The Study On Molecular Mechanism Hemopoiesis Reconstruction Of Ginsenoside Rh2 For A Mice Model After Peripheral Blood Stem Cell Transplantation

In this study, a model was formed with BALB/c mice on peripheral blood stem cell transplantation. Ginsenoside Rh2, a Chinese herb, was chosen for posttransplantation treatment, for the purpose of observing whether Ginsenoside Rh2 could accelerate the short term hemopoietic reconstitution after the transplantation. The molecular mechanism on Hemopoiesis Reconstruction of ginsenoside Rh2 was also studyed.Formed a model with BALB/c mice on peripheral blood stem cell transplantation:30 BALB/c mice were treated with cyclophosphamide at 400mg/kg intraperitoneally on day-7 and G-CSF at 5ug/kg twice subcutaneously frome day-6to day-1.The whole blood was collected on transplatation day and peripheral blood mononuclear cell(MNC) suspension was harvested after separation of blood over Ficoll-Hypaque gradient(P=1.090g/cm3). Another group of 60 BALB/c mice were given a leathal dose (7.5Gy) of total body irradiation by Co60 rays on day 0. Among them, every mice received 5 x 106 MNCs via the tail vein in 4 to 6 hours postirradiation. After transplantation, the recipients were randomly divided into four groups:group A, i.e. control group, was treated with normal saline at 0.1ml per mouse intraperitoneally, group B, i.e. low-dose ginsenoside Rh2 group, was treated with ginsenoside Rh2 at 4mg/kg intraperitoneally, group C, i.e. high-dose ginsenoside Rh2 group, was treated with ginsenoside Rh2 at 8mg/kg intraperitoneally, and group D, i.e.G-CSF group, was treated with G-CSF at 5ug/kg subcutaneously. All recipients were treated daily from day 0 to+10. Another 15 BALB/c mice were not given any treatment and serve as normal group.Observe the survival among these mice posttranaplantation and calculate survival rate. The complete blood cell count was performed per 3 or 4 days posttransplantation. Dissect spleen and observe colony-forming unit spleen(CFU-S) on day+10.Calculate the number of nucleated cell on and hematopoietic cell colony-forming unit on day+12 and day+23.The expressions of EPOR,JAK-2,STAT-3 mRNA and protein in bone marrow cells of normal mice and induced by ginsenoside Rh2 were detected by Real-time PCR method and Western blooting method.1. The survival rate was significantly higher in the medication administration groups than the saline group and significant difference was seen among these groups.2. The white blood cell count dropped rapidly postirradiation, and fell down to the bottom on day 7 to 10, then increased slowly. The medication administration group became to show a faster recovery than the saline group, theG-CSF group became to show a faster recovery than the two ginsenoside Rh2 groups. But there was no difference between the two ginsenoside Rh2 groups. The platelet count also dropped rapidly postirradiation, and fell down to the bottom on day 14, then increased slowly, but there was no difference between the four groups.The hemoglobin dropped slowly postirradiation. The hemoglobin was significantly higher in the medication administration groups than the saline group and significant difference was seen among these groups.Beacause the hemoglobin were not dropped to the bottom during the experiment, we can not explain the reconstruction of the red blood cell.3. The number of CFU-S and hematopoietic cell colony-forming unit was much higher in the medication administration group than in the saline group on day 10 posttransplantation. The G-CSF group was higher than the two ginsenoside Rh2 groups.But there was not significant difference between the two ginsenoside Rh2 groups.4. The number of nucleated cell was much higher in the medication administration group than in the saline group on day 10,day 14 and day 23 posttransplantation. The G-CSF group was higher than the two ginsenoside Rh2 groups. But there was not significant difference between the two ginsenoside Rh2 groups.5. The expressions of EPOR,JAK-2,STAT-3 mRNA and protein in bone marrow cells induced by ginsenoside Rh2 were stronger than those of the normal group. This study demonstrated that ginsenoside Rh2 can accelerate the short term reconstitution of blood cell, nucleated cell and promote the formation of the colony forming unit of stem cell/progenitor cell, and consequently facilitate the reconstruction in mice after peripheral blood stem cell transplantation. Ginsenoside Rh2 may activate JAK-STAT passway on gene and protein levels to promote the expressions of EPOR,JAK-2 and STAT-3 and consenquently to promote hemopoiesis reconstruction.