Effect and mechanism of AURKA on proliferation,apoptosis and drug sensitivity of multiple myeloma cells
Objective To explore the effects of AURKA on proliferation,apoptosis,cycle changes and drug sensitivity of multiple myeloma(MM)cells,and analyze the underlying mechanisms.Method Human multiple myeloma cell lines RP-MI8226 and U266 were transfected with smart silent NC(ssi-NC)and smart silent AURKA(ssi-AURKA),respectively.They were divided into ssi-NC group,ssi-AURKA group and blank group.Real time fluorescence quantitative polymerase chain reaction(qRT-PCR)and western blotting were used to detect the relative expression levels of AURKA mRNA and AURKA protein in the three groups of cells.The cell proliferation activity was detected by CCK8 method,and the cell cycle changes and apoptosis rate were detected by flow cytometry in the ssi-NC group and ssi-AURKA group.The drug sensitivity to bortezomib and dexamethasone was evaluated by CCK 8 assay.Western blotting was performed to detect the expression of Wnt signaling pathway related factors and epithelial mesenchymal transition(EMT)related proteins.Results The relative expression level of ssi-AURKA histone in RPMI8226 cells was 0.58±0.09,lower than that of the blank group(1.00±0.10)and ssi-NC group(1.00±0.09),F=20.84,P＜0.001;The relative expression level of ssi-AURKA histone in U266 cells was 0.57±0.07,lower than that of the blank group(1.00±0.04)and ssi-NC group(1.07±0.08),F=46.19,P＜0.001.The results of the cell proliferation activity experiment showed that there was a statistically signifi-cant difference in the relative activity of RPMI8226 cells among the blank group,ssi-NC group and ssi-AURKA group,Ftime=3 966.12,Ptime＜0.001;Fgroup=363.15,Pgroup＜0.001;Ftime×group=48.03,Ptime×group＜0.001.There was a statistical-ly significant difference in relative cell activity among the U266 cell blank group,ssi-NC group and ssi-AURKA group,with Ftime=2 452.32 and Ptime＜0.001;Fgroup=236.91,Pgroup＜0.001;Ftime×group=33.47,Ptime×group＜0.001.The S phase of the ssi-AURKA group in RPMI8226 cells and U266 cells were increased,with t-values of 12.28 and 14.64,respectively,and both P＜0.001;The G0/G1 phase showed a decrease,with t-values of 23.33 and 22.98,respectively,and both P＜0.001.In RPMI8226 cells(t=16.23,P＜0.001)and U266 cells(t=18.44,P＜0.001),compared with the ssi-NC group,the total apoptosis rate of the ssi-AURKA group was increased.The results of drug sensitivity experiments showed that there was a statistically significant difference in cell activity among the ssi-NC group,ssi-AURKA group,ssi-NC+B+D group,and ssi-AURKA+B+D group of RPMI8226 cells,with Ftime=618.82 and Ptime＜0.001;Fgroup=1 373.51,Pgroup＜0.001;Ftime×group=228.70,Ptime×group＜0.001.There was a statistically significant difference in cell activity among the ssi-NC group,ssi-AURKA group,ssi-NC+B+D group,and ssi-AURKA+B+D group of U266 cells,Ftime=617.83,Ptime＜0.001;Fgroup=1 352.84,Pgroup＜0.001;Ftime×group=155.81,Ptime×group＜0.001.The results of the Western blot experiment showed that the expression of E-cadherin was increased in the ssi-AURKA group of RPMI8226 cells,t=5.11,P=0.007.The expression of Vimentin,β-catenin,and Wnt3A were decreased,with t values of 2.86,3.87 and 7.56,respectively,with P values of 0.046,0.018 and 0.002;In U266 cells,the expression of E-cadherin was increased in the ssi-AURKA group,with t=10.02 and P＜0.001,while the expression of Vimentin,β-catenin,and Wnt3A were decreased,with t val-ues of 3.71,2.82,and 3.30 and P values of 0.020,0.047 and 0.030,respectively.Conclusions Inhibiting AURKA ex-pression can inhibit MM cell proliferation,lead to changes in cell cycle,induce cell apoptosis,and increase drug sensitivity;AURKA may affect the EMT process of MM cells and thereby influence cell proliferation and apoptosis through the Wnt/β-catenin signaling pathway.
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