Metabolic syndrome (MS) is one of the most common socially significant diseases. Around 1.9 billion people suffer from this disease, which places an enormous burden on healthcare systems around the world. This is particularly true in connection with concomitant diseases. With the progression of MS, disorders in the function of the immune system occur in the body, including those associated with mitochondrial dysfunction, leading to the development of chronic inflammation. In particular, there is an increase in the number of circulating monocytes actively recruited to inflamed adipose tissue, where there is non-specific proinflammatory activation of innate immune cells, which adopt an M1-like phenotype and become less sensitive to anti-inflammatory stimuli. This ultimately leads to a decrease in the functional activity of monocytes/macrophages and their immunoplasticity. The subject of the study was the venous blood of patients and the CD14+ monocytes/macrophages obtained from it by immunomagnetic separation. In our work, we focused on the search for significant relationships between markers of chronic inflammation and the formation of immune tolerance of monocytes/macrophages in patients with metabolic syndrome. Biochemical parameters, basal and LPS-stimulated production of proinflammatory cytokines (IL-6 and MCP-1) were investigated by culture of monocytes/macrophages in patients with metabolic syndrome. The evaluation of biochemical parameters in blood samples from MS patients and healthy donors revealed that the levels of ALAT, AST, GGT, alkaline phosphatase, uric acid, C-reactive protein, glucose and insulin were significantly higher in MS patients than in healthy donors. The levels of P-amylase and high-density lipoprotein were significantly lower than in the control group. Within the experimental model, repeated stimulation showed a decrease in cytokine production in response to LPS compared to the first stimulation on day 7. It was also found that the response to the primary stimulus was higher in cells from patients with a body mass index (BMI) 40 kg/m2, which could indirectly indicate the presence of a phenotype associated with chronic inflammation and consequently with reduced plasticity of the monocyte/macrophage immune response.

Recently, the role of vitamin imbalance in carcinogenesis has been actively discussed. Studies aimed at assessing their role in the processes of cancer development are various, and the evaluation of the initial level of vitamins is relevant when planning antitumor therapy.Objective. To determine the state of the initial level of B vitamins and homocysteine before chemotherapy in patients of different oncologic profile.Material sand Methods. The level of vitamins B1, B6, B9, active form of B12* and homocysteine in fresh frozen plasma was determined by enzyme[1]linked immunosorbent assay (ELISA) using test systems in 66 patients with verified malignant neoplasms before chemotherapy. The study included 66 patients: women n=40 and men n=26, with cancer: gastric n=12, colorectal cancer n=21, cancer lung n=11, cancer pelvic organs n=22.Results. According to the results of the study the increase in the level of holotranscobalamin (Holo-TC) and decrease in homocysteine in comparison with reference values was observed in 100% of cases. Significant difference was found only for B1: reliable differences between patients with colorectal cancer and with pelvic cancer (in the latter the mean value of B1 levels was 2.4 times higher at p = 0.0425). According to the results of correlation analysis, a weak correlation between B12 and B9 was determined. When comparing the levels of vitamin B12 and homocysteine in patients after surgical treatment and without it, no significant differences were found.Conclusion. Increased Holo-TC levels and decreased homocysteine levels by ELISA are characteristic of all varieties of solid cancer, independent of sex, age, stage and previous surgical intervention. A relative decrease in vitamin B1 is characteristic of colorectal cancer

In the present work, we carried out a comparative analysis of myocardial cytokine profile in patients with coronary heart disease (CHD) and in patients with ischemic cardiomyopathy (ICMP) associated with CHD. The concentrations of 41 cytokines secreted by 24-hour myocardial tissue culture intraoperatively sampled from the right atrial auricle (RAA, control) and peri-infarct left ventricular zone (PZ-LV) were determined by flow fluorimetry using a multiplex test system. The aim was to study in vitro cytokine profile of myocardial cells to search for possible predictors of adverse outcomes of surgical treatment of patients with CHD and ICMP. Myocardial secretion of proinflammatory molecules GM-CSF and IFN-γ increased significantly (up to 78-80 pg/g, p0.05) in patients with ICMP associated with CHD in contrast to zero values in CHD. At the same time, there was a three-fold decrease in the concentration of fractalkin 3 ligand (Flt-3L; FMS-like tyrosine kinase 3 ligand). A decrease in Flt-3L secretion was observed in the PZ-LV in comparison with the RAA. In addition, compared with RAA, concentrations of fibroblast growth factor-2 (FGF-2), platelet-derived growth factor-AB/BB (PDGFAB/BB), interleukins IL-15 and IL-4, and a regulated upon activation, normal T cell expressed and secreted (RANTES; CCL5) were strongly reduced in PZ-LV myocardial tissue culture. Differences in the course of CHD and ICMP are discussed, and possible predictors of surgical treatment risk in patients of the two groups are suggested using correlation and regression analyses. Proinflammatory cytokines (IL-5, IL-6) and chemokines (Flt-3L, IL-8), as well as angiogenesis factors (VEGF) and angiostasis (IP-10), are proposed to be considered as potential markers of adverse outcome of surgical treatment of cardiovascular disease.

Autophagy is required to maintain cellular homeostasis and organ function by selectively ridding cells of potentially toxic proteins, lipids, and organelles. Impaired homeostasis of autophagic processes is associated with metabolic disorders such as obesity and type 2 diabetes mellitus. In obesity, a violation of autophagy in adipose tissue and its inflammation contributes to the formation of type 2 diabetes mellitus. The aim of the study was to analyze the expression of autophagy genes in the adipose tissue of the greater omentum and to search for their relationship with the levels of cytokines of the IL-10 family in blood plasma in obese patients, depending on the presence or absence of type 2 diabetes mellitus. Blood plasma and visceral adipose tissue samples were studied from 347 obese patients with and without type 2 diabetes. A biochemical analysis of the patients' blood was carried out. The level of cytokines was detected by flow fluorometry. Gene expression was determined by real-time PCR, and tissue-specific protein production was determined by immunoblotting. Statistical processing of the results was carried out using GraphPad Prism 9.0.0 software. Plasma levels of IL-10, IL-20, IL-22, IL-28A, and IL-29 are increased in obese patients without type 2 diabetes compared with patients with type 2 diabetes. In patients with type 2 diabetes mellitus, the expression of the SQSTM1_p62 and MAP1LC3B genes in the greater omentum increased compared to patients without it. High plasma levels of IL-22 and IL-26 are associated with the presence of type 2 diabetes mellitus. In patients without type 2 diabetes mellitus, an increase in the level of IL-28A in blood plasma is associated with a decrease in the expression of autophagy genes SQSTM1_p62 and MAP1LC3B in the adipose tissue of the greater omentum.

A number of preclinical and clinical studies have demonstrated the efficiency of mesenchymal stromal cells to serve as an excellent base for a cell-mediated drug delivery system. Cell-based targeted drug delivery has received much attention as a system to facilitate the uptake a nd transfer of active substances to specific organs and tissues with high efficiency. Human mesenchymal stem cells (MSCs) are attracting increased interest as a promising tool for cell-based therapy due to their high proliferative capacity, multi-potency, and anti-inflammatory and immunomodulatory properties. In particular, these cells are potentially suitable for use as encapsulated drug transporters to sites of inflammation. Here, we studied the in vitro effects of incorporating synthetic polymer microcapsules at various microcapsule-to-cell ratios on the morphology, ultrastructure, cytokine profile, and migration ability of human adipose-derived MSCs at various time points post-phagocytosis. The data show that under appropriate conditions, human MSCs can be efficiently loaded with synthesized microcapsules without damaging the cell’s structural integrity with unexpressed cytokine secretion, retained motility, and ability to migrate through 8 μm pores. Thus, the strategy of using human MSCs as a delivery vehicle for transferring microcapsules, containing bioactive material, across the tissue–blood or tumor–blood barriers to facilitate the treatment of stroke, cancer, or inflammatory diseases may open a new therapeutic perspective.

Correct choice of nutrient media for culturing different types of cells in various applications is one of the most important aspects of modern biotechnology, since chemical composition of the culture media largely contains the necessary metabolites to support certain cells’ growth lines outside the body. Jurkat line of human leukemic T-lymphoblast-like cells (hereinafter Jurkat T-cells) is actively used for in vitro modeling of intracellular signaling and activation of normal blood T-lymphocytes mediated by the T-cell receptor/CD3/ CD4 complex in toxicological studies of immune and secretory responses, to test medicinal substances and ions. Also, Jurkat T-cells are widely used for ex vivo testing in immunology, oncology, toxicology, orthopedics, and traumatology. The existing standards and numerous studies are mainly based on short-term in vitro cultivation of Jurkat T-cells in RPMI 1640 nutrient medium. Meanwhile, the issues of long-term maintenance of the growth of Jurkat T-cells culture are poorly presented in the research literature. This study aimed for studying the activity of Jurkat T-cells over 7 to 14 days of in vitro culture and comparing the relative value of RPMI 1640 and αMEM media for the behavior of immunocompetent tumor cells. Using flow cytometry, multiplex analysis, and phase contrast Cell-IQ microscopy, the proportions of living cells and those dying by apoptosis and necrosis, secretion of cytokines and chemokines, and the dynamics of cell biomass propagation were studied. It was found that the αMEM medium in the complete nutrient medium, as compared with RPMI 1640, is more appropriate to in vitro promotion of cell viability (increased proportion of viable cells by 13.5% at the day 14), their secretory ability for 23 из 27 tested biomolecules, shortened adaptation time (на 32%) in culture before growth initiation, 5-fold increase of the Jurkat Т-cell cellularity by the day 7. Potential significance of the chemical components of nutrient media and secreted biomolecules for these results is discussed. As based on the results obtained, we concluded on superior properties of αMEM medium for long-term in vitro cultures of Jurkat T-cells. Consequently, the in vitro testing of medical devices intended for long-term contact with the body, including those for cancer patients, using Jurkat T-cell leukemia line in RPMI 1640 medium, may lead to wrong predictions on their biocompatibility and potential antitumor activity.

The article presents the results of evaluating growth factors, pro – and anti-inflammatory cytokine production by multipotent mesenchymal stem cell cultures under the conditions of co-cultivation with immuno-isolated beta-cells of the pancreas. β-cell transplantation is a minimally invasive therapeutic approach (compared to transplantation of entire pancreas), and it provides better metabolic control with respect to insulin administration. However, when transplanting β-cells, there is always a risk of immune rejection of the grafted cells. It is generally recognized that encapsulation is an effective means of immunological protection against the recipient’s immune system during transplantation. Regulation of the autoimmune response to transplanted cells is crucial for the treatment of type I diabetes mellitus. In recent years, along with replacement of islet cells, much attention has been paid to the use of multipotent mesenchymal stem cells with immunomodulatory and/or immunosuppressive properties, aimed for the correction of diabetes mellitus. Either in vitro and in vivo, they impact not only T-lymphocytes, but also B-lymphocytes, dendritic and NK-cells. Mesenchymal stem cells are able to inhibit proliferation of immune cells and reduce their secretion of inflammatory cytokines, acting as auxiliary cells to improve the survival of islets in the early post-transplant phase. Combined transplantation of multipotent mesenchymal stem cells and pancreatic β-cells is a promising approach to the treatment of type I diabetes mellitus. Deeper study of the mechanisms that cause their cytoprotective effect upon the transplant may be helpful for implementation of this therapeutic approach and improve its efficiency. In our study, a 1% solution of low-viscosity sodium alginate with addition of saline solution (0.9% sodium chloride) was used to create immuno-insulating scaffolds, and a 2.2% BaCl2 solution was added for polymerization. Decreased production of proinflammatory cytokines (TNFα, IL-12, IL-5) and growth factor (GM-CSF) was registered in co-cultures of β-cells with mesenchymal stem cells of bone marrow origin, and those obtained from subcutaneous adipose tissue. Anti-inflammatory activity was more pronounced in adipose stem cells and their immunomodulatory effects were shown via changes of their cytokine-producing activity. Hence, the multipotent mesenchymal stem cells obtained from adipose tissue and bone marrow have shown to exert cytoprotective effect upon pancreatic beta-cells by shifting the cytokine-producing activity towards an antiinflammatory profile.

T-lymphoblast-like human leukemia cells of the Jurkat line (Jurkat T cells) form polyploid forms with a high DNA content in suspension culture. Due to pronounced genetic instability, this contributes to further transformation and development of clonal diversity (polyclonal) of the cell line. Little information is available on the adherent subpopulation of Jurkat T cells. In this work, we analyzed the content of nucleic acids in suspension (DNA) and adhesive (DNA, RNA) subpopulations of Jurkat T cells using flow cytometry and propidium iodide dye, as well as confocal laser microscopy and acridine orange dye. The morphology and mobility of large (with a diameter of more than 15 μm) Jurkat T cells adhering to plastic were studied using Cell-IQ phase-contrast microscopy in real-time. According to the intensity of fluorescence in the conditionally green wavelength range (300–530 nm: from UV to green) and conditionally red (565–800 nm: from red to far-red), three subpopulations of adherent Jurkat T cells were identified: with high, medium, and low nucleic acid content. Thus, Jurkat-T cells adhering to the plastic surface of the plates retain a pronounced heterogeneity in the DNA content characteristic of the suspension fraction, which suggests a difference in the morphofunctional properties (polyclonicity) of this subpopulation of cell culture. With a sharp increase in the total cell mass, the proportion of large (giant, 15–50 μm or more) cells attached to the plastic remained constant for 21 days of cultivation and amounted to 1% of the adhesive fraction. It was found that large Jurkat T cells (with a median diameter of 31 µm) moved along the plastic at a linear (median) speed of 38 µm/h. The polynuclear Jurkat T cells on plastic are morphologically identified, with a linear increase being revealed in the mobility of adherent cells with an increase in their diameter (regression coefficient r = 0.33, p < 0.02, n = 52). Possible cellular and molecular mechanisms of an increased number of DNA copies in some adhering Jurkat T cells are discussed. It is assumed that the discovered new property (locomotor activity) can provide polyploid (multinucleated) adhering Jurkat T cells with a significant advantage—directed migration (chemotaxis) in a growing cell population under conditions of nutrient deficiency due to a change in the nutrient medium after 3–4 days of cultivation.

Steatosis in the liver in obesity increases the work of mitochondria to utilize excess lipids. An overload of β-oxidation of fatty acids, the tricarboxylic acid cycle, and oxidative phosphorylation leads to a decrease in ATP and an increase in the formation of reactive oxygen species. Normally, mitochondria can efficiently remove elevated levels of reactive oxygen species using the cell's antioxidant system and metabolic adaptation to altered conditions. This study aimed to investigate the role of hepatic SOD expression in the pathogenesis of NAFLD in obesity. It was found that the level of SOD1 expression in the liver in obese patients with and without type 2 diabetes with a BMI > 40 kg/m2 was lower than in healthy donors. The copy number of mitochondrial DNA (mtDNA) in the liver in all obese patients was more than two times lower than in the control group. In the liver of obese patients without type 2 diabetes, the SOD1 protein level and the mtDNA copy number were interrelated and negatively correlated with the area of fatty inclusions. Thus, in obese patients, a decrease in antioxidant defense in the liver leads to the vulnerability of mitochondria, which, in turn, contributes to the progression of steatosis and insulin resistance.

The review considers complex, controversial, and individual effects of heparin and its derivatives on the bone and circulatory systems in dependence of the dose, the state of the cells and tissues of the recipient. General data on the anticoagulant activity of heparin and its derivatives are presented; special attention is paid to the effect of heparin on mesenchymal cells and tissues and its role in angiogenesis. We also discuss the ability of heparin to bind osteogenic and angiogenic biomolecules in the context of the development of systems for their delivery and sustained controlled release and propose a schematic representation of the positive and side effects of heparin as a delivery system for biomolecules in tissue engineering.

The paper reports a study of the morphofunctional activity of T-lymphocytes in response to the in vitro contact with calcium phosphate (CP) coating in the presence of the particles bearing antibodies against CD2, CD3, and CD28 antigens. VT1-0 titanium plates (10 × 10 × 1 mm 3 ) with a double-sided microarc rough ( R a = 2–5 µm) CP coating were used as the model samples of the mineral matrix of the bone tissue. Magnetic particles (MACSiBead TM T-cell Activation/Expansion Kit human) bearing antibodies to CD2, CD3, and CD28 antigens were used as a T-cell activator (TCA) simulating the signals produced by antigen-presenting cells (APCs). Mononuclear cells (MNCs) isolated from human blood (98.8% of CD45CD3 + cells) were cultured in the presence of samples with a CP coating and/or TCA (2 × 10 6 particles in 1.5 mL of nutritive medium in the proportion of 2 : 1 to cells) for 2 and 14 days. The CP coating and TCA triggered MNC culture adaptation in a synergic way via the mechanisms of hyperactivation and subsequent death of T-lymphocytes. Immune selection occurred through the accumulation of the naive CD45RA + /RO + T-lymphocytes and memory T-cells with the simultaneous depletion of the CD4 + and CD8 + T-cell pool. The shift in the T-lymphocyte populations was observed together with the increase (after 48 h of culturing) in the cell secretion activity with its subsequent decrease by the 14th day of observation. CP coating sustained (compared with the cell culture grown on plastic) the secretion ability of Th1 (IL-12, TNFα, and IFNγ) and Th2 (IL-4, IL-6, IL-10, and IL-13) lymphocytes. At the same time, the prolonged TCA signal after the 48-h activation caused the depletion of T-cell secretion. The suggestion that the observed in vitro effects may play role in the switching of signaling between T-lymphocytes, APCs, and CP materials at the cell–foreign body interface, which may result in a change in the inflammation phase, development of immune tolerance, successful osseointegration of the implant, or bone tissue remodeling impairment, is discussed.

Background. The pathogenesis of nonalcoholic fatty liver disease (NAFLD), which develops in obesity and type 2 diabetes mellitus (T2DM), is associated with the effects of inflammatory factors on the liver parenchyma and liver mitochondrial dysfunction. Aim. To determine the role of sTNFSF14 in the regulation of liver mitochondrial biogenesis in obese patients with and without T2DM. Materials and methods. The study included 263 obese patients with and without T2DM and 42 apparently healthy donors. Quantitative determination of cytokines in the blood plasma was performed by fluorescence flow cytometry. The level of relative gene expression in the liver biopsy samples was investigated by real-time PCR. Semi-quantitative determination of proteins in the liver biopsy samples was studied by western blotting. Results. The study showed that the levels of sTNFSF14, interleukin (IL)-10, gp130 / sIL-6Rb, and sIL-6Ra in the blood plasma of the obese patients without T2DM significantly exceeded the similar values in the control patients and obese patients with T2DM. In the liver biopsy samples of the obese patients with T2DM and a body mass index (BMI) > 40 kg / m2, the expression level of the dynamin-1-like protein (DRP1 / DNM1L) gene was lower than in the control group, and the expression level of the mitofusin 2 (MFN2 ) gene tended to be higher. Compared with the control group, an increase in the expression level of the NADH-ubiquinone oxidoreductase chain 4 (MT-ND4) gene was recorded in the liver of all the obese patients. The patients with obesity showed a decrease in the amount of mitochondrial DNA (mtDNA) compared with the control group. Conclusion. Thus, sTNFSF14, interacting with IL-10 and gp130 / sIL-6Rb in the circulation, positively effects the liver in the obese patients without T2DM. A low level of sTNFSF14 in the blood plasma of the obese patients with T2DM results in decreased mitochondrial division and increased cellular respiration.

Introduction: Currently, traumatology actively uses ceramic materials with calcium phosphate coating (CP). Such materials have a stimulating effect on multipotent mesenchymal stromal cells (MMSCs) and immunocompetent cells. So, their use involves an assessment of the impact of these materials on the development of tumor cells. Methodology: The Jurkat 5332 cell line and human adipose-derived MMSCs (AMMSCs) were examined. 3D culture was simulated by adding to the cell culture the substrates from commercially pure titanium with rough (Ra=2-5 µm) CP microarc coating. The cultures were used: 2D with Jurkat T cells (JTCs) on plastic surface; 2D co-culture of JTCs and AMMSCs on plastics; 3D with JTCs and CP matrix; 3D with JTCs, AMMSCs, and CP matrix. Findings & Conclusion: Both 2D and 3D JTC cultures showed an increase in the CD45RO receptor expression that led to increasing number of CD45RO+CD45RA+ cells. Probably, JTCs restored the partial maturation and differentiation. Vice versa, the expression of CD45RO receptor and the number of CD45RO+CD45RA+ cells decreased in case of JTCs and AMMSCs co-cultivation. A similar reaction of the cells was revealed in the 3D culture of JTCs and AMMSCs. Thus, JTCs with AMMSCs co-cultivation may

The role of γc cytokines (IL-2, IL-7, and IL-15) in the regulation of apoptotic death of memory T cells under cultivation conditions in vitro was studied using the method of cytofluorimetry. It was detected that the effect of IL-2 in combination with TCR activator is aimed at increasing the number of mature CD45RO+CD95+ T cells with a simultaneous decrease in the number of viable T cells, which gives reason to assume the proapoptotic effect of IL-2 under restimulation conditions. On the contrary, the effect of IL-7 and IL-15 in the activation model in vitro leads to an increase in the number of living T cells in СD45RO cultures (as compared with the samples, in which only the activator was added) against the background of an increase in the number of CD8+CD95+ T cells with a central memory phenotype (TCM). Changes induced by IL-15 are aimed an decreasing the content of CD4+CD95+ T cells in CD45RО+ cultures of the immune memory T cells. Thus, a dose-dependent positive effect of IL-2 on the clonal expansion and negative effect on the viability of T cells was demonstrated. While IL-7 and IL-15, on the contrary, preserve the viability of T cells, participating in the generation of a pool of long-lived memory T cells.

We studied the effects of methylprednisolone (MP) on the late stages of differentiation of activated memory T cells in healthy donors and in patients with the diagnosis of rheumatoid arthritis (RA) in vitro. It was found that MP exerts a suppressing effect on the number of CD4+ T cells expressing the activation (CD25) and co-stimulation (CD28) molecules in CD3+CD45RO+ cultures of healthy donors and patients with rheumatoid arthritis (RA). Assessment of the level of expression of the U2af1l4 and Gfi1 genes controlling the alternative splicing of the Ptprc gene demonstrated unidirectional but quantitatively different changes in health and in RA. Association between the level of expression of the U2af1l4 and Gfi1 genes and the number of CD3+CD4+CD45RO–CD28– cells in the populations of TCR-activated memory T cells in RA patients treated with MP confirms the hypothesis that glucocorticoids can stimulate the formation of terminally differentiated lymphocytes (CD4+CD28–CD45RO–, TEMRA), which are supposed to play the key role in the pathogenesis of RA.

We studied the effect of the native (non-recombinant) alpha-fetoprotein (AFP) on differentiation, proliferation, and cytokine profile of activated helper T cells 17 (Th17). The object of the study was a culture of isolated by immunomagnetic separation helper T cells (CD4+), induced into the Th17 phenotype by using TCR-activator and proinflammatory cytokines (IL-1β and IL-6). AFP had not significant effect on the frequency of Th17 cells (ROR-γτ+) in the helper T cell culture, and did not affect proliferation of these cells, as measured by Ki-67 expression. Evaluation of the cytokine profile of culture supernatants by using the Luminex xMAP technology, revealed that AFP did not affect the levels of IL-4, IL-5, IL-7, IL-8, IL-10, IL-17, IFN-γ and TNF-α, but at concentrations of 50 IU/ml and 100 IU/ml it increased IL-2 production by activated helper T cells. At the same time, AFP suppressed the synthesis of G-CSF and GM-CSF (10 IU/ml), but stimulated the production of CCL4/MIP-1β (100 IU/ml) and CCL2/MCP-1 chemokines (10 IU/ml and 50 IU/ml).