Induction of Vasculo- and Osteogenesis in Spheroids Formed by Adipose-Derived Stromal Cells.

Vascularization of bioengineered bone tissue constructs remains a challenging problem of regenerative medicine. Spheroids generated in 3D culture of adipose-derived stromal cells supplemented with inducing factors demonstrate stable characteristics and express of mesenchymal, endothelial, and osteoblasts markers, and represent a prototype of vascularized microtissue. Adipose-derived stromal cells spheroids induced to both angio- and osteogenic differentiation can be used in development of new innovative technologies for in vitro fabrication of vascularized bioequivalents for repair of large bone defects.

Angiogenic potential of spheroids from umbilical cord and adipose-derived multipotent mesenchymal stromal cells within fibrin gel.

One of the essential goals in regenerative medicine is microvascularization which enables an effective blood supply within de novo constructed tissues and organs. In our study, we used two common multipotent mesenchymal stromal cell (MMSC) sources (subcutaneous adipose tissue and Wharton's jelly of the umbilical cord) where is a subpopulation of endothelial precursors. In the medium supplemented with VEGF, the 3D cultures of UC MMSCs and ADSCs promoted the endothelial cell differentiation. To evaluate their ability to form a capillary-like network, we encapsulated spheroids within non-modified and PEGylated fibrin hydrogels. The PEGylated hydrogel supported better the formation of multibranched cords than the pure fibrin gel. Analysis of tubule growth rate, length, and branching showed that the differentiated ADSCs had higher angiogenic potential than the differentiated hUC MMSCs. Our study can be a basis for the development of new strategies in tissue engineering and treatment of vascular diseases.

The technology of obtaining multipotent spheroids from limbal mesenchymal stromal cells for reparation of injured eye tissues.

It is known that stem and progenitor cells open new possibilities for restoring injured eye tissues. Limbal eye zone, formed mainly by derivatives of neural crest, is the main source of stem cells for regeneration. The current study considers development of innovative technology for obtaining 3D spheroids from L-MMSC. It was shown that under 3D conditions L-MMSC due to compactization and mesenchymal-epithelial transition self-organize into cellular reparative modules. Formed L-MMSC spheroids retain and promote undifferentiated population of stem and progenitor limbal cells, as supported by expression of pluripotency markers - Oct4, Sox2, Nanog. Extracellular matrix synthetized by cells in spheroids allows retaining the functional potential of L-MMSC that are involved in regeneration of both anterior and, probably, posterior eye segment.

[Dynamics of osteogenesis after inoculation of autogenic mesenchymal stem cells of adipose tissue].

Experiment was conducted on 40 rats of Wister line. On the artificially reproduced experimental model autogenic mesenchimal stem cells (MSC) of adipose tissue were inoculated in space between bone autograph of tibia and mandible. MSC wasn't inoculated in the comparison group. Formation of a new bone substance in space between an autograph and mandible bone was observed. It was clear that after 120 days (180 days), there was a statistically significant decline of the area occupied by an immature fibroreticular bone. Described phenomenon, presumably, could be explained as a result of decline of the number of active cells in the population of inoculated MSC according to phenomenon of limited number divisions of cells on telomeres, described by Hayflick L. and Moorhead P.S.

[Correlation of psychoemotional status and adaptation to complete dentures].

Patients with full adentia are characterized by the formation of specific psycho-emotional status. Rational psychotherapeutic support of these patients largely determines the efficiency of dental prosthetic treatment. At the same time, the definition of mental and emotional status is not included in the diagnostic examination protocol. Considering the above the purpose of the study was to evaluate mental and emotional status of patients receiving complete dentures. Prosthetic rehabilitation of 30 patients with complete teeth loss was performed and clinical evaluation and evaluation of mental and emotional status were carried out before and after treatment. Patients with negative experiences of prosthetics showed a higher level of personal and situational anxiety. There was correlation of adaptation to removable dentures and the patient's personality traits. It is determined that emotional instability during treatment tends to decrease affecting the timing of adaptation to complete dentures. It is noted that patients with repeated prosthetics have earlier recovery of coordination ability of the masticatory muscles.

3D-technology of the formation and maintenance of single dormant microspheres from 2000 human somatic cells and their reactivation in vitro.

We developed an original reproducible 3D-technology for preparation of single dormant microspheres consisting of 2000 somatic cells. The dynamics of microsphere assembly from mesenchymal and epithelial cells of retinal pigment epithelium was traced using time-lapse microscopy: formation of a loose aggregate over 24 h followed by its gradual consolidation and formation of a compact viable microsphere with a diameter of 100-150 µ by day 7. The cell number in the formed microspheres remains unchanged. Reactivation observed upon fusion of epithelial and/or mesenchymal microspheres results in the formation of a united compact microtissue. The fusion dynamics reproduces spherogenesis irrespective of the initial amount of co-cultured microspheres. Reactivation via two-step induced angiogenesis opens new prospects for production of vascularized microspheres and microtissues.

[Dynamics of formation of bone tissue in rats under the influence of new xenotransplantat "Norian"].

In experiments on rats CD comparative studies have been carried out which is the effect of osteoplastic material NORIAN CRS, the rate of bone regeneration after mandibular dental implants (titanium screw) and the combined effect of these processes NORIAN CRS protein and protein (Emdogein). Functionality and maturity of the newly formed bone tissue was evaluated by histological and biochemical methods. The results suggest that the use of osteoplastic material NORIAN CRS is the 120 days leading to almost complete regeneration of bone tissue. It was also found that the combined use of NORIAN CRS and protein Emdogein impractical because adding Emdogein not accelerate bone regenerative process, but in the early postoperative period may cause an inflammatory reaction.

[Impact of xenotransplantation of multipotent mesenchymal stem cells of the bone marrow on the recovery of the structural organization and functional activity of the retina in its damage].

Among the potential sources of cells for transplantation, the most promising autologous material for cell therapy and tissue engineering are multipotent mesenchymal stem cells (MMSC). The authors carried out an experiment dealing with MMSC xenotransplantation into the rabbit suprachoidal space on a model of laser retinal coagulation. For objective assessment of the magnitude and pattern of changes in the functional activity of different retinal cellular elements and the efficiency of MMSC transplantation, they performed electroretinographic and histological studies. The study indicated that MMSC transplantation into the suprachoroidal space is a safe and effective procedure for cell incorporation in order to treat retinal diseases. Electroretinography provided evidence that single MMSC transplantation better preserved the function of photoreceptors (a-wave). Histomorphometric studies demonstrated that the inserted cells stimulated reparative processes in the laser-induced damaged retina.

Cell biology of fetal tissues and fundamental medicine.

During the recent decade the interests of specialists in experimental mammalian embryology were focused on studies of signal networks, regulatory programs, phenotype and behavior of embryonic and mesenchymal stem cells in culture, which triggered more active use of pluripotent stem cells in cell technologies. Laboratory preparation of primary induction tissue (extraembryonic endoderm, mesoderm, hypoblast, nodule, notochord, primary streak) remains impossible, but primary and secondary organizers and antagonist tissues are essential for simulation of axial development. Only in these sophisticated heterogeneous cell system will it be possible to simulate the real physiology and bioinformatics of the embryonic leaflets, their staged transformation into definitive organs. At present, mammalian and human fetal tissues remain the carriers of unique information of the middle and late embryogenesis. For this reason, functional and therapeutic activities of fetal cells and tissues are intensely studied in foreign countries; numerous banks of fetal somatic and stem cells are created. Banks of extraembryonic umbilical blood tissue have been created in Russia. However, the biological and information potentials of fetal/embryonic tissue remain not realized at the theoretical and applied levels.

Transplantation of human embryonic myoblasts and bone marrow stromal cells into skeletal muscle of C57BL/10J-mdx mice.

We studied expression of dystrophin in skeletal muscles of C57BL/10J-mdx mice after transplantation of human embryonic and fetal myoblasts and bone marrow stromal cells. Dystrophin-positive areas corresponding to the location of transplanted cell were detected in muscles of all recipient mice after transplantation of different cell cultures, but the distribution of dystrophin characteristic of normal muscle fibers was detected only after transplantation of embryonic myoblasts. Dystrophin distribution in muscle fibers after transplantation of fetal myoblasts and bone marrow stromal cells was atypical.

In vitro development of neural progenitor cells from human embryos.

Behavior of stem/progenitor cells from the brain of human embryos during in vitro culturing was studied. Cultured cells from human embryonic brain developed and formed neurospheres heterogeneous by their cell composition. In a serum-containing medium some cells underwent differentiation by the neuronal pathway, while others remained in the stem state.

[Embryonic stem cells: from fundamental studies--to clinical application]. / Embrional'naia stvolovaia kletka: ot fundamental'nykh issledovanii--v kliniku.

A short history of embryonic stem cell (ESC) discovery, cell line isolation, phenotyping and cell behavior both in situ and in vitro is outlined. The path of ESC differentiation into restricted specialized cell types mediated by signal network in culture and by grafting cell into recipient tissues is considered. Potentialities of ESC as the generator of spare parts of organs for regenerative medicine and related bioethical issues are briefly outlined.

[Embryonic stem cells (from fundamental biology to medicine)]. / Embrional'naia stvolovaia kletka (ot fundamental'noi biologii k meditsine).

The recent achievements in isolation, primary and long term culturing, cell Immunophenotyping, SAGE--mRNA--profiling, the clonogenic growth parameters of embryonic stem cells and mesenchymal stem cells have been outlined in connection with cell pluripotency and restricted differentiation, mediated by signals both in culture and in situ. Multidisciplinary data obtained by cell biologists, embryologists with recent functional genomic data give the new way for practical implementation of bioimplants derived from embryonic stem cell for cell-replacement therapy.

[Behavior and differentiation of the neural stem cells in vivo]. / Povedenie i differentsirovka neuronal'nykh stvolovykh kletok in vitro

We studied the behavior and differentiation of human and rat neural stem cells after transplantation in the adult rat brain without immunosuppression. The rat stem cells were isolated from the presumptive neocortex of 15-day-old embryos. The human cells were isolated from the ventricular brain zone of 9-week-old embryos and cultivated for two weeks before transplantation. The results of histomorphological studies suggest that the microenvironment factors did not suppress the growth or development of transplanted stem cells. Both rat and human embryonic multipotent neural cells showed similar behavior and differentiation into neurons and glial cells. After transplantation, they continued to mitotically divide and migrated from the graft area to the surrounding tissue of a recipient brain. The presumptive glial cells migrated preferentially along the capillaries and fibrous structures of the recipient brain. Similar behavior of the rat and human neural stem cells in the microenvironment of the recipient adult rat brain and the absence of immune reaction suggest that the transplantation into the rat brain may serve as a model for studying the developmental biology of the human stem cells.

Estimation of 131I thyroid doses for the evacuees from Pripjat.

The thyroid doses of 49,360 inhabitants of Pripjat evacuated after the accident at Chernobyl were reconstructed. During their evacuation most of the evacuees passed through highly contaminated territories. The evaluation of a large-scale public survey showed that only about 50% of the evacuees had left the contaminated areas within 5 days and that 30% of them stayed there for more than 30 days. As a first step, the model of dose estimations was improved, and thyroid doses were assessed for the group of evacuees for whom the 131I activity in the thyroids was measured. The 131I incorporation during the first 5 days after the accident was described by a single-intake model (inhalation); later incorporations were assumed to be proportional to the radioiodine activity in milk. As a second step, the correlation between the calculated doses and individual parameters (place of residence in Pripjat, intake of stable iodine, and age at the time of the accident) was described by an empirical equation. This equation was applied to all evacuees who completed the questionnaires of the public survey. Previous dose assessments were found to overestimate the thyroid doses especially for the younger evacuees. On the basis of these estimations, collective doses and the resulting radiation risks for thyroid cancer were assessed for different age groups.

Dosimetric support of the International Programme on the Health Effects of the Chernobyl Accident (IPHECA) pilot project: main results and problems.

The problem of post-Chernobyl dosimetry is unique in its complexity in the history of radiation medicine and radiation protection. This is because the early experience of mass exposure of people (bombing of Hiroshima and Nagasaki, Windscale and South-Ural accidents, exposure of inhabitants of Nevada in the United States of America, the Semipalatinsk area in the former USSR, the Marshall Islands, and the Goiånia accident in Brazil, and others) differed both in the much simpler structure of the irradiation source and in the number and characteristics of exposed persons. It is obvious that post-Chernobyl dosimetry, both as an independent problem, and as a tool for epidemiological studies, requires significant expertise and economic and technical expenditures. Extensive and deep research has been carried out in Ukraine for the past 10 years. This article reviews the main results of these studies.