Aim: To search for additional molecular-biological markers of cancer stem cell (CSC) involved in the development of intra-tumor heterogeneity for the detection of features of the breast cancer (BC) pathogenesis. Materialts and Methods: Expression of estrogen receptors (ER), progesterone receptors (PR), Her2/neu, E- and N-cadherin, CD24, CD44, Bcl-2, Bax, Slug, P-gp, glutathioneS-transferase (GST) and metallothionein in cell lines was determined by the immunocytochemical method. Expression of ER, PR, Her2/neu, CD24 and CD44 in the surgical material of BC patients were determined by the immunohistochemical method. The levels of the miRNA were determined using real-time polymerase chain reaction. Results: Cells of high-grade malignancy (HGM), MDA-MB-231 and MDA-MB-468 are characterized by high expression of stem cell markers compared to the cells of low-grade malignancy (LGM), T47D and MCF-7: CD44 levels in T47D and MCF-7 cells were in range of 72–79 points, which is significantly lower than in HGM cells (p < 0.05). Also, HGM cells with the properties of CSC were characterized by high expression of antiapoptotic proteins, the transcription factor Slug, and low levels of proapoptotic protein Bax (p < 0.05) compared to LGM cells. In cells with CSC characteristics an increased expression of transferrin and its receptor, ferritin, fentorin and hepcidin was revealed indicating activation of the endogenous iron metabolism. The characteristic feature of HGM cells with CSC phenotype were the increased levels of oncogenic miR-221, -155 and -10b by 60%, 92% and 78%, respectively, and decreased levels of oncosuppressive miR-29b, -34a and -200b by 8.4 ± 0.3, 4.6 ± 0.2, and 3.4 ± 0.6 times compared to MCF-7 line cells. It has been established that the development of resistance to cytostatics is accompanied by increased aggressiveness of tumor cells, loss of expression of hormonal receptors and acquiring of stem phenotype. In particular, increased expression of P-gp was observed in BC cells during the development of resistance to doxorubicin, of GST during the development of resistance to cisplatin along with increased CD44 expression (p < 0.05). We have revealed the relation between the presence of cells with the CSC phenotype (CD44+CD24-/low) and clinical and pathological characteristics of BC patients, their survival and BC sensitivity to neoadjuvant therapy (p > 0.05). Conclusions: The dependence between the expression of CSC markers and the degree of malignancy of tumor cells, development of resistance to cytostatics in vitro was established as well as the predictive value of the detection of the CSC for the individual prognosis of the BC course and sensitivity of the tumors to the treatment.

Aim: To compare biological properties of primary tumor cells isolated from malignant effusion of cancer patients with the same cells of permanent lines established during their long-term cultivation in vitro and to assess the impact of phenotypic conversion that was caused by changes in their microenvironment on their behavioral characteristics. Materials and Methods: The study was performed on primary cell cultures from pleural effusion or ascites of breast and ovarian cancer and permanent cell lines derived from them, namely permanent ovarian cancer cell line I, permanent ovarian cancer cell line II and permanent breast cancer cell line I. Biological characteristics were studied using standard cell culture methods and immunocytochemical assays. Results: Three new cell lines were established from breast and ovarian cancer and cell morphology, migration activity, the kinetics of growth, colony forming activity in semisolid agar and sensitivity to anticancer drug were examined. These characteristics were compared with those of the primary tumor cells. It has been shown that among the primary tumor cells from malignant effusion, cells with mesenchymal characteristics were the most prevalent. Cultivation of primary cancer cells in vitro leads to a phenotypic change of their population: it becomes more homogeneous in morphology with predominantly epithelial-like cells. Also, later after a number of cell doublings in vitro, the cell population changes to include cells primarily with immunophenotypic properties characteristic of epithelial cells. These changes include increase in number of E-cadherin-positive cells and a decrease in number of vimentin and α- smooth muscle actin-positive cells. It was found that significant changes in expression of epithelial-mesenchymal transition associated proteins in cells during their cultivation in vitro in new microenvironment are accompanied by a rapid change in their sensitivity to anticancer drugs. Conclusions: The new breast and ovarian cancer cell lines were established and characterized. The induction of phenotypic transdifferentiation in malignant cells from pleural effusion and ascites can be an important approach for suppressing the progression of neoplastic process.

Introduction: The objective of the study was immunophenotypic and cytogenetic analysis of mesenchymal stem cells from equine bone marrow and foal umbilical cords during in vitro culture. Material and Methods: The mesenchymal stem cells were obtained from equine bone marrow of three horses and from foal umbilical cords of six foals. The cells were cultured in CO2 incubators by standard procedure. Quantitative abnormalities of chromosomes, i.e. aneuploidy and polyploidy, and structural aberrations, i.e. breaks in chromosomes and chromatid, were taken into account during the study. Results: The results of cytogenetic analysis of equine bone marrow mesenchymal stem cells at the third and fourth passages indicated that the level of karyotype variability of these cells corresponded to the spontaneous level of karyotype variability typical of the peripheral blood lymphocytes of this species. Equine bone marrow contained several clones of stem cells that differed in the expression of specific nuclear markers characteristic of proliferating cells. Conclusion: Mesenchymal stem cells from foal umbilical cords during in vitro cultivation are characterised by quantitative abnormalities of the chromosomal apparatus.

Background: The assessment of biosafety of pharmacologically active substances is crucial for determining the feasibility of their medical use. There are controversial issues regarding the use of substances of different origins as implants. Methods: We have conducted the comprehensive studies to determine the in vivo toxicity and in vitro genotoxicity of new generation of hydrophilic gel for implantation (production name of the substance "Activegel") to detail its characteristics and assess its biosafety. Results: In vivo studies have shown the absence of clinical manifestations of intoxication in animals and no abnormalities in their physiological condition, general and biochemical blood tests. Evaluation of the site of the gel application showed no inflammatory reaction and evidenced on normal state of tissues of animal skin. The results of the genotoxicity test indicated that the gel did not affect the parameters of DNA comets and, accordingly, had no genotoxic effect on human peripheral blood lymphocytes. When studying the effect of the gel on malignantly transformed cells in vitro, it was found that the gel for implantation did not change the proliferative activity and viability of human breast cancer cells. Conclusions: Comprehensive in vitro and in vivo study using various experimental model systems showed that the hydrophilic gel for implantation "Activegel" is non-toxic.