Vol 6, No 4 (2011)

опыт зарубежных стран, в частности, сша, где совсем недавно был зарегистрирован клеточный препарат на основе стволовых клеток пуповинной крови hemacord, третью фазу клинических испы- таний проходит инновационный лекарственный препарат prochimal, в котором в качестве дей- ствующего начала используются живые клет- ки, вынуждает формулировать представления о «клеточных препаратах», а, следовательно, решать вопрос о пересмотре существующих клас- сификаций лекарственных средств, дополнений в фармакопею, выработки правил проведения доклинических и клинических исследований с этой категорией препаратов (в частности, иссле- дование фармакокинетики, фармакодинамики, а при выпуске дженериков - биоэквивалентности), создании производственных регламентов и др. по вашему мнению, может ли такой подход решить спорные вопросы о стандартизации и регламентации в создании и применении алло- генной клеточной трансплантации, например, при негематологических показаниях?

Dystrophic epidermolysis bullosa (EB) is an inherited skin fragility disorder in which blistering occurs in the sublamina densa zone at the level of anchoring fibrils of the dermoepidermal junction. EB result from mutations in the type VII collagen gene (COL7A1) and the nature of these mutations and their positions correlate with the severity of the resulting phenotypes. Presently there are not methods of therapy with the expressed curative effect. It is known that the development of approaches to the treatment of EB is based mainly on the protein, gene and cell therapy. In mouse model it was shown that protein therapies of EB is promising since intradermally injected purified human C7 collagen corrected the EB phenotype and markedly prolonged survival of the animals. For gene therapy approaches to EB, the use of various vectors have been developed. In such a way scientists efficiently delivered cDNA or full-length type VII collagen into keratinocytes or fibroblasts and achieve correction of the genetic defect and persistent synthesis and secretion of functionally active collagen in transduced cells. The use of stem cell therapy looks like considerable progress in solving the problems of EB. For this purpose were found suitable the bone marrow mesenchymal cells as they were capable to give rise to functional development of skin cells, including keratinocytes and dermal fibroblasts. Some investigators are looking at the potential of human umbilical cord cells as a source of epithelial stem cells with appropriate ability for epidermal reconstitution. It is also shown that stem cells from human umbilical cord blood in in vitro experiments can differentiate into keratinocytes, so, cord blood is a good source of cells for ex vivo expansion and transplantation in patients with large defects of the skin. An international team of researchers recently conducted an initial phase of clinical trials of donor bone marrow stem cells in patients with the most severe form of EB. Despite the modest initial successes, the results showed that treatment with stem cells in the future will bring hope to extend and improve the quality of life of patients with EB.

The regulation development of haemopoietic stem cells by rabbit anti-human CD34+ stem cells immune globulin has been established. It was showed by experimental investigation on rats with cytostatic haemo-immunosuppression and in culture human CD34+ stem cells. Thus, in animals which received anti-CD34+ immune globulin the recovery of blood and immunity values occurred earlier than in the control animals. The experimental animals recovered haemolymphocytopoiesis after one month while control group normalized haemopoietic function after two months. In vitro colony-forming ability human haemopoietic stem cells are increased in 30 per cent by the addition specific CD34+ immune globulin in counditioned medium. Our preliminary date suggest that rabbit anti-human mesenchymal stem cells immune globulin can stimulate the differentiation of mesenchymal cells in osteoblasts and endotheliocytis the cells that form haemopoietic niche.

The study was focused on proliferative capacities of human hematopoietic cord blood stem cells at two different methods of cryopreservation. The proliferative potential was determined by measurement of telomere length and colonyforming cell assay. The telomere length was estimated by technique that combined flow cytometry and fluorescence in situ hybridization on Beckman Coulter Cytomics FS-500. We used the Telomere PNA Kit/FITC for Flow Cytometry for measuring telomeric sequences as well as MethoCult® GF H4435 in colony-forming cell assays. The programmed Planer Cryo freezer and direct transfer into liquid nitrogen vapour were used for cryopreservation of samples. The correlation between telomere length and capacity of cells to forming colony after thawing were established. The programmed freezing showed advantage over freezing by direct transfer cells into liquid nitrogen vapour by better keeping of proliferative capacity hematopoietic cells.

The authors have constructed and characterized a series of membranes based on resorbable polyhydroxyalkanoates of different compositions. Five PHA types have been studied: a homopolymer of 3-hydroxybutyric acid, copolymers of 3-hydroxybutyric and 4-hydroxybutyric acids, 3-hydroxybutyric and 3-hydroxyvaleric acids, 3-hydroxybutyric and 3-hydroxyhexanoic acids. Scanning electron microscopy and atomicforce microscopy were used to examine the microstructure of membrane surfaces, showing that membranes based on the copolymer of 3-hydroxybutyrate and 3-hydroxyhexanoate had the roughest surface, while membranes based on the copolymer of 3-hydroxybutyrate and 3-hydroxyvalerate had the smoothest surface. The contact angle for water in air was smaller and hydrophilic properties better in the copolymer membranes than in the membranes based on the high-crystallinity homopolymer of 3-hydroxybutyric acid. The culture of mouse fibroblast cell line NIH 3Т3 was used to test PHAbased membranes; results of fluorescent probes of DNA DAPI and the MTT assay show that membranes based on studied PHAs are not cytotoxic on direct contact with cells and are highly biocompatible; their adhesive properties and ability to maintain fibroblast proliferation are similar to those of polystyrene and better than those of polylactic acid membranes.

Wide dissemination of chronic hepatitis and low efficiency of their medication require development of new approaches for treatment of this pathology. One of the most perspective search directions is connected with stimulation of mesenchymal stem cells differentiation into hepatocytes by applying fibroblast growth factor 4 (FGF-4) and hepatocyte growth factor (HGF), which determine primary differentiation of anterior gut epithelial cells into hepatocytes during prenatal ontogenesis in mammals. It is known that hepatic stellate cells (HSC) are the natural source of these growth factors in the liver and they are essential component of microenvironment for differentiating epithelial cells during liver ontogenesis and regeneration. The aim of this survey was to study the ability of stimulation of bone marrow derived multipotent mesenchymal cells (MMSC) hepatic differentiation by cultivation in the hepatic stellate cells conditioned media. MMSC were cultivated in 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 with HSC separated by semipermeable membrane on the boyden chambers; 5) in the mixed co-culture. Results of the survey showed that factors and cytokines, produced by HSC in the media render significant effect on MMSC and lead to their differentiation into hepatoblasts and hepatocytes with stable expression (unlike expression of MMSC cultivated with growth factors) of epithelial markers and ƒ-fetoprotein. Direct intercellular contacts between MMSC and HSC in the mixed co-culture stimulate bulk and apparent expression of hepatic markers unlike in monoculter, but the phenomenon of cell fusion was not detected. Thus, results of this study confirm the hypothesis of the HSC role as an important element of microenvironment for MMSC hepatic differentiation in vitro.

Regeneration of the rat mandible bone tissue when using a novel osteoplastic material TIOPROST in comparison with the material «KollapAn-M» is studied by methods of light microscopy. The results obtained have shown that the inflammation rate of a bone defect volume applying TIOPROST surpasses bone tissue regeneration by the material «KollapAn-M» for more than one month. Having filled the bone tissue defect TIOPROST is determined to form a porous 3D construction which has a supportive function being a tissue-engineering matrix, have no pro-inflammatory properties. TIOPROST restrains the duration of the inflammation alteration and exudation stages in a wound. During biodegradation TIOPROST induces an active growth of blood vessels from a bone defect bed into tissue engineered matrix canals. The data obtained evolve the current knowledge on the role of free-radical processes in post-traumatic bone tissue regeneration mechanisms and substantiate the rationale for application of antioxidant compounds to optimize osteogenesis in bone tissue injury.

Stem cell therapy finds ever-widening application in restoration of neural tissue, and particularly spinal cord injuries (SCI). Still, safety problems are far from being solved and adverse effects of the therapy remain understudied. The goal of the study is to evaluate safety and to detect complications and specific characteristics of clinical application of mobilized autologous hematopoietic stem cells (AHSC) in complex therapy of SCI. The complications have been studied at all stages of SCI therapy: at the stage of AHSC mobilization to peripheral blood, separation stage and cell therapy. It was detected that overall complication rate achieved 75%. Our study showed that AHSC therapy of SCI demands specific training of the medical staff and must be administered in multi-field hospital under the supervision of hematologist, intensivist and neurologist experienced in highly technological methods of treatment.