Vol 12, No 5 (2018)

In modern cardiovascular surgery, there remains a problem associated with a lack of venous material for bypass operations. Great interest has emerged in developing small-diameter vascular grafts ensuring stable patency. One possible solution is the creation of a tissue-engineered vascular implant (TEVI) based on a biodegradable polymer scaffold. Being structural and functional features of the scaffold, cell integration and cultivation on the scaffold are the key processes in the TEVI development. The aim of this research is to identify the optimal method of adipose-derived mesenchymal stem cell seeding and cultivation on the tubular biodegradable scaffold from poly(L-lactide). Results of static and dynamic cells cultivation on the scaffold after seeding with the developed filtration method were compared. The proposed method combining filtration seeding and dynamic cultivation has proved its higher efficiency and is suitable for further TEVI development.

Thymic epithelial cells play not only an important structural role, but also create the microenvironment necessary for the maturation and differentiation of thymocytes. One of the stages of maturation of thymocytes is their adhesion to the epithelial cells of the thymus, the mechanism of which has not been sufficiently studied. The goal of the study was investigation of the interaction of mouse thymocytes with cortical cTEC1-2 and the medullary mTEC3-10 thymic epithelial cell lines, namely, the dynamics of the adhesion, apoptosis, and population composition, as well as the expression of the neuropilin-1 (Nrp-1) and plexin A1 receptors (PlexA1) among adherent, nonadherent, and control cells cultured without epithelium. The maximal adhesion index was observed for a 30-min cocultivation; after this time, the index decreased. The main population of adherent cells consisted from immature CD4⁺CD8⁺ lymphocytes, and the relative content of mature CD4⁺ and CD8⁺ thymocytes was lower than in the control. We have shown for the first time that, even at short cocultivation periods (30 and 120 min), an increase in the content of thymocytes at the early apoptosis phase was observed among the population of adherent cells. For the first time, data have been obtained demonstrating the participation of Nrp-1 and PlexA1 in the process of thymocyte adhesion to mouse thymic epithelial cells as independent molecules in the absence of their semaphorin 3A ligand. It was shown that the expression of Nrp-1 on the surface of adherent thymocytes was significantly reduced during the adhesion process compared to the control, and the expression of PlexA1 increased. Preincubation of thymocytes with antibodies to Nrp-1 increased their adhesion to epithelial cells. The data obtained contribute to a better understanding of intercellular relationships in the thymus gland.

Modeling the interaction of multipotent mesenchymal stromal cells (MMSC) and activated immune cells in vitro is an important step in the study of molecular mechanisms of inflammation and tissue regeneration. Under our experimental conditions, we demonstrated that, upon cocultivation under contact and contact-free conditions, MMSC isolated from human adipose tissue suppressed proliferation of peripheral blood mononuclear cells (PBMC) activated by phytohemagglutinin (PHA). It was found that activation leads to an increased ability of CD4⁺ T-lymphocytes to form adhesive contacts with MMSC. In MMSC, cocultivation with activated PBMC increased the synthesis and exposure on the surface of the ICAM-1 adhesion molecule and induced the synthesis of CD80, CD86, and HLA-DR molecules involved in the formation of intercellular contacts with CD4⁺ T-lymphocytes by interaction with their counterreceptors LFA1 and CTLA4/CD28. It was shown that the appearance of intercellular contacts leads to an increase in the production of interferon-gamma (IFN-γ) by CD4⁺ T-lymphocytes and induction of the synthesis of indolamine-2,3-dioxygenase (IDO) in MMSCs. The obtained data allowed clarifying the mechanism of MMSC-mediated immunosuppression, which involves contact interactions with activated CD4⁺ T-lymphocytes.

At present, it is believed that disregulation of alpha-synuclein (SNCA) metabolism and its aggregation is associated with the pathogenesis of Parkinson’s disease (PD). It is thought that the death of dopaminergic neurons in PD can be mediated by the effect of dopamine on the process of alpha-synuclein oligomerization, primarily by direct oxidation of the protein. At the same time, the effect of dopamine on the regulation mechanism of the SNCA gene expression is not excluded. It is known that transcription factors GATA-1, GATA-2, and ZSCAN21 can participate in the regulation of SNCA expression. In the present study, we evaluated the effect of exogenous dopamine (100 μM) on the mRNA level of the SNCA, GATA-1, GATA-2, and ZSCAN21 genes and the level of total alpha-synuclein in cultured peripheral blood lymphocytes (PBL) of patients with PD (n = 18) and individuals of the control group (n = 14) using real-time PCR and ELISA, subsecuently. A decrease in the SNCA expression level under the action of dopamine was first found in PBL of patients with PD (P = 0.013), of the control group (P = 0.004), and of the combined group that included patients with PD and control individuals (P = 0.001). When assessing alpha-synuclein level, a tendency toward its decrease in PBL of individuals from the control group (P = 0.068), as well as of the combined group of PD patients and control individuals (P = 0.059), was revealed by comparing the cells treated and untreated with dopamine. An increase in the mRNA level of the ZSCAN21 gene in PBL of control group individuals (P = 0.022), as well as of the GATA-1 gene in PBL of the group of patients with PD (P = 0.019) cultured in the presence of dopamine, was shown. An increase in the mRNA levels of the genes GATA-1, GATA-2, and ZSCAN21 in PBL under the treatment with dopamine was also observed in the combined group of patients with PD and control individuals (P = 0.027, 0.029, and 0.002 for GATA-1, GATA-2, and ZSCAN21, respectively). Thus, the obtained data show the effect of dopamine on the mRNA level of the SNCA, GATA-1, GATA-2, and ZSCAN21 genes in cultured PBL of PD patients and control group individuals, suggesting a possible effect of dopamine on the regulation of SNCA expression.

Long-term cultivated mammalian cell lines, as a rule, have either an epithelial- or fibroblast-like morphology. The monolayer cell line that we obtained from rat ascites Zajdela hepatoma contains cells of both types, genetically related but phenotypically different. We compared the behaviors of these cells during their migration into the free space of an experimental “wound” under the same cultivation conditions. We measured the following parameters: the dynamics of the cell number in this space, area of cell projection on the substrate, coefficient of cell spreading, rate of cell migration, and proposed parameter W reflecting the cell efficacy in the wound closure. We found that, initially (at the first stage, within 9–10 h), wound closure was due to the cell migration and increased area of cell projection on the substrate. At the second stage, the cell division contributed to a further increase in cell number within the wound. In terms of parameter W, epithelial-like cells moved less chaotically and filled the wound approximately 1.5 times more efficiently than fibroblast-like cells, the motion of which was alike to Brownian. Meanwhile, the average migration rate of epithelial-like cells was about half that of fibroblast-like cells. In the area of the wound that had just closed, the sizes of cells were larger than those of their counterparts outside the wound. We suggest that the increase in cell size was transitory and resulted from changes in the cell functional state during migration to the wound.