Effects Of CD47 Expression And Isoforms On Hematopoietic Stem Cell Transplantation And Reconstruction

BackgroundStem cell transplantation is currently one of the main treatments for more than 50 diseases,such as hematological malignancies,autoimmune diseases,and immunodeficiency diseases.Hematopoietic Stem Cell(HSC)transplantation is the longest,most extensive and most successful example of stem cell transplantation in clinical application.For certain hematological malignancies,such as Chronic Lymphocytic Leukemia(CLL),Diffuse Large B Cell Lymphoma(DLBCL)that is not effective for chemotherapy,allogeneic HSC transplantation is the only promising cure.HSC survival and function after transplantation is the key to cure.CD47 as a transmembrane protein expressed on almost all cell surfaces has important biological significance for the survival and function of hematopoietic stem cells.On the one hand,as a ligand,CD47 can bind to SIRPa on the surface of macrophages,and inhibit the phagocytosis of hematopoietic stem cells by macrophages through the CD47-SIRPa signaling pathway.Studies in mice suggest that hematopoietic stem cells up-regulate the expression of CD47 under the stimulation of mobilizing agents,reducing the risk of being engulfed by macrophages.Decreased expression of CD47 on the surface of HSC in patients with hemophilia syndrome causes HSC to be engulfed by macrophages and the decrease in whole blood cells.On the other hand,CD47 as a cell surface receptor also plays an important regulatory role in regulating apoptosis,proliferation and injury.For example,the TSP1-CD47 signaling pathway can induce apoptosis,and the absence of CD47 will induce fully differentiated adult cells to reprogram to form multipotent cells.This shows that the expression of CD47 and the signal pathways related to CD47 are of great significance to the survival and function of HSC after transplantation.Although the study found that there are four different isoforms of natural CD47,there is limited research on the effects of the expression of different isoforms of CD47 on cell fate.ObjectiveTo investigate the changes of CD47 expression level of bone marrow hematopoietic stem/progenitor cells in mice after whole body ionizing radiation.To detect the expression of CD47 isoforms in hematopoietic cells,and study the effects of CD47 expression and CD47 isoforms on hematopoietic stem cell on HSC transplantation and reconstruction through cell biology and molecular biology methods.Method1.Irradiate the whole body of mice with non-lethal dose X-ray,and detect the ratio of bone marrow hematopoietic stem/progenitor cells and the expression of CD47 at different time points after irradiation.2.Extract the RNA of bone marrow hematopoietic stem cells and bone marrow,spleen,peripheral blood mononuclear cells,and identify the isoform and relative expression of the CD47 isoform it expresses.3.Isolate the bone marrow hematopoietic stem cells of CD47 KO mice.Infect them with CD47 isoform-1 or isoform-5 lentivirus.Detect the expression of CD47 isoforms in peripheral blood and the reconstruction of different immune cells at different times after transplanting infected cells into CD47 KO mice irradiated with lethal doses.Conclusion1.Compared with mature hematopoietic cells,mouse hematopoietic progenitor cells highly express CD472.After whole body irradiation,the expression of CD47 in LSK hematopoietic stem cells is up-regulated in the short term,and the long-term reserve is decreased.3.Some mature hematopoietic cells,such as bone marrow cells,spleen cells and peripheral blood mononuclear cells(PBMC),express all isoforms of CD47,mainly isoform 2;some immature hematopoietic cells,such as LSK hematopoietic stem cells express all isoforms of CD47,mainly isoform 3 and isoform 4,and the relative expression of isoform 1 is higher than that of mature hematopoietic cells.