Comparison Of Immunoregulation Between Human Amniotic Mesenchymal Stem Cells And Human Bone Marrow Mesenchymal Stem Cells On T Lymphocytes In Peripheral Blood

ObjectIn our study,we respectively established the system of human amniotic mesenchymal stem cell(hAMSC)and human bone marrow mesenchymal stem cell(hBMSC)were co-cultured with human peripheral blood mononuclear cell which were stimulated with phytohemagglutinin(PHA-PBMSC)in vitro.To compare the immunoregulation function between human amniotic mesenchymal stem cell and human bone marrow mesenchymal stem cell on allogeneic peripheral blood T lymphocytes.This study can provide experimental basis for the future clinical application of hAMSC treatment of GVHD and solve the problem of insufficient source of hBMSC.Methods(1)The human amniotic membrane mesenchymal stem cell(hAMSC)were isolated from the amniotic tissue of fresh placenta by enzymatic digestion.The hAMSC were cultured in vitro by subculture.The bone marrow mesenchymal stem cell(hBMSC)were isolated from bone marrow by Ficoll-Hypaque density gradient centrifugation,and then the stem cells were cultured in vitro to obtain purified hBMSCs.(2)Peripheral blood mononuclear cells(PBMC)were isolated by Ficoll-Hypaque density gradient centrifugation.(3)The fourth generation hAMSC and hBMSC treated with mitomvcin C(MC)were co-cultured with PHA-stimulated PBMC for 72 hours.The immunoregulatory function of the two kinds of MSCs on peripheral blood lymphocytes was divided into experimental group(HAMSC+PBMC+PHA group and hBMSC+PBMC+PHA group),positive control III group(PBMC + PHA group),negative control group(PBMC group,hAMSC and hBMSC group treated with mitomycin alone).(4)The percentage of Treg,Thl,Th2,Tc1,Tc2 cells in each experimental group and positive control group were detected by flow cytometry.(5)The levels of interleukin-2(IL-2)and interleukin-10(IL-10)in the supernatant of the experimental group and the control group were detected by enzyme-linked immunosorbent assay(ELISA).Results(1)Compared with PBMC+PHA group,the levels of Treg,Th2 and Tc2 in hAMSC+PBMC + PHA group and hBMSC + PBMC + PHA group were significantly increased(P<0.05),the levels of Thl and Tc1 were significantly decreased(P<0.05).There was no significant difference in the T cell subsets between hAMSC+PBMC+PHA and hBMSC+PBMC+PHA group(P>0.05).(2)Compared with PBMC+PHA group,the concentration of Interleukin 10(IL-10)in hAMSC+PBMC+PHA group and hBMSC+PBMC+PHA group were significantly increased(P<0.05),the levels of Interleukin 2(IL-2)were significantly decreased(P<0.05).There was no significant difference in the T cell subsets between hAMSC+PBMC+PHA and hBMSC+PBMC+PHA group(P>0.05).ConclusionThe human amniotic membrane mesenchymal stem cell(hAMSC)and the human bone marrow mesenchymal stem cell(hBMSC)can increase the proportion of Treg,Th2 and Tc2 subpopulations in PHA-stimulated allogeneic lymphocytes in vitro and promote the secretion of IL-10,which can down-regulate the ratio of Thl and Tcl subpopulations and inhibit IL-2 secretion.hAMSC and hBMSC have similar immunoregulatory functions.