真核始动因子4E反义寡核苷酸对人膀胱癌BIU-87细胞中eIF4E和肝素酶表达的影响

目的 探讨真核始动因子4E(eIF4E)反义寡核苷酸(ASODN)对人膀胱癌BIU-87细胞株中eIF4E及肝素酶(HPA)蛋白、mRNA表达的影响. 方法 采用脂质体介导方法分别将2.5、5.0及7.5 μg/ml eIF4E ASODN转染膀胱癌BIU-87细胞,并设立细胞对照组(不转染)、空白对照组(转染空脂质体)及无关对照组(转染无关对照ASODN),分别转染24、48及72 h,采用原位杂交、免疫细胞化学方法分别检测各组BIU-87细胞中eIF4E及HPA蛋白、mRNA表达. 结果 eIF4E ASODN各转染组细胞中eIF4E蛋白及mRNA表达量均较对照组降低(蛋白:2.5 μg/ml组分别为3.55±0.52、3.52 ±0.51、3.22 ±0.44；5.0 μg/ml组分别为3.43±0.47、3.41±0.46、3.19±0.41；7.5 μg/ml组分别为2.36±0.39、2.33±0.37、2.05±0.30.mRNA:2.5 μg/ml组分别为3.19±0.41、3.13±0.39、2.90±0.38；5.0μg/ml组分别为3.07±0.39、2.94 ±0.38、2.27±0.37;7.5 μg/ml组分别为2.22±0.37、2.06±0.30、1.95 ±0.29),与细胞对照组、空白对照组和无关对照组相应时间段、相应浓度组两两比较差异均有统计学意义(P＜0.05),且具有时间和浓度依赖性.eIF4E ASODN各转染组细胞中HPA蛋白及mRNA表达量均较对照组降低(蛋白:2.5 μg/ml组分别为4.44±0.55、4.40±0.54、3.99±0.52;5.0μ g/ml组分别为4.41 ±0.55、4.21±0.53、3.77±0.50；7.5 μg/ml组分别为4.02±0.52、3.98±0.52、2.32±0.37.mRNA:2.5μg/ml组分别为4.12±0.51、3.75±0.50、3.63±0.45；5.0 μg/ml组分别为4.00 ±0.51、3.71±0.50、3.54 ±0.44;7.5 μg/ml组分别为3.87±0.52、3.61 ±0.49、2.08 ±0.30),与细胞对照组、空白对照组和无关对照组两两比较差异均有统计学意义(P＜0.05),且具有时间和浓度依赖性.eIF4E ASODN对HPA蛋白、mRNA表达的抑制效应和对eIF4E蛋白、mRNA表达的抑制效应呈正相关关系(HPA蛋白r=9.23,mRNA r=9.59,P值均＜0.01). 结论eIF4E ASODN可下调膀胱癌BIU-87细胞中eIF4E、HPA蛋白及mRNA的表达,抑制肿瘤细胞的生长、浸润及转移.

Effects of antisense oligodeoxyribonucleotides of eIF4E on the expression of eIF4E and heparanase in human bladder carcinoma BIU-87 cells

Objective To explore the influence on the expression of elF4E and heparanase by antisense oligodeoxyribonucleotides (ASODN) transfection in human bladder carcinoma BIU-87 cells.Methods After transfecting the 2.5,5.O,7.5 μg/ml eIF4E ASODN into BIU-87 cells,at 24 h,48 h and 72 h,a cell control group (no transfection),a blank control group (empty liposomes) and a non-sense control group (transfected with non-sense ASODN) were established.The expression of eIF4E,HPA and mRNA were detected by in situ hybridration.The expression of eIF4 and HPA protein were detected by immunocytochemistry.SPSS 13.0 statistical software was used for statistical analysis.Results The expression quantity of eIF4E protein and mRNA were lower in the experimental groups ( protein:2.5 μg/ml ASODN treated 24 h,48 h,72 h were:3.55 ±0.52,3.52 ±0.51,3.22.±0.44 respectively; 5.0 μg/ml group:3.43 ±0.47,3.41 ± 0.46,3.19 ± 0.41 respectively ; 7.5 μg/ml group:2.36 ± 039,2.33 ± 0.37,2.05 ± 0.30 respectively.mRNA:2.5 μg/ml treated 24 h,48 h,72 h were:3.19 ±0.41,3.13 ±0.39,2.90 ±0.38 respectively ; 5.0 μg/ml group:3.07 ± 0.39,2.94 ± 038,2.27 ± 0.37 respectively ; 7.5 μg/ml group:2.22 ± 037,2.06 ± 0.30,1.95 ± 0.29 respectively.All data were less than those in the control groups (P ＜0.05).The expression quantity of HPA protein and mRNA were lower in experimental groups (protein:2.5 μg/ml ASODN treated 24 h,48 h,72 h were:4.44 ±0.55,4.40 ±0.54,3.99 ±0.52 respectively; 5.0 μg/ml group:4.41 ±0.55,4.21 ±0.53,3.77 ±0.50 respectively; 7.5 μg/ml group:4.02 ±0.52,3.98 ±0.52,2.32 ±0.37 respectively.mRNA:2.5 μg/ml treated 24 h,48 h,72 h were:4.12 ±0.51,3.75 ± 0.50,3.63 ± 0.45 respectively ; 5.0 μg/ml group:4.00 ± 0.51,3.71 ± 0.50,3.54 ± 0.44respectively ; 7.5 μg/ml group:3.87 ± 0.52,3.61 ± 0.49,2.08 ± 0.30 respectively).All data were less than in control groups ( P ＜ 0.05 ).There was a positive correlation on expression of HPA protein and eIF4E protein by inhibitory effect of eIF-4E ASODN (protein r=9.23,mRNA r=9.59,P ＜0.01).Conclusion eIF-4E ASODN might be used to inhibit the expression of eIF-4E gene and HPA gene in bladder cancer BIU-87 cells.