Introduction. One of the stages in the development of medicinal products (MP) is the study of safety assessment. Determination of possible toxic effects, establishment of the nature and severity of the damaging effect of drugs under development during their administration is carried out, including studying the effect on hematologic parameters and hematopoiesis. Aim. To assess the possibilities of PEG-HYAL impact on hematologic parameters and hematopoiesis when administered per os. Material and methods. Hyaluronidase immobilized by electron beam synthesis on polyethylene oxide (PEG-HYAL). Laboratory animals – white mature conventional outbred rats and chinchilla rabbits. Hematological parameters were determined using an automatic hematological analyzer Abacus (Diatron, Austria) and conventional manual methods of research. The effect of PEG-HYAL on the state of medullary hematopoiesis was determined by counting the total number of myelokaryocytes per femur (106/femur) and myelograms on smears (in rats). The percentage of individual cell forms in rat myelogram counts was converted to absolute numbers, x106 cells per femur. In rabbits, the myelogram indices of bone marrow, taken from the sternum segment, were determined in relative units (%) only. Results. The analysis of the obtained results of the effect of PEG-HYAL administration on hematologic indices in rats (male and female) did not show any pathological changes. The revealed changes in hematologic parameters in rats are neither significant, nor systemic and dose-dependent. PEG-HYAL administration to rabbits has no toxic effect on hematologic parameters with multiple intragastric administration. Administration of PEG-HYAL for 28 days to rats in all studied doses does not produce a toxic effect on the main studied bone marrow parameters. While the revealed changes in the quantitative composition of myelokaryocytes are non-systemic and reversible. PEG-HYAL administration has no toxic effect on bone marrow indices of rabbits when administered intragastrically in the studied doses during 28 days. Conclusion. Interpretation of the obtained results, concerning the effect of the per os PEG-HYAL administration to experimental animals on hematologic parameters and hematopoiesis, can be used as a preclinical dossier of the drug.

Introduction. Testicular hyaluronidase preparations are widely used in medical practice. Modification of native biologically active molecules by electron beam PEGylation allows to improve their pharmacokinetic properties, which determines a subsequent improvement of pharmacodynamic effects. The development of an original oral drug with a pharmacologically active core, which is hyaluronidase, is promising, but requires studying the safety of use at the preclinical stage. Aim. Ultrastructural evaluation of hepatotoxic effects of PEGylated hyaluronidase in vitro. Materials and methods. The object of a study was testicular hyaluronidase PEGylated on polyethylene oxide (PEG-HYAL) using electron beam synthesis. The culture medium is a continous culture of human liver cells Chanq liver. The cytotoxic effect was detected by the MTT assay. Ultrastructural changes were evaluated by electron microscopy. Results. The original drug PEG-HYAL in the studied concentrations of 37, 75 and 150 U/ml has no cytotoxic effect on the human liver cell culture (hepatocytes). Cell viability is virtually at the control level when PEG-HYAL administration at concentrations of 37 and 75 U/ml, and in the maximum concentration of 150 U/ml, cell proliferation is stimulated significantly, as evidenced by an increase in cell viability up to 106%. The results of evaluation of ultrastructural changes in hepatocytes showed that exposure to PEG-HYAL in all concentrations leads to the enhancement of metabolic processes in cells, development of autophagy, which is one of the main homeostatic processes. Conclusion. The drug PEG-HYAL in all studied concentrations has no toxic effect on the human hepatocyte culture. The results obtained can be used in further research in the development of PEG-HYAL.

The widespread use of testicular hyaluronidase preparations determines the particular interest in the development of oral dosage form. The drug under development, which is hyaluronidase pegylated using electron beam immobilization technology (PEG-GIAL), requires an assessment of its safety. Identification of possible neurotoxic action, as well as the influence on cardiac function seems to be very relevant. Aim of the study was to investigate the effect of PEG- GIAL on behavioural reactions and electrical activity of the heart of experimental animals. Material and methods. Eight hundred of white outbred rats (male and female) were used as experimental animals. The effect of PEG-GIAL on the central nervous system was determined using the «open field with holes» test, examining such characteristics as emotional reactions and orientation-research behaviour. The functional state of the cardiovascular system was assessed by electrocardiography data. Results. PEG-GIAL administration in all studied doses does not lead to changes in the functional activity of CNS, does not have a pronounced effect on the indices of orientation-research behaviour and emotional reactions in laboratory animals. No statistically significant changes in the cardiogram of rats were revealed, including no intersex differences. Conclusions. PEG-GIAL administration in all studied doses has no toxic effect on the electrical activity of the heart and has no pronounced effect on changes in the functional activity of the CNS and behavioural reactions.

The features of the participation of Smad3 in the functioning of neural stem cells (NSC), neuronal committed precursors (NCP), and neuroglial elements were studied in vitro. It was found that this intracellular signaling molecule enhances the clonogenic and proliferative activities of NCP and inhibits specialization of neuronal precursors. At the same time, Smad3 does not participate in the realization of the growth potential of NSC. With regard to the secretory function (production of neurotrophic growth factors) of neuroglial cells, the stimulating role of Smad3-mediated signaling was shown. These results indicate the promise of studying the possibility of using Smad3 as a fundamentally new target for neuroregenerative agents.

Comparative study of phenolic compound composition was performed by HPLC-UV and HPLC-MS techniques using raw materials of Hedysarum alpinum L. plants introduced into the dendrological ecosystem of the Siberian Botanical Garden of Tomsk State University as well as the callus culture of the same species. The callus culture of Hedysarum alpinum was obtained on Murashige and Skoog nutrient medium with the addition of 1.0 mg/L 2,4-D and 0.1 mg/L 6-benzylaminopurine (6-BAP) and was subsequently cultivated on the medium containing α-NAA (2.0 mg/L) and 6-BAP (0.5 mg/L). The culture was characterized by activated phenylpropanoid pathway, which gave rise to the formation of flavonoids, isoflavones, xanthones, and other substances. The total content of flavonoids (calculated on a per quercetin basis) and formononetin in the extracts of callus culture was lower than in the extracts from introduced plants by 35 and 11

We studied the effectiveness of Xe/O2 mixture inhalation (30% Xe and 70% O2, 20 min for 5 days) in a model of experimental thromboplastin pneumonitis. Inhalation of the studied mixture decreased the intensity of the inflammatory process in the lung tissue assessed by the temperature response of animals, changed lung weight and lung weight coefficient. At acute stage of pneumonitis, an increase in xenon consumption was recorded due to its retention in the gas exchange zone and a natural decrease in oxygen consumption due to partial alveolar/capillary block. The formation of pneumonitis was accompanied by a pronounced procoagulant shift in the regulation system of the aggregate state of blood. The Xe/O2 inhalations ensured physiologically optimal levels of prothrombin and activated partial thromboplastin time against the background of a moderate decrease in fibrinogen level throughout the experiment. At the same time, the activity of the natural anticoagulant antithrombin III increased from day 5 to day 14.

The duration and severity of prothrombotic effects of the maximum tolerated dose of paclitaxel (40 mg/kg) were evaluated in intact outbred mice. Hemostasis was assessed before and on days 1, 2, 5, 7, 10, 15, 20, and 30 after a single injection of paclitaxel using standard coagulation tests (activated partial thromboplastin time, prothrombin time, fibrinogen concentration, and antithrombin III) and a “global” method, low-frequency piezothromboelastography. A pronounced prothrombotic effect of paclitaxel was revealed starting from the first day postinjection that consisted in intensification of fibrinogenesis up to the 7th day in parallel with activation of the anticoagulant mechanisms. On days 7-30 after paclitaxel administration, decompensation of its anticoagulant activity due to paclitaxel-induced damage to the endothelium was observed with the formation of a procoagulant status of the hemostatic potential of the blood. A single administration of the maximum tolerated dose of paclitaxel forms a powerful thrombogenic stimulus during the first week and provides a long-term/trace procoagulant shift in the hemostasis system (days 10-30).

The effects of inhalations of an oxygen-xenon (70%/30%) mixture were studied in two models of acute respiratory distress syndrome caused by intratracheal administration of 0.5 mg/kg LPS or 0.04 ml acidin-pepsin (pH 1.2). Inhalation of the oxygen-xenon mixture inhibited the development and reduced the intensity of the inflammatory process in the lung tissue, which was assessed by the dynamics of lung weight and body weight of animals: the therapeutic exposure decreased both parameters. It was found that the thrombogenic stimulus, pathognomonic for the development of acute respiratory distress syndrome, decreased under the effect of oxygen-xenon inhalations, while the level of natural anticoagulant antithrombin III increased.

We studied the effect of an anthocyanin-containing complex from the fruits of S. aucuparia L. on doxorubicin-induced genotoxicity in bone marrow cells of C57BL/6 mice. The complex reduced the genotoxic effect doxorubicin in metaphase plates of bone marrow cells in 24, 48 h, and 10 days after the administration of the cytostatic. The mean number of single fragments and the fraction of cells with gaps and aberrant metaphases also decreased.

The development of safe drugs occupies a special place in the pharmaceutical industry. One of the main tasks of its preclinical phase is to evaluate possible toxic effects of the developed drug on the body and on various systems, including the immune system. The aim of our work was to study immunotoxic properties of pegylated hyaluronidase (PEG- HYAL). Material and methods. Mice F1(CBA/C57Bl/6) were divided into subgroups which were intragastric and intraperitoneally administered with PEG-HYAL in different dosages (50, 500, 1250, 2500 and 5000 U/kg). The number of antibody-producing cells in the spleen, the mass and cellularity of central and peripheral immune organs, phagocytic activity of peritoneal macrophages and neutrophils, delayed hypersensitivity reaction (DHR), level of hemagglutinin to sheep erythrocytes, spontaneous and mitogen-induced splenocyte proliferation were determined. Results. PEG-HYAL did not induce DHR, did not suppress phagocyte activity of peritoneal macrophages, at a dose of 50 ED/kg did not significantly affect the hemagglutinin content to erythrocytes, but at a dose of 500 ED/kg did statistically significantly reduce the titer of specific antibodies. When experimental animals were exposed to PEG-HYAL at doses of 50 and 500 U/kg, spontaneous and mitogen-induced proliferation of splenocytes decreased. Conclusions. The PEG-HYAL trial produced results that can be used to substantiate the administration of the PEG-HYAL-based medication.

Anthocyanins are flavonoid compounds belonging to the group of polyphenols. A. melanocarpa ( Michx. ) Elliott chokeberry is known to be rich in these bioactive substances. The previously conducted chemical analysis showed that an anthocyanin-containing complex obtained from A. melanocarpa fruits comprise anthocyanins, flavonoids, phenolic acids, and catechins, with anthocyanins being the dominant components. A large amount of data indicates that Aronia fruits exhibit a wide spectrum of pharmacological activity. In this work, we assess the safety of an anthocyanin-containing complex obtained from A. melanocarpa fruits by its genotoxic study followed by an analysis of its effect on mutagenesis. To this end, a model of doxorubicin-induced genotoxicity in bone marrow cells of C57Bl/6 mice was used. The plant complex under study at a dose of 225 mg/kg had no effect the cytogenetic parameters of animal bone marrow cells after a single or double administration. The use of the anthocyanin-containing complex led to a decrease in DNA damage caused by the administration of doxorubicin, 24 and 48 hours after the introduction of a cytostatic agent. Hence, the data obtained can serve as the basis for the creation of a drug corrector for cycplasms.

The study aims to analyze furocoumarin extracts isolated from poison hemlock cell culture influencing the correction of cisplatin-caused cytostatic myelosuppression. The experiments were carried out on 160 CD1 female mice weighing 18–25 g. Cytostatic myelosuppression was simulated by administering cisplatin intraperitoneally once at a maximum tolerated dose of 10 mg/kg. The control group received physiological saline injections. Poison hemlock (Conium maculatum L.) cell culture extract was administered at a dose of 30 mg/kg to prevent the development of disorders. 4 mg/kg of Warfarin Nycomed was used for comparison examination. The correctors were administered intragastrically on the sixth day after cisplatin injected for four days. The indicators were examined on the 1st, 2nd, 5th, 7th, 10th, 15th, 20th and 30th days following the administration of cisplatin. The bone marrow and peripheral blood values were examined using the standard method. Statistical analysis was carried out using Stat Plus Pro (build 7.3.0.0). Under conditions of cytostatic myelosuppression caused by the administration of 10 mg/kg (maximum tolerated dose) of cisplatin, the use of the poison hemlock cell culture extract contributes to the restoration of myeloid and erythroid hematopoietic germs, as well as the normalization of bone marrow and peripheral blood values. With increase in the number of mature neutrophils and lymphocytes, erythroblasts, and normoblasts, the total number of myelokaryocytes increased. The content of erythrocytes and leukocytes increased in the peripheral blood as the number of segmented neutrophils and lymphocytes increased. The dynamics of hematopoietic sprout recovery with poison hemlock cell culture extract are similar to those of Warfarin.

ЭКСПЕРИМЕНТАЛЬНОЕ ОБОСНОВАНИЕ ПРИМЕНЕНИЯ ПЕРОРАЛЬНОГО ТРОМБОЛИТИКА НА ОСНОВЕ ИММОБИЛИЗИРОВАННЫХ СУБТИЛИЗИНОВ И АНТИКОАГУЛЯНТА НА ОСНОВЕ НИЗКОМОЛЕКУЛЯРНОГО ГЕПАРИНА ДЛЯ ЛЕЧЕНИЯ ТРОМБОЗА СОСУДОВ ЛЕГКИХ

The effects of JAK and STAT3 inhibitors on the production of neurotrophic growth factors by different types of neuroglial cells were studied under conditions of in vitro and in vivo models of ethanol-induced neurodegeneration. It was shown that these signaling molecules do not participate in the secretion of neurotrophins by intact astrocytes and oligodendrocytes. The inhibitory role of JAK in the regulation of this function of microglial cells was revealed. We also revealed significant changes in the role of JAK and the presence of STAT3 specifics within the framework of JAK/STAT signaling in the production of growth factors by various glial elements under the influence of ethanol. Neurodegeneration modeled in vitro led to the appearance of a “negative” effect of STAT3 on the production of neurogenesis stimulants by all types of glial cells. Moreover, the role of STAT3 in oligodendrocytes and microglial cells generally corresponded to that of JAK/STAT signaling. In astrocytes, only selective blockade of STAT3 (but not JAK) led to stimulation of their function. In mice subjected to prolonged peroral alcoholization, the neuroglial responses to the pharmacological regulation of JAK/STAT signaling were different. An inversion of the role of JAK and STAT3 in the production of neurotrophins by oligodendrocytes was noted. In addition, JAK inhibitor did not stimulate secretory function of microglial cells under conditions of prolonged exposure to ethanol in vivo.

The psychopharmacological effects of a stimulator of functions of progenitor cells of the nervous tissue STAT3 inhibitor (STAT3 Inhibitor XIV, LLL12) were studied under conditions of modeled alcoholic encephalopathy in C57BL/6 mice. The pharmacological agent corrected the parameters of exploratory behavior (characterizing predominantly cognitive activity) in the experimental animals at the late terms of observation. At the same time, the reproducibility of the conditioned passive avoidance response developed at the beginning of the course STAT3 inhibitor administration decreased. These effects developed against the background of a significant increase in the content of neural stem cells and their proliferative activity in the paraventricular zone of the brain.

We studied the participation of ERK1/2 and p38 in secretion of neurotrophic growth factors by various types of neuroglia under conditions of in vitro and in vivo modeled ethanol-induced neurodegeneration. The inhibitory role of these protein kinases in the production of neurotrophins by intact astrocytes and the absence of their participation in the regulation of functions of oligodendrocytes and microglial cells were shown. Under conditions of ethanol neurotoxicity, the role of ERK1/2 and p38 in the production of growth factors by glial elements was significantly changed. Neurodegeneration modeled in vitro led to inversion of the role of both protein kinases in the secretion of neurotrophins by astroglia and inhibition of the cytokine-synthesizing function of oligodendrocytes and microglial cells by ERK1/2 and p38. In mice receiving ethanol per os for a long time (as well as in cells in vitro exposed to ethanol), mitogen-activated kinases stimulated the function of astrocytes and inhibited the production of growth factors by microglial cells. At the same time, chronic alcoholization was accompanied by the appearance of the stimulating role of ERK1/2 and p38 in the implementation of the secretory function by oligodendrocytes.

The purpose of the study was to evaluate in vitro cytotoxicity and in vivo sub-acute toxicity of conditioned medium obtained from rat bone marrow-derived mesenchymal stem cells directed to osteogenic differentiation. Material and methods . Conditioned medium was obtained by culturing rat bone marrow-derived MSCs induced under osteogenic condition. Results and discussion . Conditioned medium from rat bone marrow-derived MSCs was shown to have no significant cytotoxic effect on Ehrlich adenocarcinoma cell culture and in vitro expanded peripheral blood mononuclear cells from healthy donors. The use of MSC-CM did not have a significant effect on the state of experimental SD rats, the total body weight and growth rate of animals. A pathomorphologic study revealed no any abnormalities associated with MSC-CM injection. Conclusion . The use of conditioned medium from rat bone marrow-derived mesenchymal stem cells stimulated under osteogenic condition was found to be safe for both in vitro studies (with respect to Ehrlich adenocarcinoma cell line and peripheral blood mononuclear cells from healthy donors) and in vivo studies (injection of MSC-CM to animals).

The genotoxic effect of paclitaxel on the chromosomal material of differentiated bone marrow cells in male and female mice was revealed in the early and long-term periods of the study. It was found that during the same observation period paclitaxel causes bone marrow hypoplasia and reduces the number of early erythropoiesis progenitor cells in the bone marrow of experimental animals, and also contributes to a decrease in their proliferative potential regardless of gender.

Выявлено генотоксичное воздействие паклитаксела на хромосомный материал дифференцированных клеток костного мозга самцов и самок мышей в ранние и отдаленные сроки исследования. Установлено, что в эти же сроки наблюдения паклитаксел вызывает гипоплазию костного мозга и снижает в костном мозге экспериментальных животных количество ранних клеток-предшественников эритропоэза, а также способствует уменьшению их пролиферативного потенциала независимо от пола. The genotoxic effect of paclitaxel on the chromosomal material of differentiated bone marrow cells in male and female mice was revealed in the early and long-term periods of the study. It was found that during the same observation period paclitaxel causes bone marrow hypoplasia and reduces the number of early erythropoiesis progenitor cells in the bone marrow of experimental animals, and also contributes to a decrease in their proliferative potential regardless of gender.