The Effect Of Abnormal Expression Of RCAS1 On The Biological Behavior Of Prostate Cancer Cells

Objective: Investigate the influence of the abnormal expression of RCAS1 on the biological behavior of prostate cancer cells and to reveal the possible mechanism of RCAS1 in promoting the occurrence and development of prostate cancer,we hope to determine the initial clinical value of RCAS1 in the early diagnosis of prostate cancer,and explore the possibility of RCAS1 targeted therapy in prostate cancer gene therapy.Methods: We cultured prostate cancer cell lines such as PC3,LNCa P,C42,DU145 in vitro,and detected the expression of RCAS1 by q PCR and Western Blot.RCAS1 high expression plasmid was constructed and determined by the gene sequencing technology,expression of RCAS1 was detected by q PCR and Western Blot after transient transfection to make sure that the plasmid was constructed successfully.Sh RCAS1 kit purchased was transfected transiently to the RCAS1 high expression cells,then we detected the expression of RCAS1 by q PCR and Western Blot,and finally the sh RCAS1 plasmid with the strongest inhibitory effect was screened.The final sh RCAS1 and RCAS1 high expression plasmid were transfected into RCAS1 high and low expression cell line respectively,using Muse flow cytometry to detect the effect in cell apoptosis and cycle;using the MTT method to detect the influence on cell proliferation;using the Transwell method to evaluate the change in invasion after 24-48 h.Results: 1.The expression of RCAS1 m RNA and protein was the lowest in PC3 cell line,and highest in LNCa P cell line,the expression of RCAS1 m RNA and protein in C42 was not significantly different from LNCa P;2.LNCa P and C42 cell lines were transfected with sh RCAS1-406,sh RCAS1-561,sh RCAS1-598,sh RCAS1-690,sh GAPDH,sh RCAS1-NC respectively,and sh RCAS1-406 was screened the strongest inhibitory effect of plasmid by q PCR and Western Blot;3.On the basis of p FLAG-CMV,we tried to constract RCAS1 high expressionplasmid named RCAS1-high,confirmed by double enzyme digestion and gene sequencing to make sure the correct sequence of inserted fragments without mutation.Expression of RCAS1 was detected by q PCR and Western Blot,and we found that the expression of RCAS1 m RNA and protein increased significantly,RCAS1-high was constructed successfully;4.After transfection of RCAS1-high in PC3 cells,the expression of RCAS1 was dramatically increased,which could significantly improve the ability of cell proliferation,apoptosis rate of PC3 cells was significantly lower than that of the control group,and promote the cell transferred from G0/G1 phase to S phase,Transwell experiment showed that the cell invasion of C42 and LNCa P cells enhanced;after transfection of sh RCAS1-406,the decreased expression of RCAS1 resulted in significant inhibition of cell proliferation,higher percentage of cells in G0/G1 phase compared with the control group,and weakened ability of invasion;Conclusion: 1.RCAS1 expression was the lowest in PC3 and highest in LNCa P and C42;2.sh RCAS1-406,which got the strongest inhibitory effect of RCAS1 plasmid,was screened;3.RCAS1 high expression plasmid,RCAS1-high,was constructed successfully;4.The high expression of RCAS1 significantly increased PC3 cell proliferation,promoted cell invasion,inhibited apoptosis,and promoted transformation from G0/G1 phase to S phase;and the low expression of RCAS1 could significantly reduce LNCa P,C42 cell proliferation,inhibit cell invasion,promote cell apoptosis,and the cells were arrested in G0/G1 phase.