MiR34a Regulates The Expression Of Tim-3 Protein And Affects The Killing Ability Of NK Cells In Acute Myeloid LSCs-MV

Objective: To investigate the effect of miR34 a on the NK cells by regulating the expression of Tim-3 protein.Methods: Stem cells were extracted and cultured from acute myeloid leukemia cells and identified by flow cytometry;LSCs-MV were isolated from stem cells by differential centrifugation and observed by transmission electron microscope;the fusion process of LSCs-MV and NK cells was observed by laser confocal microscope;the purpose of low expression and high expression of miR34 a in LSCs-MV was achieved by transfection,and the efficiency of transfection was tested by qRT-PCR.QRT-PCR was used to detect the expression of granzyme B m RNA in NK cells,the expression level of Tim-3 protein,and the expression of PEC by NK cells in the three groups A killing experiment was performed to verify the effect of miR34 a derived from acute myeloid leukemia stem cell-derived microvesicles on the killing function of NK cells.Results: The main results were as follows:(1)Stem cells were successfully extracted from acute myeloid leukemia cells by flow cytometry,and the heterogeneous lipid vesicles(MV)with the size of 0.4ml 0.9 μ m were observed by transmission electron microscope.(2)the transfection efficiency of miR34 a mimics and si RNA miR34 a silencing was evaluated by qRT-PCR.The expression level of miR34 a after transfection with miR34 a mimics(2.27 ± 0.76)was significantly higher than that in miRCtrl group(1.04 ± 0.52)and si-miR34 a group(0.28 ± 0.09).The difference between the two groups was statistically significant(P < 0.05),indicating that the transfection was successful.(3)compared with miRCtrl group,the expression of granzyme B m RNA in NK cells in miR34 a transfection group increased significantly[(4.16 ± 0.43)VS(1.01 ± 0.27)],while that in si-miR34 a group decreased significantly[(0.36 ± 0.11)VS(1.01 ± 0.27)](P < 0.05).(4)compared with miRCtrl group,the killing rate of NK cells in miR34 a transfection group was significantly higher [(0.309 ± 0.154)VS(0.251 ± 0.082)](P < 0.05),and that in si-miR 34 a group was significantly lower [(0.147 ± 0.036)VS(0.251 ± 0.082)](P < 0.05).(5)compared with miRCtrl group,Tim-3 protein expression in miR34 a transfection group was significantly down-regulated [(0.493 ± 0.014)VS(1.225 ± 0.079)](P < 0.05),and Tim-3 protein expression in si-miR34 a group was significantly up-regulated [(1.496 ± 0.133)VS(1.225 ± 0.079)](P < 0.05).Conclusion:(1)Acute myeloid leukemia stem cell-derived microbubble miR34 a can reduce the killing ability of NK cells;(2)Microbubble miR34 a may reduce the killing ability of NK cells by inhibiting the expression level of Tim-3 protein.