UC-MSC Alleviates The Effect Of Acute Graft-Versus-Host Disease By Promoting MDSC Proliferation And Its Mechanism

Background:Mesenchymal stem cells(MSCs)have low immunogenicity and biological characteristics such as self-replication,multidirection differentiation,tissue repair and immune regulation,which have a great effect on immune cells.MSC can utilize multiple mechanisms to constrain both innate and adaptive immune responses,to prevent graft-versus-host disease(GVHD).Myeloid-Derived Suppressor Cells(MDSCs),as a heterogeneous population of early myeloid progenitor cells originating from the bone marrow.It not only has a strong inhib itory effect on CD4~+T cells,CD8~+T cells and NK cells,but also can promote the proliferation of Treg cells.Therefore,it is believed that MDSC can reduce the severity of GVHD.Our previous work was found that MSC can promote the proliferation of MDSC in peripheral blood after mobilization.However,as an important cell of immune regulation,the mechanism of MSC promoting the proliferation of MDSC has been rarely reported.An in-depth study of the mechanism can further understand the immunomodulatory effect of MSC and lay a better theoretical foundation for MSC treatment of aGVHD.Objective:The purpose of this study was to investigate the mechanism of MSC which promotes the proliferation of MDSC,and use the influence of MSC on MDSC to reduce the severity of aGVHD after HSCT.Methods:26 donors were selected to obtain Peripheral blood mononuclear cells(PBMC)by mobilization and isolation for individual culture and co-culture with Human umbilical cord blood-derived mesenchymal stem cells(UC-MSC).The amount of MDSC in PBMC was determined by flow cytometry.At the same time,Fluorescent activated cell sorting(FACS)was used to isolate the co-cultured MDSC,which protein and m RNA expression levels of NOS-2,IDO and Arg-1 were determined by Western Blot and RT-PCR.Furthermore,the function of MDSC was verified by co-culture with activated T cells.UC-MSC were co-cultured with HLA-DR-cells which sorted by FACS.The levels of IL-6,IL-10,HGF and HLA-G in the supernatant of cells were detected by enzyme-linked immunosorbent assay(ELISA).Further cell experiments showed that UC-MSC could promote the proliferation of MDSC by secreting HLA-G.Therefore,in animal experiments,UC-MSC with overexpression of HLA-G was given to aGVHD mouse model and GVL mouse model and then analyze the effects of UC-MSC on aGVHD,graft-versus-leukemia(GVL)and overall survival(OS).The bone marrow and spleen cells of mice at 2 and 4 weeks after HSCT were further isolated to analyze the effect of UC-MSC with overexpression of HLA-G on the number and function of MDSC cells.Results:The results of in vitro experiments confirmed that UC-MSC can promotes and maintains the proliferation and activity of MDSC through HLA-G/ILT4 pathway and then exert UC-MSCs to mediate the function of MDSCs to inhibit T cells and promote the proliferation of Treg cells.In animal experiments,constructing UC-MSCs over-expressing HLA-G can not only induce MDSCs production in recipient mice,but also improve the function of MDSCs to suppress T cells,and further reduce acute GVHD(aGVHD)symptoms and survival time without affecting the GVL effect.This study demonstrated that UC-MSCs expanded MDSCs via HLA-G/ILT4 reducing the severity of aGVHD without affecting GVL.Conclusions:The immunosuppressive potential of MSCs in the treatment of aGVHD can significantly affect the development of myeloid cells,thus consolidating the position of MSCs in the prevention and treatment of aGVHD.