Inhibitory Effect Of Par-4 Combined With Cisplatin On Human Wilms' Tumor Cells

Objective:To study the effects of prostate apoptosis response-4(Par-4)combined with cisplatin(CDDP)on the growth of hunan wilms' tumor cells SK-NEP-1 in vitro and in vivo.Methods:1.IHC-SABC was used to determine the expression of Par-4,GRP78 and BAX protein in both nephroblastoma and control groups.2.The plasmid pEGFP-Cl-Par-4 was transfected into nephroblastoma cell line SK-NEP-1.The flow cytometry analysis was used to assess the cell apoptosis.Indirect immunofluorescence double staining was employed to detect the distribution of Par-4 and GRP78.The protein levels of Par-4,GRP78,ATF-4 and BAX were analyzed by western blot.3.Wilms' tumor SK-NEP-1 cells were cultured and transplanted into subcutaneous tissues of BALB/c nude mice.Then,these nude mice were randomly divided into five groups,including Group Par-4+CDDP,Group Par-4,Group CDDP,Free-Par-4 group and blank control group.The volumes and weights of the transplanted tumors were measured.The expressions of Par-4,GRP78 and BAX protein in transplanted tumor tissues were detected by HE staining and immunohistochemical analysis.Results:1.Expressions of Par-4 and BAX were higher in nephroblastomas than that in normal renal tissues(P<0.05).Expression of GRP78 was lower in nephroblastoma than that in normal renal tissues(P<0.05).2.Cooperative synergy between Par-4 overexpression and CDDP inhibited SK-NEP-1 cells growth and induced cellular apoptosis.Indirect immunofluorescence staining showed co-localization of Par-4 and GRP78 in the endoplasmic reticulum.Par-4 overexpression increased the expressions of GRP78,ATF-4 and BAX protein.3.Among all of Group Par-4+CDDP,Group Par-4 and Group CDDP,growth inhibitions of nude micexenografts were obvious(P<0.05).The reduction of tumor weight in Par-4+CDDP group was more significant than these of Par-4 group and CDDP group.Nevertheless,between the tumor weights of Free-Par-4 group and blank control group,differences were no statistically significant.Increased expressions of Par-4,GRP78 and BAX protein were observed in Group Par-4+CDDP(P<0.05).Conclusion:Ectopic Par-4 combining with CDDP suppressed the growth of human wilms'tumor cells in vitro and in vivo,which might be related to the activation of ER apoptosis pathway and resulted in the interraction of extracellular Par-4 and GRP78.