Biotechnological market in harvesting and application ofumbilical cord blood stem cells, is permanently developing inthe world. Efficiency of regenerative therapy, based on useof cord blood stem cells depends on the number and diversityof HLA-typed cord blood units. Human umbilical cord blood isa genuine source of the stem cells rich biological material,important for human. 114 patients with neurodegenerativediseases were transplanted with allogenic umbilical cordblood. Cord blood stem cells were used without selection byHLA antigens, but only by ABO blood group and Rh factor. Thepossibility of using of allogenic umbilical cord blood stem cellsin regenerative therapy, and the lack of immunosupressionwill overcome significant obstacles existing in cell therapy todate. In practical terms, its becoming clear that extensiveapplication of allogenic umbilical cord blood stem cellsregenerative therapy will seriously changes approaches to thedevelopment and delivery of umbilical blood stem cells-baseddrugs existing nowadays and approved by many generationsof doctors.
A series of reports about pro-angiogenic and procancerogenicactivity of thyroids hormones (TH) and itsanalogues sold through nongenomic action. The nongenomicactions of TH require a plasma membrane receptor or nuclearreceptors located in cytoplasm. The plasma membranereceptor is located on integrin бVв3 at the Arg-Gly-Asprecognition site important to the binding by the integrinof extracellular matrix proteins. TH differ in their ability toinfluence the бVв3: L-thyroxine (T4), linking through the mainsite of the receptor, it activates mainly mitogen-activatedprotein kinase (MAPK; ERK1/2) and 3,5,3-triiodo-L-thyronine(T3) binding by other site affects to phosphatidylinositol-3-kinase (PI-3-K). This messengers transduce the hormonesignal into complex cellular/nuclear events including increasedexpression of basic fibroblast growth factor (bFGF), vascularendothelial growth factor (VEGF), hypoxia-induced factor 1-б(HIF-1б) and consequent leads to activation of angiogenesisand cell proliferation. Treatment of cell with thyroidhormones caused expression of inflammation-associatedgenes: cyclooxygenase-2, matrix metalloproteinase-9.Tetraiodothyroacetic (tetrac) blocks thyroid hormone effectson angiogenesis and cancer cell proliferation.
The regulation development of haemopoietic stem cells byrabbit anti-human CD34+ stem cells immune globulin has beenestablished. It was showed by experimental investigation onrats with cytostatic haemo-immunosuppression and in culturehuman CD34+ stem cells. Thus, in animals which receivedanti-CD34+ immune globulin the recovery of blood andimmunity values occurred earlier than in the control animals.The experimental animals recovered haemolymphocytopoiesisafter one month while control group normalized haemopoieticfunction after two months. In vitro colony-forming abilityhuman haemopoietic stem cells are increased in 30 per centby the addition specific CD34+ immune globulin in counditionedmedium. Our preliminary date suggest that rabbit anti-humanmesenchymal stem cells immune globulin can stimulate thedifferentiation of mesenchymal cells in osteoblasts andendotheliocytis the cells that form haemopoietic niche.
The levels of DNA-damage and 8-oxiguanine were estimatedusing the comet assay in multipotent mesenchymal stromalcells (MSC) on different passages. Twenty eight cultures MSCfrom bone marrow of healthy donors have been analyzed. Thelevel of DNA-damages in MSC, estimated as percent of DNA incomet tail (%DNA in comet tail), didnt change in the processof cultivation (3,9±0,4 % on 3-4 passages and 3,8±0,6 %on 10-12 passages). No significant differences in the contentof 8-oxiguanine in the DNA of cells at different stages ofcultivation (1,9±0,24 p.u. 3-4 passages and 2,1±0,22 p.u.on 10-12 passages) havent been revealed. The higher levelsof DNA damage and apoptotic comets observed in two culturesof MSC.
The study was focused on proliferative capacities ofhuman hematopoietic cord blood stem cells at two differentmethods of cryopreservation. The proliferative potential wasdetermined by measurement of telomere length and colonyformingcell assay. The telomere length was estimated bytechnique that combined flow cytometry and fluorescencein situ hybridization on Beckman Coulter Cytomics FS-500.We used the Telomere PNA Kit/FITC for Flow Cytometry formeasuring telomeric sequences as well as MethoCult® GFH4435 in colony-forming cell assays. The programmed PlanerCryo freezer and direct transfer into liquid nitrogen vapourwere used for cryopreservation of samples. The correlationbetween telomere length and capacity of cells to forming colonyafter thawing were established. The programmed freezingshowed advantage over freezing by direct transfer cellsinto liquid nitrogen vapour by better keeping of proliferativecapacity hematopoietic cells.
The aim of the study was to asses morphologic andmorphometric lung tissue changes and peritoneal macrophagesactivity (MA) after allogeneic multipotent mesenchymalstromal cells (MMSC) systemic transplantation in acuteelastase model of pulmonary emphysema(PE) in rats.Methods. Forty 3-months old Wistar rats were randomizedinto 4 groups. Control group (1 group) was injectedintratracheally 0,4 ml of normal saline, other animals (2-4groups) received one intratracheal injection of 20 units (U)porcine pancreatic elastase in 0,4 ml of saline. Next day (3group) and 7 day (4 group) rats were intravenously injected2106 autologous MMSC in 0,5 ml of saline. 2 group wasused as emphysema control. Before euthanizing at the 21stday rats were undergone peritoneal lavage with analysis ofhemi-luminescent macrophages activity in the obtained fluid.Results. The lungs of 2-4 groups had various degreesof PE. The imean linear intercept in group 2 experimentalemphysema increased by 231% versus the control group. Thetransplantation of MSCs in a day after elastase decreasedmean linear intercept by an average of 50.3% compared to thecontrol of experimental emphysema. In animals MMSC injectedat the 7 th day of study, this sizes decreased more greater -by 64,5%, but was higher by 40.7% compared to the firstcontrol group. Another quantitative measure of PE alveolarindex was significantly increased by 149% in group 2.The transplantation of MMSC at 1-st and 7-th day of experimentleaded to alveolar index decreasing, respectively on 163.4and 237% . The peak Index of hemi-luminescent macrophagesactivity (Mv/106 cells) has made 28,8+1,1 (1 group),57,3+1,3 (2 group), 35,8+1,6 (3 group), 31,9+1,9(4 group).Conclusions. Our study confirmed the possibility ofregenerative lung tissue effect of autologous MMSCintravenous injected in experimental rat models of PE
The authors have constructed and characterized a seriesof membranes based on resorbable polyhydroxyalkanoates ofdifferent compositions. Five PHA types have been studied: ahomopolymer of 3-hydroxybutyric acid, copolymers of 3-hydroxybutyricand 4-hydroxybutyric acids, 3-hydroxybutyricand 3-hydroxyvaleric acids, 3-hydroxybutyric and 3-hydroxyhexanoicacids. Scanning electron microscopy and atomicforcemicroscopy were used to examine the microstructureof membrane surfaces, showing that membranes based onthe copolymer of 3-hydroxybutyrate and 3-hydroxyhexanoatehad the roughest surface, while membranes based on the copolymerof 3-hydroxybutyrate and 3-hydroxyvalerate had thesmoothest surface. The contact angle for water in air wassmaller and hydrophilic properties better in the copolymermembranes than in the membranes based on the high-crystallinityhomopolymer of 3-hydroxybutyric acid. The culture ofmouse fibroblast cell line NIH 3Т3 was used to test PHAbasedmembranes; results of fluorescent probes of DNA DAPIand the MTT assay show that membranes based on studiedPHAs are not cytotoxic on direct contact with cells and arehighly biocompatible; their adhesive properties and ability tomaintain fibroblast proliferation are similar to those of polystyreneand better than those of polylactic acid membranes.
In mammalian heart, cardiac myocyte division ceases withinthe first week of postnatal life posing one of the most intriguingquestions of evolution. Stimulation of cardiac myocyte proliferationduring postnatal period would have profound impact on cardiacregeneration in humans. It has been shown that the primaryculture of cardiac myocytes obtained from newborn rat couldserve as an appropriate model for the investigation of the processestaking place in the heart during early postnatal ontogenesis.Similarly to the in vivo situation, the increased mitotic activityobserved during the first 2-4 days after birth is diminished incardiac myocyte culture within 4-5 days of culturing. Besides,60% of cultured cells underwent mitotic division followed by increasein their volume which also closely resembles the processof cardiac myocyte hypertrophy in vivo. The cardiac myocyte volumewas progressively increased during culturing and equaled to81968, 1532212, and 3246190 m3 on the 1st, 3rd, and6th day of culture, respectively. Furthermore, the rate of cardiacmyocyte growth in culture was similar to the pattern of myocytehypertrophy observed in the in vivo settings. Myocyte hypertrophyin culture was associated with the formation of polyploid andmultinucleated, most commonly binucleated, cells which is generallyanalogous to the in vivo myocyte hypertrophy. The analysisof myocyte volume distribution suggested that during cultivationnearly 60% of cells are switched from hyperplasia to hypertrophy,stopping at the G2/M boundary of the cell cycle.The results obtained indicate that cell growth patterns inprimary cardiac myocyte culture share a lot of similarity with thein vivo cardiac myocyte growth. Primary cardiac myocyte cultureis a valuable tool for investigation of the processes responsiblefor cessation of cardiac myocyte division in adult mammals.
Wide dissemination of chronic hepatitis and lowefficiency of their medication require development of newapproaches for treatment of this pathology. One of the mostperspective search directions is connected with stimulationof mesenchymal stem cells differentiation into hepatocytesby applying fibroblast growth factor 4 (FGF-4) and hepatocytegrowth factor (HGF), which determine primary differentiationof anterior gut epithelial cells into hepatocytes during prenatalontogenesis in mammals. It is known that hepatic stellate cells(HSC) are the natural source of these growth factors in theliver and they are essential component of microenvironmentfor differentiating epithelial cells during liver ontogenesis andregeneration. The aim of this survey was to study the ability ofstimulation of bone marrow derived multipotent mesenchymalcells (MMSC) hepatic differentiation by cultivation in thehepatic stellate cells conditioned media. MMSC were cultivatedin various models: 1) in the media free of growth factors;2) in the media by sequential exposure of growth factors;3) in HSC conditioned media; 4) co-cultivation together withHSC separated by semipermeable membrane on the boydenchambers; 5) in the mixed co-culture. Results of the surveyshowed that factors and cytokines, produced by HSC in themedia render significant effect on MMSC and lead to theirdifferentiation into hepatoblasts and hepatocytes with stableexpression (unlike expression of MMSC cultivated with growthfactors) of epithelial markers and -fetoprotein. Directintercellular contacts between MMSC and HSC in the mixedco-culture stimulate bulk and apparent expression of hepaticmarkers unlike in monoculter, but the phenomenon of cellfusion was not detected. Thus, results of this study confirmthe hypothesis of the HSC role as an important element ofmicroenvironment for MMSC hepatic differentiation in vitro.
Induced pluripotent stem cells (iPSCs) are analogousto embryonic stem cells in their properties, and might bederived from adult somatic cells. There exist two mainapproaches to practical application of iPSCs technologies -they are regenerative medicine and human diseases modeling.Nowadays the process of iPSCs obtaining is being investigatedin many world laboratories, but there are no standards in theiPSC obtaining technology. This work was aimed to develop asimple, cheap and practical protocol for obtaining iPSCs andto establish a collection of iPCS lines derived from cells ofpatients with a Parkinson disease to make a cell model of thisdisease. The work resulted in a development of the protocolto produce iPSCs based on a lentiviral delivery of transgenesinto cells. iPCS lines from 3 patients with inherited Parkinsondisease forms have been obtained.
Regeneration of the rat mandible bone tissue when using anovel osteoplastic material TIOPROST in comparison with thematerial «KollapAn-M» is studied by methods of light microscopy.The results obtained have shown that the inflammation rateof a bone defect volume applying TIOPROST surpasses bonetissue regeneration by the material «KollapAn-M» for morethan one month. Having filled the bone tissue defect TIOPROSTis determined to form a porous 3D construction which has asupportive function being a tissue-engineering matrix, haveno pro-inflammatory properties. TIOPROST restrains theduration of the inflammation alteration and exudation stagesin a wound. During biodegradation TIOPROST induces anactive growth of blood vessels from a bone defect bed intotissue engineered matrix canals. The data obtained evolvethe current knowledge on the role of free-radical processesin post-traumatic bone tissue regeneration mechanismsand substantiate the rationale for application of antioxidantcompounds to optimize osteogenesis in bone tissue injury.
Stem cell therapy finds ever-widening application inrestoration of neural tissue, and particularly spinal cord injuries(SCI). Still, safety problems are far from being solved andadverse effects of the therapy remain understudied. The goalof the study is to evaluate safety and to detect complicationsand specific characteristics of clinical application of mobilizedautologous hematopoietic stem cells (AHSC) in complextherapy of SCI. The complications have been studied at allstages of SCI therapy: at the stage of AHSC mobilization toperipheral blood, separation stage and cell therapy. It wasdetected that overall complication rate achieved 75%. Ourstudy showed that AHSC therapy of SCI demands specifictraining of the medical staff and must be administered inmulti-field hospital under the supervision of hematologist,intensivist and neurologist experienced in highly technologicalmethods of treatment.
Nowadays we are dealing with a new level of biomedicalscience, that is «regenerative medicine», which providesopportunities for new treatments and the prolongation ofhuman life. It demands that new evaluative criteria, bothethic and regulatory should be worked out and the society,scientific institutions and local authorities should respond in aquick and adequate way.The authors, who are working in the frame of MegagrantProject of the Russian Government on creating Research,Education and Clinical Center of Regenerative Medicine,consider that one of the major tasks is regenerative medicineapproaches to be governed by precise ethical guidelines,and social and cultural aspects. So, the first target of thiscollaboration is to define regulations and address ethicalissues governing the field of bioengineered organs and tissues(simple or complex), both in the field of research and clinic.They present information about clinical application of tissueengineering in the world together with the summary ofexperience how regulations and authorities in Europe involvingregenerative medicine approaches for Human Subjects.The authors presented the list of ethical rules for scientificresearch and clinical application in the field of regenerativemedicine.