Expression Of Survivin And Livin In Gastric Cancer And Induction Of Apoptosis Of BGC-823 Cells By SiRNA-mediated Silencing Of The Survivin Gene

AIM:1.To study the expression of Survivin,Livin in gastric cancer tissues and their correlation with Bcl-2,Ki-67,clinical pathological features and prognosis.2. To investigate the expression of Survivin and Livin in the human gastric cancer cell line BGC-823 and SGC-7901.3.To investigate the effect of a specific small Survivin interfering RNA(SiRNA) on the expression of Survivin and cell apoptosis in human gastric carcinoma cell line BGC-823.METHORDS:The experiment involves two parts.Part 1:Immunohistochemical staining(S-P method) for Survivin,Livin,bcl-2 and Ki-67 was performed in 70 cases of gastric cancer tissues,48 cases of adjacent tissues.Immunocytochemical staining(S-P method) was performed in BGC-823 cells and SGC-7901.Part 2: To knockdown Survivin expression,a small interfering RNA targeting against Survivin was synthesized and transfected into BGC-823 cells with lipofectamine TM2000.The expression of Survivin protein was detected by Immunocytochemical staining(S-P method)and Westernern blot analysis.The effect of Survivin SiRNA and SiRNA combined with 5-Fu on cell apoptosis was detected by TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling).RESULTS:Part 1:1.Immunohistochemical analysis showed Survivin and Livin were detected both in the cytoplasm and/or nucleus of GC cells.The rates of high expression of both N-Survivin and C- Survivin in GC tissues were 28.57%. The rates of high expression of N-Livin and C- Livin in GC tissues were 42.86% and 40%respectively.Bcl-2 was expressed in the cytoplasm of GC cells.The positive rate of Bcl-2 in GC was 52.86%.The positive rates of Survivin,Livin and Bcl-2 in GC tissues were significantly higher than those in adjacent tissues (P＜0.01).2.There was a statistical difference of the expression of C-Survivin among well,moderate and poor differentiated adenocarcinama(P＜0.05).The expression of C-Survivin in papillary adenocarcinoma was significantly higher than that in poor differentiated adenocarcinoma(P＜0.01).The high expression of C-Survivin and N-Survivin positively correlated with distant metastasis(P＜0.05).The high expression of N-Survivin positively correlated with infiltrating growth in GC(P＜0.05).There was a statistical difference of the expression of C-Livin among well,moderate and poor differentiated adenocarcinoma(P＜0.05). The expression of C-Livin in well differentiated adenocarcinoma was significantly higher than that in poor differentiated adenocarcinoma,nucoid carcinoma(P＜0.05).The positive rate of bcl-2 was significantly related with clinical TMN stage (P=0.05).3.The 5 years survival rate in N-Survivin high expression group (40%) was positively lower than that in the N-Survivin low and negative expression groups(66%,P=0.04).4.Of 70 cases of GC,The average index of Ki-67 was 34.47±24.58,which was significantly higher than that in adjacent tissues(P＜0.01).Ki-67 indexes were significantly higher in N-Survivin,N-Livin high and low expression groups than that in N-Survivin,N-Livin negative expression groups in GC(P＜0.05).5.There was no correlation between the expression of Surviving,Livin and bcl-2 proteins.6.Multivariate Cox regression analysis indicated that clinical TMN stage was the only independent prognostic factor that affected survival in GC.7.Immunocytochemical analysis showed Survivin and Livin were detected both in the cytoplasm and nucleus of BGC-823 and SGC-7901 cells.Part 2:RNA interference could efficiently suppress the Survivin expression in BGC-823 cells.At 48 h after transfection,the Survivin protein expression was significantly inhibited by Immunocytochemical and Westernern blot analysis.SiRNA inhibiting Survivin expression increased apoptosis rate of BGC-823 cells.SiRNA-Survivin combined with 5-Fu strongly sensitized BGC-823 cells to apoptosis. independently.The high expression of N-Survivin was positively correlated with proliferation,distant metastasisin GC.The high expression of N-Survivin can serve as an important parameter of malignant degree and prognosis of GC.2. Survivin and Livin were expressed both in the cytoplasm and nucleus of BGC-823 and SGC-7901 cells.3.Si-RNA could efficiently silence endogenous Survivin expression,which was associated with a strongly increased apoptotic rate of BGC-823 cells.SiRNA combined with 5-Fu strongly sensitized BGC-823 cells to apoptosis.