Savchenkova I.P*, S.V. Korjikova1, L.P. Dyakonov, A.S. Teplyashin1
(All Russian Kovalenko’s Institute of Experimental Veterinary, Russian Agricultural Academe Science, 109428, Moscow, Ryazansky pr. 24/1, Russia; 1 - Institute of stem cell, 107031 Mosсow, St. Kuznetsky most, 17/1. E-mail*: s-ip@mail.ru)
The efforts of isolation of multipotent mesenchymal stromal cells (ММSCs) from a bone marrow (BM) and a adipose tissue (AT) are continued. For last 6 years only from BM have been received 4 subpopulations of ММSCs: RS (rapidly self-renewing) - cells (Colter, 2000), MAPCs (multipotent adult progenitor cells) (Reyer et al., 2001), MIAMI (D'Ippolito et al., 2004) and SD (serum-deprivated) - cells (Pochampally et al, 2004). In all listed efforts the cells having different morphological characteristics and basic differences on expression of surface antigens (Ags) depending on conditions of isolation and cultivation are derived. It testifies to that, the biological basis and the molecular nature of mesenchymal stem cells up to the end are not studied. Manipulations on isolating of a mesenchymal stem cell - parent of all forms of its progenitors demand the further careful research. Vacanti with co-authors (Vacanti, et al., 2001) reported on presence small, spore-like cells (d-3µm) in all tissue and organs of mammals which possessed high viability and were kept in extreme conditions: cryogenic and high-temperature shocks (-860C; +860C), oxygen starvation during 5 days. In this connection the purpose of our researches was to establish extreme conditions for viability and normal functioning of a cell in a tissue to confirm or deny hypothetical model of existence in an adult organism of reserve stem cells. For creation of an extreme situation of the leader to mass destruction of cells used cryogenic shock. For this purpose freshly isolated AT received during lipoaspiration under local anesthesia at five patients (from their consent) was frozen in 50 ml test tubes (Coster) at -700C without a cryoprotector. Tissue was thawed through 48 h and isolated from it cells by a technique described by us earlier (Teplyashin et al., 2005). Results of the lead researches have shown, that in AT, subjected cryogenic shock is kept a some cell population of small cells (d-10µm) with fibroblast-like morphology. Cells were positively stained with Abs (BD Bioscience) for the Ags: CD29, CD44, CD49a,b,d, CD73, CD90, CD105, CD166, HLA ABC. Cells were negative for CD34, CD45, CD133 - markers of hematopoietic stem cells, CD31, Flk - markers of endothelial cells, CD117 - receptor of stem cell factor, STRO-1, as well as for HLA DR, DP, DQ (flow cytometer analysis Epics Elite Coulter) (Fig.). On induction to differentiation in vitro cells were able to differentiate to cells of bone, adipose and cartilage tissues. These cell population had the characteristics similar earlier isolated us ММSC from AT (Teplyashin et al., 2005).Unlike them the derived cell population was homogeneous on the structure. Thus, it has been established, that in AT in extreme conditions for an organism the population of reserve cells with a phenotype similar AT ММSC is kept. These cells can have special value for preservation of genetic resources in extreme conditions
Colter D.C., Class R., DiGirolamo C.M., Prockop D.J. Rapid expansion of recycling stem cells in cultures of plastic-adherent cells from human bone marrow // PNAS. 2000. V. 97. P. 3113-3218.
Reyes M., Lund T., Lenvik T., et al. Purification and ex vivo expansion of postnatal human marrow mesodermal progenitor cells // Blood. 2001. V. 98. P. 2615-2625.
D’Ippolito G., Diabira S., Howard G.A. et al. Marrow-isolated adult multilineage inducible (MIAMI) cells, a unique population of postnatal young and old human cells with extensive expansion and differentiation potential // J. Cell. Sci. 2004. V. 117. P. 2971-2981.
Pochampally R, Prockop D.J. et al. Serum deprivation of human marrow stromal cells (hMSCs) selects for a subpopulation of early progenitor cells with enhanced expression of OCT-4 and other embryonic genes //. Blood. 2004. V. 103. P. 1647-1652
Vacanti M.P., Roy A., Cortiella J. еt al. Identification and initial characterization of spore-like cells in adult mammals // J. of Cellular Biochemistry. 2001. V.80. P. 455-460.
Teplyashin A.S., Korjikova S.V., Sharifullina S.Z. et al. Characteristic of human mesenchymal stem cells isolated from bone marrow and adipose tissue // Tsitologia. 2005. V. 47. № 2. P. 130-135.
Teplyashin A.S., Tchyupikova N.I., Korjikova S.V., et al. Comparative analysis of cell populations with phenotype similar to mesenchymal stem cells derived from subcutaneous fat // Tsitologia. 2005. V. 47. №7. P. 637-643.
Fig. Ag expression on the surface of reserve cells isolated from adipose tissue subjected cryogenic shock. Grey color - control staining with Ig labeled with phycoethythrin; white color--stainig with specific Abs.
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