Study On Apoptosis In Raji Cells And Its Molecular Mechanism Induced By Down-regulated Of PLK1 Expression

[Objective] Polo-like kinase 1(PLK1)is an enzyme involved in mitotic processes that regulates multiple aspects of cell-cycle control.In this study,RNA interference technology was used to observe the effects by down-regulated PLK1 expression on biological behaviors such as apoptosis of Raji cells in vitro and the formation of xenograft tumor in nude mice.Molecular mechanism of the effects was investigated.The study will explore the role of PLK1 in the pathogenesis of lymphoma and provide some experimental evidence for the study on the molecular mechanism and target therapy of lymphoma.[Methods] TUNEL,Hoechst 33258 staining,flow cytometry,animal xenografts,were used to observe the effects of down-regulated PLK1 expression on biological behaviors of Raji cells in vivo and nude mice.Then,q RT-PCR and Western blot were used to detect expression of mRNA and protein about apoptosis-related molecules and cyclerelated molecules such as Bax and CyclinD1 by decreasing PLK1 expression in Raji cells.[Results] The results of TUNEL showed that the apoptotic cell increased significantly in the experimental group(Raji-PLK1-shRNA)(P <0.0001)compared to negative control groups(Raji-NC-shRNA and Raji).Compared to negative control groups,the results of Hoechst 33258 staining showed that the percentage of apoptotic cell increased significantly in Raji-PLK1-shRNA(P <0.05).The results of flow cytometry showed that the the percentage of early apoptotic cell increased significantly in Raji-PLK1-shRNA(P <0.001),compared to negative control groups.It suggested that the down-regulation of PLK1 expression could increase the apoptosis rate in Raji cells.Compared to negative control groups,the results of xenograft tumor assay showed that the tumor volume decreased significantly in Raji-PLK1-shRNA.It suggested that the down-regulation of PLK1 expression in Raji cells could inhibit xenograft tumor growth.Compared to negative control groups,the results of Real time fluorescence quantitative PCR showed that mRNA expression level of wtp53,Bax,Rb and Caspase3 increased significantly and mRNA level of mutp53,CyclinD1,CDC2,Bcl-2 and Survivin decreased significantly in Raji-PLK1-shRNA(P<0.05).Compared to negative control groups,the results of Western blot showed that the expression of protein wtp53,Bax,Rb and Caspase3 increased significantly and the expression of protein mutp53,CyclinD1,CDC2,Bcl-2 and Survivin decreased significantly in Raji-PLK1-shRNA(P<0.05).[Conclusions] 1.Down-regulation of PLK1 expression can induce cell apoptosis in Raji cells of human lymphoma by activating wtp53,then down-regulation of Survivin.On the other hand,the active p53 can up-regulation of Bax and down-regulation of Bcl-2,then activate Caspase3.2.Down-regulation of PLK1 expression can induce cell cycle arrest in Raji cells by inhibiting genes of CDC2 and CyclinD1 and activating genes of p53 and Rb.