Effects Of Target Inhibiting NACK On Proliferation And Apoptosis Of SK-N-BE(2) Cells And Related Mechanisms In Neuroblastoma

Objective Neuroblastoma is a common extracranial solid tumor in children.Notch signaling pathway is closely related to the occurrence of neuroblastoma.Notch activation complex kinase is an important rate limiting enzyme in Notch signaling pathway.The purpose of this study was to investigate the effects of down-regulation of NACK expression by siRNA transfection on proliferation and apoptosis of neuroblastoma SK-N-BE(2)cells and its mechanism.Methods Human neuroblastoma cells SK-N-BE(2)are routinely cultured.Design and synthesize NACK siRNA and NC siRNA according to relevant experiments,transfect SK-N-BE(2)cells through liposome-mediated siRNA,and divide into SK-N-BE(2)cells according to different treatment methods: experimental group(NACK siRNA group),negative control group(NC siRNA group)and control group.Observe the fluorescence of the transfected cells under a fluorescent microscope;qRT-PCR and Western blot methods to detect the silencing efficiency of the NACK gene after transfection;CCK-8 method was used to detect the cell proliferation inhibition rate of each group.Flow cytometry was used to analyze the apoptosis of each group.Finally,after NACK knockdown,the expression of downstream target genes Hes1,c-Myc and n-Myc in Notch1 signaling pathway was detected by qRT-PCR and Western blot.Results Liposome-mediated siRNA transfected human neuroblastoma SK-N-BE(2)cells showed green fluorescence under a fluorescence microscope.In the experimental group,the expression level of NACK m RNA and protein of cells which successfully transfected with NACK siRNA was significantly reduced(P <0.05).Compared witch the negative group and the control group,CCK-8 method detected that the cell proliferation in the experimental group was significantly inhibited(P <0.01),and the flow cytometry detected that the apoptosis in the experimental group was also significantly increased(P<0.01).Western blot and qRT-PCR detected protein levels and m RNA expression of related target genes Hes1,c-Myc,and n-Myc downstream of the Notch1 signaling pathway were significantly lower than those of the negative control group and the control group,the difference was statistically significant(P <0.05)Conclusion After neuroblastoma SK-N-BE(2)cells which transfected with liposome-mediated NACK siRNA,the expression of NACK gene was targeted to suppress,which further inhibited Notch1 signaling pathway,resulted in cell proliferation was inhibited and cell apoptosis was increased,and on this basis,exerts an anti-neuroblastoma effect.