摘要
目的探讨小分子干扰RNA(siRNA)对绒毛膜癌(绒癌)细胞中人类白细胞抗原G(HLA-G)基因表达的影响,了解HLA-G基因在绒癌发生、发展中的作用。方法设计并合成HLA-GsiRNA,分为两组。实验组以不同浓度(分别为1.0、2.5、5.0μg/L,下同)的HLA-GsiRNA转染HLA-G基因高表达的绒癌细胞系JEG-3细胞,对照组以等量阴性对照siRNA转染JEG-3细胞。实时荧光定量RT-PCR技术检测转染后JEG-3细胞中HLA-GmRNA的表达,HLA-GmRNA的表达以PCR循环数阈值(Ct值)表示;蛋白印迹法测定JEG-3细胞中HLA-G蛋白的表达;显微镜观察并计数转染后存活的JEG-3细胞数。结果HLA-GsiRNA转染JEG-3细胞后,明显下调JEG-3细胞中HLA-GmRNA及蛋白的表达水平。实验组不同浓度的HLA-GsiRNA转染后,JEG-3细胞的Ct值[分别为(20.67±0.02)、(21.37±0.03)、(21.43±0.02)个循环数],分别与对照组[分别为(20.33±0.01)、(20.37±0.02)、(20.40±0.03)个循环数]比较,差异有统计学意义(P<0.05)。上述浓度转染后,实验组JEG-3细胞的HLA-G蛋白表达水平[分别为0.42±0.03、0.37±0.02、0.12±0.04],分别与对照组[分别为1.69±0.23、1.62±0.31、1.36±0.22]比较,差异有统计学意义(P<0.01)。实验组JEG-3细胞生长较对照组明显受到抑制(P<0.05)。结论HLA-GsiRNA可以下调HLA-GmRNA及蛋白的表达水平,抑制细胞生长,表明HLA-G基因在绒癌的发生、发展中具有重要作用。
Objective To study the role of human leukocyte antigen-G （HLA-G） in the carcinogenesis and development of choriocarcinoma. Methods We designed and synthesized a double strand small interference RNA （siRNA） of HLA-G, then transfected it into a HLA-G overexpressed choriocarcinoma cell line, JEG-3 by lipofectine; HLA-G mRNA level was detected by real time RT-PCR; HLA-G protein level was detected by western blot. The living cells of JEG-3 were counted under microscope after transfection by siRNA. Results Double strand siRNA of HLA-G effectively downregulated the mRNA level and the protein level of HLA-G. The mRNA levels by Ct value at different concentration （ 1.0, 2. 5,5.0 μg/L） were 20. 67 ±0. 02, 21.37 ±0. 03,21.43 ±0. 02, respectively. They were significantly different compared with the control groups which were 20. 33 ± 0. 01, 20. 37 ± 0. 02,20.40 ± 0. 03, respectively （P 〈 0.05 ）. The protein levels were down regulated in different concentration compared with the control groups（0.42 ±0.03 vs 1.69 ±0.23, 0.37 ±0.02 vs 1.62 ±0.31, 0. 12 ±0.04 vs 1.36 ±0.22; P 〈0. 01 ）. Through downregulation of HLA-G expression, the growth of choriocarcinoma cells was inhibited.Conclusions Small interference RNA of HLA-G can effectively down regulate HLA-G gene in mRNA andprotein levels and inhibit tumor growth in choriocarcinoma. It shows that HLA-G plays an important role in the development of choriocarcinoma.
出处
《中华妇产科杂志》
CAS
CSCD
北大核心
2005年第8期549-552,共4页
Chinese Journal of Obstetrics and Gynecology
