^(125)I标记反义肽核酸对SKOV3细胞CerbB-2 mRNA和Her-2蛋白表达的抑制作用

Depressed Expression of CerbB-2 mRNA and Her-2 Protein in Human Ovarian Cancer Cell Line SKOV3 by ^(125)I Labeled Antisense Peptide Nucleic Acid

目的探讨利用脂质体转染125I标记的CerbB-2反义肽核酸(asPNA)阻断人卵巢癌细胞株SKOV3中CerbB-2mRNA翻译并抑制Her-2蛋白表达的作用。方法 (1)125I用CH-T法标记CerbB-2-asPNA,测定其标记率和放化纯。(2)分对照组、非标记分子探针组和125I标记反义肽核酸组,运用RT-PCR法检测各组细胞CerbB-2mRNA的翻译。(3)运用流式细胞仪检测各组细胞的Her-2蛋白表达率。结果 (1)125I标记CerbB-2-asPNA的标记率为(56.90±4.38)%,放射化学纯度为(87.00±0.99)%,放射性浓度为3.7MBq/ml,化学浓度为5.00μmol/L。(2)125I标记CerbB-2-asPNA作用SKOV3细胞24h后,SKOV3细胞CerbB-2在mRNA水平上明显下降,与SKOV3细胞对照组比较,差异有统计学意义(P<0.01)。(3)125I标记CerbB-2-asPNA作用SKOV3细胞24h后,细胞的Her-2蛋白表达率降低幅度较大,与SKOV3细胞组对比,两者间差异有统计学意义(P<0.01)。结论 125I-L-CerbB-2-asP-NA对人卵巢癌细胞株SKOV3CerbB-2基因表达在mRNA翻译和蛋白表达水平上均有抑制作用。

Objective To investigate the inhibitory effect of 125I labeled antisense peptide nucleic acid （ 125I-asPNA） transfected by liposome to block CerbB-2 mRNA translation and Her-2 protein expression in the human ovarian epithelial cancer cell lines SKOV3. Methods （ 1） The method of 125I labeled CerbB-2 antisense peptide nucleic acid with the CH-T was used; meanwhile its tagging ratio and radiochemical purity were detected. （ 2） Three experimental groups were divided in the experiment： the control group,no labeled molecule probes group and 125I labeled antisense peptide nucleic acid group; The translation of CerbB-2 mRNA in SKOV3 was detected by reverse transcription-polymerase chain reaction （ RTPCR） . （ 3） The expression ratio of Her-2 protein in cell by flow cytometry was detected. Results （ 1） 125 I-asPNA’s labeled ratio was （ 56. 90 ± 4. 38） % ,radiochemical purity was （ 87. 00 ± 0. 99） % ,radioactive concentration 3. 7 MBq/ml and chemistry concentration 5. 00 μmol/L. （ 2） After 24h treated by 125 I-L-asPNA,the CerbB-2 in SKOV3 cell lines declined obviously on mRNA level,their differences were significant （ P 0. 01） compared with those in the SKOV3 cell control group. （ 3） Flow cytometry indicated that the expression ratio of Her-2 protein in SKOV3 cell lines which was treated for 24 h by 125I-L-asPNA declined greatly,their differences were significant compared with those in the SKOV3 cell control group（ P 0. 01） . Conclusions 125I-L-asPNA can depress the CerbB-2 gene expression of the human ovarian epithelial cancer cell lines SKOV3 on the mRNA translation and protein expression level.