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Currently, the mortality rate in pediatric population resulted from such diseases as respiratory distress syndrome, pneumonia and infections is extremely high. It cannot be ruled out that this is due to impaired functioning of the children's immune system. The thymus gland appears to be the central organ of immunopoiesis in children, that is why pediatricians pay special attention to thymic hyperplasia. To date, there is no consensus on the nature and clinical significance of the thymic enlargement. The aim of research was to study morphometric parameters of the thymus gland in children of the first year of life having thymic enlargement. Material and methods . To analyze the incidence of thymic enlargement in deceased children, the authors selected 764 autopsy protocols of children for the period from 2012 to 2017; of these, 117 were children who died in their first year of life. Autopsies were performed in Saratov Centralized Pediatric Prosectorium. The thymic enlargement in deceased children of the first year of life was detected in 34% (40 cases). Autopsy material of 32 children was selected (15 cases with and 17 cases without the thymic enlargement) for histological examination. The thymic index was calculated to specify correlation between the thymus mass and the age and body weight of a child. Morphometric studies were performed using an mVizo-103 microvisor. Results. As demonstrated, infectious diseases are a risk factor for the formation of thymomegaly in children of the first year of life. A morphological study detected that the thymic enlargement occurred mainly due to the enlargement of the cortical layer with the reduction of the lobule area. The lobule area in the studied group of children reduced by 15%, and the number of lobes bordering in the field of view increased by 50%, respectively to the similar parameters in children of the comparison group. Conclusion. Thus, infectious pathologies appear to be the most common cause of death in children with the thymic enlargement. The study results support the fact that histological signs of thymomegaly are the increased number of bordering thymic lobes in the field of view and the increased number of immature thymocytes.

Using fluorescence methods for visualization of human tumor cell apoptosis and necrosis in vitro, the antitumor effects of flavonoid-containing Gratiola officinalis L. and green tea extracts at ultralow concentrations on A498 human kidney carcinoma cells after 24 and 48 h were compared. Fluorescent dyes (propidium iodide and acridine orange) were used in the “living and dead” test, which made it possible to identify the total number of cells that died by necrosis and apoptosis, as well as cells in which apoptosis has started and apoptotic bodies or pycnosis of the nucleus were detected. Gratiola officinalis L. extract had the most pronounced cytostatic activity 48 h after exposure; the number of cells per field of view decreased by a factor of 1.5 compared with the control. Green tea extract at a concentration of 0.0288 mg/mL had the most pronounced cytotoxic activity; its manifestation was growing 48 h after exposure. Gratiola officinalis L. extract exhibited cytotoxic activity at lower concentrations (0.0036 mg/mL).

The goal of our research was to assess the vascularization degree of tumor to predict the efficiency of plasmonic photothermal therapy (PPT) and photodynamic therapy (PDT) in tumor-bearing rats. Before any treatment, 3D Doppler ultrasound imaging was used for assessment of the vascularization degree of transplanted rat cholangiocarcinoma. For PPT, the gold nanorods with aspect ratio of 4:1, functionalized with thiolated polyethylene glycol, were used. After multiple fractional intravenous (IV) injections in rats with cholangiocarcinoma, the tumours were irradiated through the skin by an 808-nm NIR diode laser at a power density of 2.3 W/cm2 for 15 min. For PDT, galactose – Luphthalocyanine, as a photosensitizer, was applied by intratumoural injection, in a dose of 2 mg/kg. Then, tumors were irradiated through the skin by 670 nm-diode laser source with power density 200 mW/cm2 applied for 1000 sec in three non-overlapping zones covering whole tumour surface to obtain a total irradiation dose of 200 J/cm2 for each lesion. The withdrawal of the animals from the experiment and sampling of tissues for morphological study were performed before and 72 hrs after PPT and PDT. The vascular microdensity in tumors was assessed on histological sections as vessel counts or vessel area per unit of assessed tumor area. It was shown that efficiency of PPT and PDT therapy was mostly due to the sufficient accumulation of photothermosensitizers in the tumor, therefore preliminary assessment of tumor vascularization degree was necessary before starting a therapy.

The influence of long-term peroral injection of gold nanoparticles with various sizes on the liver, spleen, and lymph nodes of adult laboratory rats and their progeny was studied. The morphometric analysis was conducted using a µVizo-103 medical digital microviser. The conglomerated gold nanoparticles were detected in tissues via the dark-field microscopy. Gold nanoparticles with sizes of 2 and 50 nm were found to cause the mostly pronounced morphological alterations in organs that testifies to their size-dependent impact. The liver in a group receiving 50-nm gold particles exhibit noticeable dystrophy of hepatocytes and activation of a monocyte macrophage system. The spleen and lymph nodes reveal the activation of proliferation and differentiation of immunocompetent cells and macrophages. Peroral injection of gold particles during gestation of females led to an increase in a number of hepatocytes with necrobiotic changes as well as to elevated amount of non-parenchymatic elements in the liver of the progeny, especially in groups containing the 2- and 50-nm gold nanoparticles.

The possibilities of using fluorescence-based analytical methods and their advantages for visualization and identification of the type of programmed cell death in human tumor cells exposed to flavonoids have been analyzed in experiments in vitro. HeLa cervical cancer cells and A498 kidney carcinoma cells were used as the objects of the study, and exposure to the flavonoid-containing extract of common hedgehyssop (Gratiola officinalis L.) was tested as the experimental treatment. The following fluorescence-based techniques were used: the “live and dead” test with double staining by propidium iodide and acridine orange and double staining with annexin and propidium iodide. Autophagy induction was confirmed by fluorescence-based tests implemented in a Muse cell analyzer with a Muse Autophagy LC3-Antibody Based Kit. The use of double staining with acridine orange and propidium iodide fluorescent dyes in the “live and dead” test and comparison with phase contrast microscopy enables the visualization of apoptotic body and autophagosome formation processes in the cells and can, therefore, be used as a method of screening assessment of the efficiency of various chemotherapy drugs.

The extract of G. officinalis was separated by five major fractions and the antitumor activity of hydrophobic fractions has been established. For the most active fractions spectral investigations of the main component were performed using UV-Vis electronic spectra, circular dichroism (CD), Fourier-transform infrared spectroscopy (FTIR) and nuclear magnetic resonance (NMR) spectroscopy. On the basis of spectral characteristics, it was demonstrated that the main components of the fractions 4 and 5 of G. officinalis providing its biological activity, highly likely, are of flavonoid nature. Based on the NMR data we propose the structure of the aglycon moiety of the main component from fraction 4 as 4,2’,5’-trihydroxychalcone. The fraction 5 is the most active, the antitumor effect of which is implemented through cytotoxic, cytostatic, apoptotic and autophagosomal activity in respect to human kidney cancer tumor cells.

The purpose of the research was to clarify the effect of chicken broilers feeding in the composition of mixed fodder with Vitamins E and C on the cellular link of non-specific resistance of the organism and, in particular, the parameters of pseudoiesinophils phagocytosis in blood. The research was carried out on chickens broiler – 308 grown in the farm “Fedyuk M” in the Zolochiv district of Lviv region. The keeping of the chickens was cellular with free access to feed and water. Experiments were conducted in 4 groups of broiler chickens of 100 chicks in each according to the scheme: the control group was fed with the standard feed (SC) according to the existing norms recommended for the cross ROSS-308; the first experimental group in addition to the SC received vitamin Е 1g/10 kg of mixed fodder; the second experimental group received vitamin C 2.5 g/10 kg of mixed fodder; the third experimental groups in addition to the ration received Vitamin C 2.5 g/10 kg and Vitamin E 1 g/10 of mixed fodder. For carrying out of immunological researches in chickens blood was taken in different age periods: 27-, 34- and 41-day-old age. In the whole blood, the phagocytic activity of pseudoiesinophils (FA), phagocytic index (FI) and phagocytic number (FF) were determined. For research, the daily culture of Escherichia coli (strain VKM-125) was used. The stimulatory effect of vitamins E and C in the composition of mixed fodder for chicken broiler broilers on the indices of pseudoiesinophils phagocytosis of blood was determined. As evidenced by a significantly higher phagocytic activity, phagocytic index and phagocytic count of pseudoiselinophils in blood of chickens in the experimental groups by comparison with the control group. At the same time there was a direct correlation between phagocytic activity and phagocytic index and index in blood of chickens broiler in experimental groups. At the same time, the ability of blood pseudoiesinophils to phagocytosis of microbial cells was higher in chicken broilers, which were fed with vitamin E and C.

Objective of the study: We used fluorescence imaging methods of apoptosis and necrosis in human renal carcinoma A498 tumor cells in vitro to reveal the indicated forms of cell death under the combined effect of flavonoid-containing extract of Gratiola officinalis and cytostatic (cyclophosphamide). Materials and methods: The dyes were propidium iodide and acridine orange, which were used in the “alive and dead” test. This test helped us to identify the total number of dead cells in the forms of necrosis and apoptosis and the number of cells in which apoptosis had started, it was characterized by the appearance of apoptotic bodies or nucleus pyknosis. Results: We found the most pronounced cytotoxic activity at the ratio of extract of Gratiola officinalis and cyclophosphamide concentrations of 1:1. The number of living cells decreased when exposed to the ratio of extract and cytostatic concentrations of 2:1. When the ratio of concentration of the extract relative to the cytostatic increased to 3:1, the cytostatic activity of the extract began to appear, the total number of tumor cells decreased. The number of cells with nucleus pyknosis and the number of cells with apoptosis signs significantly increased at a 3:1 ratio of extract and cytostatic concentrations, which confirms the presence of pro-apoptotic activity of the studied combination. This trend indicates the dependence of a certain form of cell death (apoptosis, necrosis) on the ratio of extract and cytostatic doses, and it also demonstrates the cytostatic and cytotoxic effects of this combination. Conclusion: Fluorescence methods of investigation in the “alive and dead” test allowed us to visualize the forms of cell death of human kidney carcinoma A498 by combined exposure to the flavonoid-containing extract of Gratiola officinalis and cytostatic (cyclophosphamide) 24 h after exposure. We found that the combination with a concentration ratio of the extract and cyclophosphamide of 3:1 has the greatest effectiveness due to stimulation of the cytostatic effect and cytotoxic effect.

To establish the mechanisms of pathomorphism of transplantable kidney cancer in rats that used flavonoid-containing hedge hyssop (Gratiola officinalis L.) extract in an in vivo experiments.The experiment was carried out on 30 male Wistar rats with transplantable kidney cancer PA. At 72 hours after tumor inoculation, the rats in the experimental groups received hedge hyssop extract at an oral or intramuscular dose of 110 mg/kg/day for 12 days. A comparison group consisted of the animals with a tumor, but without exposure. The investigators used the immunohistochemical markers of apoptosis (p53, bax, bcl-2, Fas-receptor, Fas-ligand), autophagy (LC3B), proliferation (Ki-67), and angiogenesis (VEGF). During statistical data processing, the Shapiro-Wilk test was used to test the normality of the indicator distribution in the groups. Cramer-Welch's t-test was also employed to compare the groups.Histological examination of tumor tissue under the action of hedge hyssop extract showed the emergence of extensive areas of damage (necrosis and apoptotic bodies). With both routes of hedge hyssop extract administration, there was a sharp decrease in the expression of the proliferation markers Ki-67 and the angiogenesis marker VEGF and a high expression of the apoptosis markers p53, bax, CD95 (Fas/APO-1), and FAS-ligand in tumor cells and its absence in the comparison group. All the described changes were more pronounced with intramuscular administration. The expression of the autophagy marker LC3B increased with the oral administration of hedge hyssop extract and decreased with its intramuscular administration.A pronounced pathomorphism of kidney cancer develops due to consumption of hedge hyssop extract. This suppresses the proliferation, angiogenesis, and activation of apoptotic signaling and mitochondrial pathways and blocks protective autophagy. The autophagy marker LC3B can be used as an additional criterion for evaluating the therapeutic pathomorphism of tumors.Цель исследования - в экспериментах in vivo установить механизмы патоморфоза перевитого рака почки под влиянием флавоноидсодержащего экстракта аврана. Материал и методы. Эксперимент проводили на 30 самцах крыс Wistar c перевитым раком почки РА. В опытных группах крысы через 72 ч после перевивки опухоли на протяжении 12 сут получали экстракт аврана перорально или внутримышечно в дозе 110 мг/кг/сут. Группу сравнения составили животные с опухолью, но без воздействия. Использовали иммуногистохимические маркеры апоптоза (p53, bax, bcl-2, Fas-receptor, Fas-ligand), аутофагии LС3b, пролиферации Ki-67 и ангиогенеза VEGF. При статистической обработке данных нормальность распределения показателей в группах проверяли при помощи критерия Шапиро-Уилка. Для сравнения групп использовали критерий Крамера-Уэлча (Т). Результаты. При гистологическом исследовании в ткани опухоли под действием экстракта аврана наблюдали появление обширных зон повреждения (некроз и апоптозные тельца). При обоих путях введения экстракта аврана отмечали резкое снижение экспрессии маркеров пролиферации (Ki-67), ангиогенеза (VEGF) и высокую экспрессию маркеров апоптоза (p53, bax, CD95 (Fas/APO-1), FAS-ligand) в опухолевых клетках и отсутствие ее в группе сравнения. Все описанные изменения были более выражены при внутримышечном введении. Экспрессия маркера аутофагии LC3B увеличивалась при пероральном и уменьшалась при внутримышечном введении экстракта аврана. Заключение. Под влиянием экстракта аврана развивается выраженный патоморфоз рака почки. Происходят подавление пролиферации, ангиогенеза и активация сигнального и митохондриального пути апоптоза, а также блокирование протекторной аутофагии. Маркер аутофагии LC3b может использоваться в качестве дополнительного критерия оценки лечебного патоморфоза опухолей.

Objective ― The discovery of the apoptosis-inducing effects of flavonoid vagonin allowed to make an assumption of existence of similar effect in others flavonoids. This study is devoted to the effects of Gratīola officinālis extract on cell culture of the human kidney cancer. Methods ― Cell cultures of human kidney carcinoma – Caki-1 and SN12c were used in the study. The cells were stained with Hoechat 33258 dye. The number of living cells, cells in a state of apoptosis and mitosis were counted. The Cramer-Welch criterion (T) was used to compare the obtained data. Results ― The activation of apoptosis was noted at all concentrations of the Gratiola officinalis L. extract during the first day of exposure. The apoptotic activity increased with increasing of extract concentration. After 48 hours, this activity was maintained only at a Gratiola officinalis L. extract concentration of 0.9 mg/ml. After 24 hours, the apoptotic activity of the extract was more expressed in the culture of CaKi-1. However, after 48 hours the extract induces more pronounced apoptosis in the culture of Sn12c cells. The cytotoxic activity of the extract was not differ after 24 hours in both cultures, after 48 hours it was more pronounced in the culture of CaKi-1. Conclusion ― We revealed a pronounced antitumor and apoptotic activity of the Gratiola officinalis L. extract against the cultures of the kidney cancer Caki-1 and Sn12c. Apoptosis of tumor cells can be manifested in the form of pycnosis of the nucleus, the formation of apoptotic bodies and the emergence of cellular debris resulting from complete degradation of tumor cells. The apoptotic activity of the Gratiola officinalis extract depends on their concentration.

Upconversion nanoparticles (UCNPs) are of interest as novel luminescent probes for numerous applications in nanobiophotonics. The small size of UCNPs enables the particles to overcome biological barriers, thereby ensuring a deep penetration into the tissues and accumulation in a number of organs. In addition, particles are known to possess high surface chemical reactivity as well as a large surface-tovolume ratio, which affect their biocompatibility. In this paper, we present data of the dark toxicity of uncoated UCNPs in the course of their interaction with tissues and organs. It was found inflammatory reaction that developing in the organism when UCNPs are introduced under the skin. The signs of damage and necrosis in all layers of the skin were not observed. The dense connective tissue capsule was formed around the particles through a week after their administration. The particles do not diffuse and dissolve, but remain at the introduction site. UCNPs cause an inflammatory reaction in skin, in a week the skin site, where particles were administrated, is almost not seen. The changes in blood supply and blood circulation in the organs were developed in 2 days mainly. This in turn led to insignificant dystrophic changes. All changes were reversible and disappear or their degree of severity decreases after 7 days after the administration of the particles. The particles enter the total blood flow, but they did not show a pronounced toxicity.

This study focuses on the effect of the flavonoid-containing Gratiola officinalis L. extract with antitumor activity on the intensity of peroxidation and the content of vitamin E in the blood serum of animals with transplanted liver cancer PC-1. Intramuscular and oral administrations of the Gratiola officinalis extract in a dose of 110 mg/kg reduce MDA concentration (more than 20 times) and lipid hydroperoxide (more than 1.5 times) in rats with transplanted tumors. This effect leads to decrease in intensity of lipid peroxidation processes in animals. The Gratiola officinalis extract administration increases the vitamin E concentration (more than 1.3 times) in the serum of rats. This result enables to suggest that the extract of Gratiola officinalis contains the tocopherols. Thus, the study of mechanisms of the Gratiola officinalis extract influence on the activity of peroxidation processes and on the activation of the antioxidant system is promising.

Influence of flavonoids on the intensity of peroxidation processes in the blood is investigated by numerical modeling and by experiment in vivo. As an example we consider the effects of flavonoid-containing extract of Helichrysum arenarium L. with antitumor activity on serum of rats with transplanted liver cancer PC-1. It was found that the content of malondialdehyde, lipid hydroperoxides and average mass molecules were decreased in animals with transplanted liver cancer after intramuscular and oral administration of Helichrysum arenarium L extract in a dose of 1000 mg/mL. The extract reduces the intensity of lipid peroxidation processes in animals. The compound formation possibility of flavonoids and products of lipid peroxidation is investigated by numerical simulations. Using the density functional theory method of molecular modeling, we analyze hydrogen bonds formation and their influence on IR - spectra and structure of molecular complex which is formed due to interaction between flavonoids and products of lipid peroxidation processes on example of naringine and malondialdehyde. We have found that naringine can form a steady molecular complex with malondialdehyde by hydrogen bonds formation. Thus, the application of Helichrysum arenarium L. extract for suppression processes of lipid peroxidation and activation of enzymatic and non-enzymatic antioxidant systems is promising.

In the present work the effectiveness of antioxidants quercetine (a pure chemical) and Gratiola officinalis extract, which is obtained by a new method of extraction from plant material, is investigated on the model of photodynamic haemolysis that is a rather convenient method to monitor the rate of cell membranes oxidative destruction. The effect of these antioxidants on the rate of photodynamic haemolysis is considered as a measure of membranoprotective efficiency.

The data of local microbiologic monitoring was used to study the profiles and mechanisms of resistance to beta-lactams antibiotics of gram-negative isolates of bacteria, pathogens of hospital-acquired infections in hospital reanimation and surgical departments. The study included 210 clinical isolates of pathogens of hospital-acquired infections: Pseudomonas aeruginosa--86 (40.9%), Acinetobacter baumannii--45 (21.4%), Klebsiella pneumoniae--52 (24.8%), Escherichia coli-23 (11.0%), Enterobacter spp.--4 (1.9%). The profiles of resistance to antibiotics were analyzed using the technique of serial micro-dilutions. The detection of the most common and clinically significant gens of beta-lactamases of gram-negative bacteria was implemented using the PCR technique and sequence analysis. The most activity was detected among carbapenems and cephaperazone/sulbactam. The local characteristics of prevalence of gens coding beta-lactamases (TEM, SHV, CTX) in K. pneumoniae and E. coli were established The study detected 11 isolates of P aeruginosa resistant to carbapenems and having genetic determinants of VIM-group and coding metal-beta-lactamases. The discussed data permits to assess the indicators of resistance to beta-lactams antibiotics and basic mechanisms of resistance of causative agents of hospital-acquired infections. The studies of this kind are unique for every type of hospital-acquired infection. The results can be used in the development of the concept of etiotropic and empiric therapy.

A significant increase of resistance in out-of-hospital and hospital pathogens to antimicrobials is being lately observed all over the world, including the Russian Federation. Within 2005-2007 clinical strains of potential producers of extended spectrum beta-lactamases were isolated in diverse medical centres. The isolates suspected to produce extended spectrum beta-lactamases by the criteria of CLSI (Clinical and Laboratory Standards Institute, USA) were included into the test with the polymerase chain reaction and sequence for detecting the genes of beta-lactamases TEM, SHV, CTX, OXA and MBL by the original primers. The total of 99 isolates of gramnegative bacteria suspected to produce extended spectrum beta-lactamases by the findings of the local laboratories were tested at the National Research Centre of Antibiotics: 44 strains of the family Enterobacteriaceae, 38 strains of P. aeruginosa and 17 strains of A. baumanii. The Enterobacteriaceae isolates were mainly multiple resistant (29.5%) carrying simultaneously the genes of 4 groups of beta-lactamases (TEM, SHV, CTX and OXA). The investigation revealed 1 strain of E. coli having the gene of metal-beta-lactamase VIM. The strains of A. baumanii mainly contained the genes of beta-lactamases OXA (23%). Group CTX-M1 prevailed (95%) among the detected genes CTX, 2 strains of group CTX-M9 were also isolated. The mechanisms of antibiotic resistance were not elucidated in 35 strains of P. aeruginosa (92%), gene TEM being detected only in 1 strain and gene MBL (metal-beta-lactamase) being detected in 2 strains. The prevalence of multiple resistant strains carrying the beta-lactamase genes OXA and TEM in the medical centres was shown. The most negative fact was isolation of the E. coli strain producing metal-beta-lactamase.

The method of chicken reovirus strain differentiation was worked out on the basis of RT-PCR and heteroduplex mobility assay (HMA). The S3 gene cDNA (633-896 b.p.) of some Russian and Italian chicken reovirus isolates was amplified by RT-PCR. The analysis of these cDNA samples was carried out by HMA. The relation between nucleotide differences and relative mobility of compared cDNA heteroduplex was reflected by the regression curve. The equation of linear regression was derived (y = 91.726-0.89x; where y is the level of nucleotide difference of compared cDNA (%), x is the relative mobility of compared cDNA heteroduplex (%)). This method made it possible to take correct results within 5-35% of nucleotide difference in heteroduplex sequences.