Objective: to study the cellular mechanisms of activation of regenerative processes in the liver when using total RNA (tRNA) of bone marrow cells (BMCs) based on an extended liver resection (ELR) model. Materials and methods. Male Wistar rats (n = 80) with ELR model (70%) were divided into 2 groups: group 1 (control group) had a single saline injection, while group 2 (experimental group) received a single tRNA injection at a 30 μg/100 g dose of animal weight. The biochemical parameters of liver function and weight were monitored over time. Also monitored were microstructural changes in hepatocytes 48 hours after ELR by examining mitotic activity, caspase-9 expression and morphometric parameters. Results. It was found that in group 2, in comparison to group 1, there was faster normalization of biochemical parameters (by 10–14 days), a higher mitotic index of hepatocytes (23.45‰ versus 5.37‰), and initially sharper decrease and then faster recovery of liver mass (by 10–12 days versus 18–20 days). Both groups showed almost total expression of caspase-9, including in mitotically splitting hepatocytes. Group 1 demonstrated decreased values of morphometric parameters of single and binuclear cells, decreased number of binucleated hepatocytes and increased total density of hepatocytes as compared to the intact liver. Intraperitoneal administration of tRNA increased morphometric parameters of mononuclear hepatocytes, did not affect their number, but increased the area of the nuclei of binuclear hepatocytes as compared to the control group. Conclusion. The proven capability of cell-bone marrow total RNA to simultaneously support apoptosis in liver cells after ELR and induce mitotic activity indicates that tRNA can switch activated apoptosis to cell proliferation at the early phase of the regenerative process. This effect may be due to the presence of regulatory RNA molecules in tRNA, including numerous non-coding RNAs.

Aim : to conduct a comparative assessment of the effectiveness of liver regeneration occurring after induction of chronic fibrosing liver disease (CFLD) using bone marrow mononuclear cells (BMMCs) and total RNA (tRNA) extracted from BMMCs. Materials and methods . The study involved 140 Wistar rats. CFLD was modeled in 100 rats, of which 25 died. The surviving 75 rats (CFLD formed by the third month) were divided into 3 groups: Group 1 – control (administered with physiological saline); Group 2 – a single injection of tRNA from BMMCs at a dose of 30 μg/100g body weight; Group 3 – a single injection of BMMCs at a dose of (30–35) × 106 cells. The dynamics of regenerative processes in the liver was evaluated based on the animal mortality, dynamics of restoration of biochemical markers (ALAT, ASAT, alkaline phosphatase and total protein) and morphological picture of the liver on the seventh day and after three, six and nine months. The significance of differences in the compared values was determined through Student’s t-test for <0.05. Results . Mortality in Group 1 was 12%, in Groups 2 and 3 – 4%; In Group 1, ALAT and ASAT were restored to normal values after two months, alkaline phosphatase after 3 months, and total protein remained low for over 4 months. In Groups 2 and 3, all hepatic homeostasis markers returned to the values they were before CFLD modeling faster than in Group 1 (after two months). However, in Group 2, the regeneration rate was higher than in Group 3. It was revealed that normalization of functional liver parameters in all groups were ahead of restoration of the histological structure of the liver. Liver defibrotic processes in Group 2 were activated after 3 months, and in Groups 1 and 3 – after 6 months. The histological structure of the liver was restored in Group 2 after 6 months, and in Groups 1 and 3 after 9 months. Conclusion . BMMCs and tRNA extracted from them in biologically effective doses trigger liver regeneration in CFLD. However, regulatory effect from the use of tRNA appears earlier and is more effective.

Aim: to determinate the most effective liver cells and multipotent mesenchymal stromal cells of bone marrow (MMSC BM) ratio into implantable cell engineering constructions (CECs) used for chronic liver failure (CLF) correcting. Materials and methods. For creating liver CECs it was used a biopolymer implant – a composition of a heterogeneous collagen-containing gel (BMCG) (Sphero®GEL trademark) containing viable liver cells and MMSC BM in the following ratios – 1 : 1; 5 : 1 and 10 : 1 respectively. CECs with different ratios of liver cells and MMSC BM were implanted into liver of rats in which chronic liver failure (CLF), was modeled by using CCl4. The effectiveness of the regulatory effects of CECs (with different cell ratios) on regenerative processes in livers were assessed by using biochemical, morphological and morphometric methods at different periods after their implantation. Results. Corrective effect of CECs with different cell composition on biochemical and morphological parameters of livers at chronic liver failure was established. During studying the liver CECs with various cell ratios of liver cells and MMSC BM (1 : 1; 5 : 1 and 10 : 1 respectively), it was found that the most optimal ratio of cells into the CECs is 5 : 1, because at this ratio of cells, there were a more distinct normalization of the morphological and functional liver parameters within 365 days after modeling CLF and maintenance of the structural homeostasis into the CECs. Themselves, which allows predicting their long-term regulatory effect on the liver tissue in CLF and maintaining its normal structural and functional state. Conclusion. The effective correction of chronic liver failure can be carried out by using the implanted liver CECs, in which donor liver cells and MMSC BM where presented in ratios – 1 : 1; 5 : 1 and 10 : 1. But analysis of prolonged correction of liver morphological and functional parameters at CECs using it was allow to recommend the preferences using of CECs with ratio 5 : 1, because prolonged preservation of structural homeostasis into these CECs makes possible to prognosticate their prolonged regulatory action on the liver tissue at CLF, especially for recipients on a waiting list for liver transplantation.

Разработана методика определения анилокаина в плазме крови крыс методом ВЭЖХ c УФ-детектированием. Пробоподготовку проводили жидкость-жидкостной экстракцией смесью 1-бутанол — гексан (2:98 об./об.) и последующей реэкстракцией 0,1 % водным раствором муравьиной кислоты. Хроматографический анализ осуществляли в изократическом режиме на колонке Nucleodur HILIC с подвижной фазой ацетонитрил — 25 мМ раствор формиата аммония (85:15) при длине волны 205 нм. Методика валидирована по следующим параметрам: специфичность, эффект переноса, линейность, прецизионность, правильность, пределы количественного определения, допустимость разбавления, стабильность. Аналитический диапазон методики составил 25 – 1000 нг/мл. Разработанная методика может быть применена для исследования фармакокинетики препаратов анилокаина.

This review briefl y discusses the experience of organizing and carrying out experimental studies in our center in order to increase the validity of the researches. The rules of organizational and methodological approaches aimed at increasing the suitability of the results of experimental studies are also given. Each research work in which experiments on laboratory animals are supposed to be carried out should be organized, planned and carried out in accordance with national and international ethical standards. Discussion and publication of both planned and conducted experimental work makes it possible to make the research process more open and objective. The experimental investigation is carried out in the following algorithm: the formulation of the problem and the hypothesis on the basis of the literature data; development of the purpose and objectives of the study with adequate methods and selection of equipment; distribution of material to control and trial groups; creation of a plan for monitoring the indicators during the experiment; processing and interpretation of results; preparation of a scientifi c report.

Разработана методика определения ацетилсалициловой кислоты и ее основного метаболита — салициловой кислоты в модельном растворе и плазме крови кроликов методом ВЭЖХ с УФ-детектированием. Пробоподготовку плазмы проводили методом высаливания. Хроматографический анализ осуществляли в изократическом режиме на колонке Hypersil BDS C18 с подвижной фазой ацетонитрил — вода (pH 2,5, 30:70) при длине волны 230 нм. Предел количественного определения ацетилсалициловой и салициловой кислот в модельном растворе составил 0,05 мкг/мл, в плазме крови — 0,2 мкг/мл. Разработанная методика применена для изучения новых форм ацетилсалициловой кислоты на основе биосовместимых полимерных носителей, в том числе для изучения фармакокинетики после внутримышечной имплантации.

Разработана методика определения антидота угарного газа ацизола в крови человека методом ВЭЖХ на колонке HILIC с УФ-детектированием после осаждения белков ацетонитрилом. Хроматографический анализ проводили в изократическом режиме на колонке Nucleodur HILIC с подвижной фазой ацетонитрил — вода (95:5) при длине волны 225 нм. Нижний предел количественного определения составил 116,2 нг/мл, верхний предел — 11111 нг/мл. Валидация методики подтвердила соответствие ее характеристик (специфичность, эффект переноса, линейность, пределы количественного определения, прецизионность и правильность, степень извлечения, стабильность) установленным требованиям.

On an experimental model of chronic fibrotic liver damage (male rats Wistar (n=60), damage of CCl 4 , the duration of the experiment 90 days) it was studied the effectiveness of cell therapy for the correction of chronic liver failure. These rats were divided into 3 experimental groups: in the I st group (control, n=10) isotonic saline (650 mkl.) was injected; in the II nd group (n=20) suspension of liver cells was applicated in a dose 8-10×10 6 cells; in the III rd group (n=30) suspension of liver cells and bone marrow cells (mesenchymal stromal cells) in ratio 5:1 were used as cell associates on microparticles injectable heterogeneous biopolymer hydrogel «SpheroGEL» (cell-engineering design) in common dose 8-10×10 6 cells . It was ascertained that in the 2 nd and in the 3 rd groups the accelerated normalization of disturbed liver functional indices (ALT, AST, ALP) took place – to 30 days, but in the control group only to 90 days. The reliable differences in rats of normalization of functional indices were absent between the II nd and the III rd groups. But in 90 days by using special histological dyeing it was found out that defibrotic processes in liver tissue were more expressed in the III rd group in comparison with the III nd group. Received results were consequence of prolonged vital activity of cells (liver cells and mesenchymal stromal bone marrow cells) into cell-engineering designs, which were transplanted in the III rd group. The obtained effect can be explained by that the developed cell-engineering designs provide adequate conditions for prolonged vital activity of the transplanted cells.