Comparison Of Biological Characteristics And Immunosuppressive Activity Between HAMSC And HBMSC

BackgroundAllogeneic hematopoietic stem cell transplantation (allo-HSCT) is an effective method for the treatment of a variety of diseases, including hematologic malignancies, severe aplastic anemia and severe mediterranean anemia. However, the transplant related complications are problems urgently need to be addressed. And acute graft-versus-host disease (aGVHD) is still the most severe cause of non-relapse mortality after allo-HSCT. As we known, the occurrence of aGVHD need to have three conditions below: ①The immune cells that contain sufficient amounts of host heterologous antigens to identify and attack the host; ②The host allogeneic antigen should be different from the allograft explants; ③The host is in a state of immune incompetence and does not produce rejection of the graft. At present, many kinds of drugs, such as anti-human thymocyte globulin (ATG), anti TNF antibody infliximab, cyclosporine A (CSA) and tacrolimus (FK506), are used for the treatment of GVHD. But these drugs have side effects and are easy to cause the infection of virus, fungi and so on, which reduce the curative effect. So, how to reduce the mortality rate, improve the survival rate and the quality of life in patients with allo-HSCT is consider to be the challenge for clinical work.Mesenchymal stem cells (MSC) are multipotent cells that can be isolated from various tissues, such as adult bone marrow, amniotic fluid, placenta, umbilical cord and so on. And the major criteria for the identification of MSC are the morphologic, phenotypic, expansion capacity and in vitro differentiation potential. As for the characteristics of promoting engraftment of donor cells and accelerating the speed of hematological recovery, avoiding T-cell recognition, inhibiting the T-cell proliferation, MSC might become a potential tool in the therapeutic options to treat and cure various of diseases, for example the autoimmune disease, mervous system injuries, Alzheim-er’s disease and cardiovascular diseases. And MSC isolated from boon marrow has been proven to posess the potential in the treatment of GVHD. Nevertheless, the clinical applications of human bone marrow mesenchymal stem cell (hBMSC) are limited for the reduction in cell number, proliferation and the potential of differentiation with age. Moreover, the collection of the hAMSC should be performed with invasive bone marrow puncture. All these issues above limited for doctors and researchers to obtaining a sufficient number of hBMSC.The mature T lymphocytes of donor activated in the patient’s body, causing a series of reaction including the production of IL-2, IFN-y, TNF-a, IL-12 and their receptors, and so on. The IL-2 and TNF-a are produced by Thl, which mediated the cell immune response and played important roles in the aspects of delayed hypersensitivity, autoimmune disease, anti-infection and GVHD. With the immunosuppressive of hAMSC, it seems to be a new promising tool in the prophylaxis and treatment of GVHD.Previous studies have reported that mesenchymal stem cells exist in almost all tissues and organs, and the ability of expansion in vitro of mesenchymal stem cells derived from different tissues is not exactly similar. Therefore, more and more researchers are now committed to finding another MSC for which that the cell procurement does not provoke ethical controversy, and is more easily accessible and safer compared with hAMSC. In 2004, hAMSC was first distributed from the amnion epithelium, which is derived from the embryonic mesoderm of amnion. FACS analysis was performed to determinete phenotypical characterization of hAMSC, which demonstrated that hAMSC expressed similar phenotypes reported for hBMSC. Moreover, hAMSC expressed phenotypes of Oct-3, Oct-4, SSEA-3 and SSEA-4, which were similar to stem cells. In recent years, human amniotic stem cells have been contributed to the treatment of corneal repair, spinal cord injury and so on. So, as the hAMSC more available than hBMSC, and might be considerd to be a better sourse of MSC with any ethical controversy, if it can be a alternative to hBMSC for prevention and treatment of GVHD?ObjectTo compare the biological characteristics and immunosuppressive activity between human amniotic mesenchymal stem cell (hAMSC) and human bone marrow mesenchymal stem cell (hBMSC).MethodsMSC from human amnion and bone marrow were isolated using enzymatic digestion and Ficoll-Hypaque density gradients, respectively. Their biological characteris-tics were compared by morphology, cell growth curves, cell cycle profile analysis, immunophenotype and immunofluorescence assay. Their immunosuppre-ssive activities were studied on total activated T-cell with phytohemagglutinin (PHA-PBMSC).An in vitro co-culture system was performed to study:the comparison of lymphocyte proliferation and supernatant level of IFN-y were measured by CCK-8 assay and ELISA, respectively.Results(1) Both hAMSC and hBMSC demonstrated fibroblast-like morphology. hAMSC were able to expand for at least 15 passages, while hBMSC for only 6-7 passages;(2) There was no significant difference in the proportion of G2/M phase cells in the two cells types (P>0.05);(3) By FACS analysis for immunophenotype, both MSC were shown to be positive for CD105,CD90, CD73 and negative for CD34, CD45, CDllb, CD19, HLA-DR, but hAMSC were positive for Oct-3/4, which was in contrast to hBMSC;(4) Both of hAMSC and hBMSC were positive for vimentin by immunofluoresc-ence assay;(5) Both cells exhibited similar responses to inhibit lymphocyte proliferation induced by PHA in co-culture conditions, and with the increasing proportion of MSC: lymphocyte, the suppressing effect was more obvious;(6)The supernatant IFN-y secretion of hAMSC co-cultured with lymphocyte in a proportion of 1:1 was measured by ELISA after 72 hours, and the level of IFN-y was significantly lower than that in the same co-culture system of hBMSC. In contrast with the IFN-y in the PHA-stimulated group, the level of IFN-y in both co-culture groups was significantly lower.ConclusionMSC from amnion had a higher proliferative capacity and stem cell properties, compared with that of hBMSC. Both MSCs could inhibit lymphocyte proliferation and suppress IFN-y secretion induced by PHA in vitro.