Effects Of NPRL2 On Proliferation And Migration Of Human Prostate Cancercells In Vitro And In Vivo And Its Mechanism

To clarify the expression of NPRL2 in prostate cancer and its effect on the proliferation of prostate cancer DU145 and PC3 cells.Explore the metastasis of interfering NPRL2 expression on apoptosis and invasion in DU145 and PC3 prostate cancer cells;The metastasis of interfering NPRL2 expression on the invasion of DU145 and PC3 prostate cancer cells in vivo.Methods一 vitro experiment1.In vitro apoptosis detection: 1)the expression of NPRL2 in the normal prostate cell line rwep-1 and prostate cancer(DU145 and PC3 cell lines)were detected by rt-pcr and Western Blot.2)the PC3 prostate cancer and DU145 cell lines are slow NPRL2 sh virus RNA transfection,first of all,determined by fluorescence transfection is successful,then after the transfection DU145 and PC3 cell line respectively from the level of mRNA and protein expression of NPRL2 in quantity for testing;3)cck-8 method was used to detect the proliferation ability of DU145 and PC3 cell linesafter transfection;4)use flow cytometry to detect the apoptosis and cycle of DU145 and PC3 after transfection;5)Hoechst staining was used to detect the apoptosis of cells after transfection.6)the apoptotic proteins were detected by RT-PCR and Western Blot.7)use the scratch and Transwell to detect the invasion and metastasis of the cells after transfection;8)use Western Blot and rt-q PCR to detect the downstream PI3K/AKT/ gsk-3β signaling pathway of NPRL2 and its regulation;2.Detection of invasion and metastasis in vitro: 1)detection of invasion and metastasis of prostate cancer DU145 and PC3 cells by Transwell and scratchings;2)Western Blot and RT-PCR were used to detect NPRL2 and its downstream NF-kappa B/ MMP-9 signaling pathway.二 vivo experimentsIn vivo experiment: the tumor rate of cells in the body after transfection was detected,and the tumor reduction rate after the tumor was formed.HE staining was used to observe the morphological changes of tumor cells in the tumor.The changes of apoptosis-related proteins in the tumor were detected by immunofluorescence.The expression of apoptotic protein in the tumor was detected by immunohistochemistry.Western blot and rt-pcr were used to detect NPRL2 and apoptosis and metastatic proteins and genes.Results1.Compared with the normal prostate cell line,RWPE-1,the expression level of NPRL2 in the prostate cancer cell line DU145 and PC3 was significantly increased.2.After interfering with NPRL2 expression,the proliferation ability of human prostate cancer cells DU145 and PC3 decreased significantly.3.Interference with NPRL2 expression can effectively inhibit the apoptosis and cycle of human prostate cancer cells DU145 and PC3.4.Interfering with the expression of NPRL2 can effectively inhibit the expression of PI3K/AKT/ gsk-3 in the apoptotic signaling pathway of human prostate cancer cells DU145 and PC3.5.After interfering with the expression of NPRL2,it can effectively inhibit the invasion and metastasis of human prostate cancer cells DU145 and PC3 effectively,and have an effect on the related pathway NF-kappa B/ MMP-9.6.Internal experiments confirmed that interfering with NPRL2 expression can effectively inhibit tumor metastasis in nude mice.Conclusion: NPRL2 in prostate cancer DU145 and PC3 high expression,the expression of interference NPRL2 can effectively reduce the in vitro proliferation of prostate cancer DU145 and PC3,and can inhibit the invasion and metastasis,and in vivo experiments have established that interfere with the expression of NPRL2 can effectively inhibit DU145 andPC3 prostate cancer cells and the proliferation of the invasion and metastasis.