of proteins were used in the research. Titration of extracts with investigated probes revealed differences in the peptides properties that are a part of them, to bind probes, reflecting tissue specificity of peptide drugs. Adding the extracts to albumin solution triggered quenching of intrinsic fluorescence of albumin and affected the state of tryptophan residues of the protein. Investigated probes show a high sensitivity to the interaction with blood serum albumin substances of peptide nature and could be recommended as new fluorescent probes for use in the biomedical researches. Since fluorescence of these probes is excited in the long wavelength of spectrum, it enables to reduce the artifacts by their use that are introduced by chromophore groups of the biomacromolecules absorbing in the visible region. [ABSTRACT FROM AUTHOR]