Analysis of circulating tumor cells in patients with triple negative breast cancer during preoperative chemotherapy.

The presence of circulating tumor cells in the blood of patients with triple negative breast cancer (early and locally advanced cancer) before and after preoperative chemotherapy was assessed using expression markers. Before therapy, circulating tumor cells were detected in 5 of 13 (38%) patients with early cancer and in 7 of 17 (41.2%) patients with locally advanced cancer. After therapy, the circulating immune cells were detected in one patient with locally advanced cancer, who had no circulating cells before therapy. The tumor was resistant to chemotherapy and the disease progressed. The detected circulating tumor cells were HER-2-positive, while the primary tumor was HER-2-negative. It was concluded that the circulating immune cells can be a potential marker of the efficiency of therapy and predictors of the disease course, while their phenotype can differ from the phenotype of the primary tumor.

[Comparative transcriptome pairwise analysis of spontaneously transformed multipotent stromal cells from human adipose tissue].

Potential markers and the mechanism of the spontaneous transformation of multipotent stromal cells (MSC) with perivascular immunophenotype have been determined. A transcriptome comparative study was performed involving six paired specimens of normal and spontaneously transformed MSC with perivascular immunophenotype, obtained in the first passages following the isolation of adipose tissue of six healthy donors. According to the results obtained using the microarray Illumina HT-12 v4 with the Significance Analysis of Microarrays software, differentially expressed transcripts were revealed and a statistical analysis using Gene Ontology and Molecular Signatures Databases was conducted. The association of the spontaneous transformation of isolated MSC with perivascular immunophenotype with previously identified oncogenic cell transformation pathways (E2F, ATR/ATM, RAS, and RHOA) is suggested and further aims for more detailed study are set. Potential transformation markers, including largely unknown genes and those previously not associated with cancer genes (HSPB6, PLAC9, FEZ1, DTWD1, APH1A, and ATP5L) are described.

Adenoviral transduction of multipotent mesenchymal stromal cells from human adipose tissue with bone morphogenetic protein BMP-2 gene.

We determined conditions for effective transduction of multipotent mesenchymal stromal cells from human adipose tissue with adenoviral constructs carrying the gene of human bone morphogenetic protein BMP-2. The peak of transgene transcription and BMP-2 protein secretion in the transduced cultures was observed on day 6 after infection. The maximum transcription of BMP-2 gene and genes of osteogenic markers (bone sialoprotein, osteopontin, and osteocalcin) was observed in the medium containing sodium ß-glycerophosphate and ascorbic acid. Addition of D 3 vitamin did not enhance the expression of BMP-2 gene in transduced cells. The obtained cell cultures with high osteogenic potential can be used in bone tissue engineering.

[Molecular-genetic analysis of DNA pol and TK of HSV-1 population using NGS technology].

It was determined the ratio of viral DNA and DNA from Vero cells using the polymerase chain reaction in real time in Vero cell lysate, infected with L2 strain of the herpes simplex virus type 1. Copy number of the virus reached a maximum after 24 hours of incubation of infection. Total DNA was isolated and sequenced using NGS technology by Ion Torrent device. Nucleotide sequences of the thymidine kinase gene (UL23) and DNA polymerase (UL30) were determined for a population of HSV-1 strain L2. Comparison of the primary structure of these genes with the corresponding nucleotide sequences of known strains of HSV-1 KOS and 17 was conducted. Differences in the structure of genes UL23 and UL30 between strain L2 and reference strains KOS and 17 are not important, because changes are found in non-conservative regions.

[A comparative evaluation of bone tissue formation by tissue scaffold and osteoplastic material «Bio-Oss¼ transplantation in the maxillary sinus floor].

The effectiveness of bone tissue formation by transplantation of tissue-engineering scaffold and frequently used osteoplastic material «Bio-Oss¼ combined construction in maxillary sinus floor was evaluated and marked differences in regeneration terms by this treatment mode were found out.

[Bone tissue restoration after tooth removal by means of tissue-engineering construction based on multipotent stromal adipose cells].

The paper contains clinical examples of tissue-engineering construction based on multipotent stromal adipose cells application for tooth socket healing. The method do not only prevents the bone tissue resorption but allows recreating its volume.

Improvement of cardiac contractile function in rats with postinfarction cardiosclerosis after transplantation of mononuclear and multipotent stroma bone marrow cells.

We compared the efficiency of autologous mononuclear cells and multipotent stromal cells of the bone marrow after their non-selective intracoronary transplantation on day 30 after acute coronary infarction in rats. Improvement of hemodynamic parameters of myocardial contractility (rates of left ventricular pressure rise and drop) in comparison with the initial values and deceleration of postinfarction prolongation of QRS and QT intervals were observed in rats of the experimental group in contrast to controls in 4 weeks after transplantation. These functional changes were more intensive after transplantation of multipotent stromal cells and were accompanied by more pronounced morphological signs of reverse myocardial remodeling: thickening of the scarred left ventricular wall, shrinkage of the scar, and decrease in left ventricular dilatation index.

[Actual problems of cell therapy for cardiac diseases].

This article reviews the literatury and own dates of cell therapy for cardiac diseases. The principal unresolved issues were formulated.

Tissue engineering construct on the basis of multipotent stromal adipose tissue cells and Osteomatrix for regeneration of the bone tissue.

We developed a new method of creation of tissue engineering constructs for regeneration of the bone tissue based on the principle of free distribution of cells in a fibrin clot within a scaffold. The tissue engineering construct includes multipotent stromal adipose tissue cells committed in osteogenic lineage, platelet-rich plasma, and resorbed material on the basis of xenogeneic bone collagen. The culture of bone progenitor cells was characterized by the main markers of osteoblastic differon. The material meets all requirements for materials intended for tissue engineering. An innovative high-technological tissue engineering product for clinical application is prepared.

Reparation of the myocardium after transplantation of mononuclear bone marrow cells.

Safety and efficiency of intracoronary transventricular transplantation of autologous mononuclear bone marrow cells in rats with postinfarction cardiosclerosis were studied. The cells migrated to the damaged area and were detected only in the cicatricial tissue; they have fibroblast-like phenotype and some of them were stained with Fapα (marker of reactive fibroblasts). More active proliferation of non-muscular cells and formation and maturation of collagen fibers in the cicatricial tissue were observed after transplantation of mononuclear cells. This led to thickening of the cicatricial wall, but the size of the scar and index of dilatation of the left ventricle remained unchanged. The number and volume density of newly formed blood vessels in the damaged area increased after transplantation, but no labeled cells were seen in the vascular walls. It can be hypothesized that stimulation of neoangiogenesis is mediated by paracrine mechanisms, which also explains improvement of global contractility of the left ventricle (increased contractility index in functional tests). Thus, transplantation of mononuclear bone marrow cells leads to thickening and strengthening of the cicatricial wall, stimulates angiogenesis, and improves global myocardial contractility. However, no morphological signs of reverse remodeling of the left-ventricular myocardium were revealed.

Biocompatibility of tissue engineering constructions from porous polylactide carriers obtained by the method of selective laser sintering and bone marrow-derived multipotent stromal cells.

We studied the biocompatibility of porous polylactide carrier matrices obtained by means of surface selective laser sintering. Carrier matrices had no cytotoxic activity, but maintained adhesion and proliferation of cells. Subcutaneous transplantation of tissue engineering constructions from these carriers and bone marrow-derived multipotent stromal cells did not cause the inflammatory response and pathological changes in rats. The conditions for organotypic regeneration were provided at the site of transplantation (high degree of blood supply and considerable amount of immature precursor cells).

Regeneration of skull bones in adult rabbits after implantation of commercial osteoinductive materials and transplantation of a tissue-engineering construct.

We performed a comparative study of reparative osteogenesis in rabbits with experimental critical defects of the parietal bones after implantation of commercial osteoinductive materials "Biomatrix", "Osteomatrix", "BioOss" in combination with platelet-rich plasma and transplantation of a tissue-engineering construct on the basis of autogenic multipotent stromal cells from the adipose tissue predifferentiated in osteogenic direction. It was found that experimental reparative osteogenesis is insufficiently stimulated by implantation materials and full-thickness trepanation holes were not completely closed. After transplantation of the studied tissue-engineering construct, the defect was filled with full-length bone regenerate (in the center of the regenerate and from the maternal bone) in contrast to control and reference groups, where the bone tissue was formed only on the side of the maternal bone. On day 120 after transplantation of the tissue-engineering construct, the percent of newly-formed bone tissue in the regenerate was 24% (the total percent of bone tissue in the regenerate was 39%), which attested to active incomplete regenerative process in contrast to control and reference groups. Thus, the study demonstrated effective regeneration of the critical defects of the parietal bones in rabbits 120 days after transplantation of the tissue-engineering construct in contrast to commercial osteoplastic materials for directed bone regeneration.

Directions of migration of bone marrow mononuclears after intracoronary transventricular injection.

Directions of migration of mononuclear bone marrow cells after intracoronary transventricular injection procedure developed by us were experimentally studied. After nonselective injection of cells into the right and left coronary arteries in rats, the labeled cells were detected only in the damaged zone of the myocardium. Localization of transplanted mononuclears in the scar attests to their homing into the damaged zone. Numerous cells were found in the red pulp of the spleen and solitary cells were detected in the liver and lungs. In the heart, the labeled transplanted cells were detected only in the scar zone at all terms of the study; they were not incorporated into the vascular walls, but were surrounded by thick bundles of collagen fibers and probably underwent differentiation into fibroblasts. No data on possible differentiation of the transplanted cells into vascular cells or cardiomyocytes were obtained.

Morphological changes in paraurethral area after introduction of tissue engineering construct on the basis of adipose tissue stromal cells.

We studied morphological changes in the paraurethral area of Wistar rats after introduction of tissue engineering constructs on the basis of multipotent mesenchymal stem cells and gelatin sponge. The tissue engineering construct containing autologous culture of the stromal fraction of the adipose tissue was most effective. After introduction of this construct we observed more rapid degradation of the construct matrix and more intensive formation of collagen fibers.

Chromosome variability of human multipotent mesenchymal stromal cells.

We elaborated a method of preparing cytogenetic preparations of cultured multipotent mesenchymal stromal cells from the adipose tissue. It was found that karyotypic changes (monosomy, translocations) appear in some samples during culturing. Clones with changed karyotype were detected in 11-14-passage cultures from 2 of 7 individuals. The percent of aberrant cells in cultures from different individuals varied from 1.5 to 5.95 per 100 cells, which attested to karyotype instability. These data substantiate the need for cytogenetic control of cells before their transplantation into donor organism and further investigation of chromosome variability in stem cells.

[Cytochalasin B induced changes in cytoplasmic Na+/K+ balance in 2-cell mouse embryo].

This work was performed to study changes in intracellular elemental (Na/K) concentrations caused by Cytochalasin B in two-cell mouse embryo using Electron Probe Microanalysis. The presence of Cytochalasin B is required to transfer a somatic cell nuclear into an early embryo cell. The direct effect of this chemical is cytoskeleton transformation, which would be able to cause the increase of potassium channel activity resulting in cytoplasmic Na/K imbalance. In our study Cytochalasin B was shown to decrease the intracellular sodium concentration. The Na/K balance in the cytoplasm of mouse embryos reverted to its intact level after treatment them with Cytochalasin B free Dulbecco's solution. Possible mechanisms responsible for the changes in the intracellular sodium concentration observed in the embryo cells are discussed.

[Prospects for using combined three-dimensional injection of gel transplants with autologous and allogenic cell cultures in reproductive medicine].

A combined cellular transplant for injection has been designed, by using lowly differentiated stromal fat tissue cells obtained from Wistar rats and it preserved biological and physical properties after passing through a small-diameter injection needle (an insulin syringe). A morphological study of the results of its transplantation into paraurethral tissues in 28 female rats showed that the transplant cells, autologous ones in particular, activated the proliferation of local connective tissue cell populations and retained their viability and proliferative and synthetic activities up 20 days or more. At the end of the experiment (on days 27-35), marked focal connective tissue enlargement at the site of injection of both allo- and autografts results in the narrowing of the urethral lumen. The injection implantation of combined allo- and autografts opens up new avenues for the treatment of pathological processes of the reproductive and urinary tracts.

Electron probe microanalysis of potassium and sodium in clonogenic culture of human neural stem cells.

The mean potassium and sodium concentrations and distribution of potassium in clonogenic culture of human neural stem cells (neurosphere) were estimated by means of electron probe microanalysis. High sodium concentration was typical of undifferentiated cells. Potassium was irregularly distributed in the test structure. Our results confirm published data on heterogeneous morphological structure of neurospheres.

Stimulation of reparative osteogenesis after xenotransplantation of human prenatal mesenchymal stem cells and chondroblasts.

Regeneration of the bone tissue after unilateral xenogeneic transplantation of a suspension of human mesenchymal cells and chondroblasts was studied in rats with experimental damage to both femurs. The state of animals was satisfactory and non-depressed in the early and late postoperation period. No local pathological reactions and complications were seen. Administration of mesenchymal stem cells and chondroblasts into the area of bone defect accelerated regeneration on days 10 and 30 (compared to the control group). Implantation of mesenchymal stem cells more significantly stimulated reparative osteogenesis compared to treatment with chondroblasts. This was mainly associated with increased ratio of mature lamellar bone tissue. Unilateral transplantation of the cell suspension stimulated regeneration in the contralateral limb due to accelerated maturation of the bone tissue. The bone tissue formed after transplantation of mesenchymal stem cells and chondroblasts was integrated into bone organ and underwent complete remodeling. Xenotransplantation of prenatal mesenchymal stem cells and chondroblasts without immunosuppression was not followed by the development of early and delayed complications or local reactions of graft rejection.

[Electron probe microanalysis of potassium and phosphorus in mouse oocytes and zygotes].

Intracellular concentrations of potassium and phosphorus were determined by Electron Probe Microanalysis in mouse mature oocytes and zygotes. The oocytes were characterized by insignificant variations in the concentrations of these elements in the cytoplasm: 60 +/- 4 and 103 +/- 6 mM, respectively. In zygotes, on the contrary, significant variations were observed: 64 +/- 16 and 84 +/- 14 mM, respectively. Changes in the potassium homeostasis during the first cell cycle have been discussed.