Preliminary Study On Culturing And Hematopoietic Supporting Of Human Amniotic Mesenchymal Stem Cells

[Objective]Applying well-established isolation and culture of technology,to study the biological characteristics and the hematopoietic support function of the human amniotic mesenchymal stem cells(A-MSC) were conducted to promote its clinical application in a wide range.It also provides experimental basis in helping clinical co-transplantation of MSC and HSC and improving the success rate of HSCT.[Methods]1.Using established isolation and culture system of mesenchymal stem cells(MSC),amniotic mesenchymal stem cells were isolated,cultured,purificated and amplificated.2.Subcultured cells of the third generation were collected at a number of 10⁶ level.Using RT-PCR method,the expression of many factors,such as LIF,SCF,M-CSF,G-CSF,MG-CSF,IL-3,IL-6,IL-11,were analyzed in vitro cultured human amniotic mesenchymal stem cells(A-MSC) on the level of mRNA expression.3.Nonadherent mononuclear cells of umbilical cord blood were plated onto a feeder single-layer formed by the irradiated A-MSC at third generations,the independently cultured one was used as a control,suspended cells were collected at days 3,7,14 and 21,using a long-term cultures system(MethoCult^TMH5100 ),half of the medium was replaced weekly for the next three weeks.By counting the cells,the function of supporting hematopoietic for amniotic membrane MSC in vitro was evaluated.[Results]In this report we characterize multiple hematopoietic cytokines expression by human amniotic mesenchymal stem cells(A-MSC),including LIF,SCF,M-CSF, G-CSF,MG-CSF,IL-3,IL-6,IL-11.By comparing the number of the suspension cell obtained at the days 3,7,14 and 21 in culture medium,the results showed that the human amniotic MSC has the function to support hematopoiesis in vitro.[Conclusion]It is concluded that tissue explant culture is a fairly ideal method in vitro isolation,cultivation,purification,amplification of human amniotic mesenchymal stem cells.In addition,we also observed that there is the correlation between the human amniotic organization and hematopoietic function in this review. Moreover,it is preliminary proved that human amniotic MSC have possessed the functions of supporting hematopoiesis.Finally,it is suggested that combined transplantation of human amniotic mesenchymal stem cells with hematopoietic stem cell will effectively promote hematopoietic recovery and implantation.