The Role Of WAVE1on Autophagy And Drug Sensitivity In Leukemia Cells

Objective:To investigate the role of WAVE1on autophagy and drug sensitivity to ADM on HL-60cells by down-regulating WAVE1expression.Methods:Lentivirus mediated WAVE1shRNA affected HL-60cells, HL-60WAVE1shRNA and HL-60ControlshRNA were successfully constructed via CCK-8、Flow cytometry analysis/RT-PCR/Western blot.In lentivirus-transfected HL-60cells,The standard of autophagy was assessed by Western blot and Electron microscopy;the chemotherapy sensitivity of ADM was determined by CCK-8kit.Results:1. CCK-8showed that HL-60cells were affected in different MOI,The cell viability was nearly alike the unaffected.2.HL-60cells were observed expression of GFP under Fluorescence microscope after48hours’ infection.there were many green fluorescent cells in WAVE1shRNA and Control shRNA groups,the uninfection group had not seen any green fluorescence.The transfected rate is highest when MOI was100.Compared with The normol group and Control shRNA group,the expression levels of WAVE1mRNA and protein were decreased confirmed that HL-60WAVE1shRNA was successfully constructed.3.CCK-8reveal that the best screening concentration of puromycin is8ug/ml.4.After treatment of ADM for48hours,compared with Control group,the protein levels of LC3-Ⅱ in WAVE1shRNA were declined evidently; The number of autophagic vacuoles was decreased. Down-regulation of WAVE1can inhibit the ADM-induced autophagy.5. After pretreated with caspas-3inhibitor (Z-DEVE-FMK), HL-60WAVE1shRNA cell viability was decreased considerablly when compared with The normol group and the control,P<0.05.Conclusion:Down-regulation WAVE1can inhibie the cells’s level of autophagy. Knockdown the WAVE1can increased HL-60sensitivity to chemotherapy by suppressing autophagy.