Research Of The Effect Of Decitbine On The Function Of Mesenchymal Stem Cells From Patients With Myelodysplastic Syndromes

Objective:Increasing evidence that myelodysplastic syndromes(MDS)are a heterogeneous group of malignant diseases with abnormal hematopoietic stem cells and bone marrow microenvironment.Mesenchymal stem cell(MSC)is a key component of bone marrow microenvironment.Although animal experiment has demonstrated that the primary genetic disruption in bone marrow MSC can induce DNA damage of hematopoietic stem cells and MDS-like pathological hematopoiesis,knowledge about MSC in humans with MDS was limited,and the results are not always consistent.Therefore,the functional characteristics of MSCs derived from MDS and its changes treatment with dicitabine were studied in this study.Methods:Bone marrow specimens of 40 newly diagnosed MDS patients and 17 nutritional anemia patients were collected from Guangdong People’s Hospital.MSCwere isolated from bone marrow and in vitro culture.The proliferation and growth characteristics of MSC were observed.Under appropriate conditions,MSC was induced to differentiate into osteoblasts and the formation of calcium nodules was observed by alizarin red staining.The cloning potential of MSC was evaluated by fibroblast colony forming unit experiment,the proliferation ability of MSC was calculated by formula,and the proliferation curve of MSC was determined by MTT experiment.The gene expression in MSC was analyzed by real-time quantitative PCR,and the cell cycle and apoptotic characteristics of MSC were analyzed by flow cytometry.The gene expression in MSC was analyzed by real-time quantitative PCR,and the cell cycle and apoptotic characteristics of MSC were analyzed by flow cytometry.Changes of proliferation,cell cycle,apoptosis and gene expression of MDS-derived MSCs after dicitabine treatment were analyzed.By co-culture of MDS-derived MSC with peripheral blood mononuclear cells in vitro,flow cytometry was used to analyze the effect of dicitabine on differentiation and regulation of activated T cells induced by MDS-derived MSC.Results:MSCs in all control groups could grow in vitro,and MSCs in 9 out of 40 MDS patients could not proliferate in vitro.The control group showed fine fibrous appearance,while the MSC from MDS showed flat and polygonal appearance,and its volume was larger than that of the control group.The cloning and proliferation ability of MDS-derived MSCs were lower than those of control group.The number of calcium nodules induced by MDS-derived MSC and the expression level of transcription factors regulating osteogenic differentiation before differentiation were lower than those of the control group.The expression of Dicerl and Jagged-1 genes involved in the transformation of hematopoietic stem cells to MDS and AML is increased in MSCs derived from MDS.The proportion of MSCs derived from MDS in G0/G1 phase was higher than that in control group,while that in S phase was lower than that in control group(P<0.05).The early apoptotic cells in MSC derived from MDS were significantly higher than those in control group(P<0.05).Dicetabine(0.25 μM)had no significant effect on the proliferation and apoptosis of MDS-derived MSCs,but it could change the cell cycle distribution of MDS-derived MSCs,reduce the proportion of G0/G1 cells and increase the proportion of G2/M cells.Dicetabine can weaken the ability of MSC derived from MDS to induce activated T cells to differentiate into Tregs,and reduce the expression of immunosuppressive molecule PD-L1 in MSC derived from MDS.Conclusions:The biological characteristics of MSCs derived from MDS changed,the cell volume increased,the proliferation ability decreased,most cells arrested in G0/G1 phase,the proportion of early apoptotic cells increased,the osteogenic differentiation ability of MSCs derived from MDS decreased,and the gene expression of Decerer 1 and Jagged-1 involved in regulating the transformation of hematopoietic stem cells into MDS increased.Dicetabine can partially alter the biological characteristics of MDS-derived MSCs,reduce the proportion of MDS-derived MSCs in G0/G1 phase,weaken the ability of MDS-derived MSCs to induce activated T cells to differentiate into Tregs,and reduce the expression of immunosuppressive molecule PD-L1 in MDS-derived MSCs.Therefore,the functional changes of MSCs derived from MDS may play an important role in the progression of MDS and may become a potential therapeutic target.