| Objective:(1)To investigate the expression and clinical significance of SCN4B(Voltage-gated sodium channel ?4 subunit)in prostate cancer(PCa)tissues,adjacent tissues and cells;(2)To investigate the effect of lentivirus transfection on the proliferation,migration and invasion ability in prostate cancer;(3)To explore the potential mechanism of SCN4B in EMT progression of prostate cancer.Methods:(1)Immunohistochemistry(Elivision TMplus method)staining to detect the expression of SCN4B in prostate cancer tissues and corresponding adjacent tissues(103 cases),the relationship between SCN4B expression and clinicopathological characteristics and prognosis of patients with prostate cancer was analyzed.(2)Real-time reverse transcription-polymerase chain reaction(RT-qPCR)and western blotting were used to detect the mRNA and protein levels of SCN4B in three prostate cancer cell lines(DU145,PC-3,LNCAP).(3)The expression of SCN4B in prostate cancer cells DU145 and LNCAP was enhanced or inhibited by lentivirus mediated exogenous force,and the transfection effect was verified by RT-qPCR and Western blotting test.(4)To construct the stable overexpression of SCN4B DU145 and SCN4B knocking down LNCAP prostate cancer cell lines,the effect of SCN4B on the proliferation,migration and invasion of prostate cancer cells was verified by cell colony cloning experiment,CCK-8 experiment,cell scratch test and Transwell invasion experiment.(5)A subcutaneous xenograft model of prostate cancer in nude mice was established to prepare exogenous forced overexpression of SCN4B cells and RNA interference silencing of SCN4B cells,the successfully transfected cells were inoculated under the skin of nude mice,and the tumor growth rate and volume were recorded regularly,all nude mice will be killed 35 days later.(6)Western blotting and immunohistochemical staining were used to detect the expression of EMT marker molecules in prostate cancer cells before and after transfection.Results:(1)Immunohistochemical staining showed that the SCN4B staining intensity of prostate cancer tissues was significantly lower than that of adjacent tissues,the expression of SCN4B in cancer was related to Gleason grade,T stage and M stage,and the under-expression of SCN4B was closely connected to bad prognosis(P<0.05).(2)RT-qPCR and Western blotting showed that SCN4B expressed differently at the levels of RNA and protein in prostate cancer cells(highly malignant DU145,moderately malignant PC-3,and low malignant LNCAP),and the expression of SCN4B decreased with the increase of cell malignancy(P <0.05).(3)Overexpression of SCN4B significantly promoted the proliferation,migration,and invasion of prostate cancer cells,while downregulation of SCN4B significantly inhibited the proliferation,migration,and invasion of prostate cancer cells.(4)In animal experiments,the tumor volume was smaller in the group with high SCN4B overexpression,while the tumor volume was larger in the group with low SCN4B down-expression,and the growth rate was faster than in the group with overexpression.(5)Western blotting results showed that after RNA interference with SCN4B up-regulated expression,E-cadherin expression was down-regulated in Sh-SCN4B-1of prostate cancer cell,while N-cadherin,vimentin were up-regulated.After SCN4B overexpression,E-cadherin expression was increased in the prostate cancer cell PCDH-SCN4B,while N-cadherin,vimentin were down-regulated and EMT process was obstructed.At the same time,the above views were verified by immunohistochemistry of two groups of tumor tissues.Conclusion:(1)SCN4B was lowly expressed in prostate cancer tissues and was associated with higher TNM stages and shorter survival of prostate cancer.(2)SCN4B can be used as a therapeutic target to inhibit the proliferation,migration and invasion of prostate cancer.(3)Up-regulated of SCN4B can inhibit EMT progression in prostate cancer. |
PDF Full Text Request