小干扰RNA与反义寡核苷酸技术抑制肝星状细胞RhoA表达的效果比较  

Effect comparison of small interfering RNA and antisense oligonucleotide in inhibition of RhoA expression of rat hepatic stellate cells

作　　者：张玉峰[1] 叶坤英[1] 钟丹[2] 

机构地区：[1]河南中医药大学第二临床医学院,郑州450002 [2]井冈山大学医学院

出　　处：《山东医药》2017年第34期5-8,共4页Shandong Medical Journal

基　　金：河南省中医药科学研究专项课题(2014ZY02027)

摘　　要：目的对比观察小干扰RNA(siRNA)和反义寡核苷酸(ASODN)技术抑制肝星状细胞RhoA表达的效果。方法将培养的大鼠肝星状细胞株HSC-T6随机分为siRNA组和ASODN组,两组又分别分为A、B、C、D各4个亚组;siRNA组和ASODN组中的A亚组不转染质粒,C、D、B亚组分别采用siRNA、ASODN技术转染大鼠RhoA特异性Rat1、Rat2质粒及HK-A阴性对照质粒。转染48 h取各组细胞,以RT-PCR技术检测细胞中的RhoA、Ⅰ型胶原(ColⅠ)mRNA,酶联免疫吸附法检测细胞上清液中的透明质酸(HA)及层粘连蛋白(LN)。结果 siRNA组与ASODN组中RhoA、ColⅠmRNA均以C亚组表达量最低,组内A、C亚组比较差异有统计学意义(P均<0.05),组内其余亚组间比较差异无统计学意义(P均>0.05);siRNA组C亚组RhoA、ColⅠmRNA相对表达量低于ASODN组(P均<0.05),两组A、B、D同亚组间比较差异无统计学意义(P均>0.05)。siRNA组与ASODN组细胞上清液HA、LN水平均以C亚组最低,组内A、C亚组比较差异有统计学意义(P均<0.05),组内其余亚组间比较差异无统计学意义(P均>0.05);siRNA组C亚组细胞上清液HA、LN水平低于ASODN组(P均<0.05),两组A、B、D同亚组间比较差异无统计学意义(P均>0.05)。结论 siRNA和ASODN均可抑制大鼠HSC-T6细胞的RhoA表达,但siRNA下Rat1质粒所介导的RNA干扰技术相对于ASODN能更有效抑制HSC-T6细胞外基质HA、ColⅠ、LN的生成。Objective To compare the effects of small interfering RNA（siRNA） and antisense oligonucleotide（ASODN） in inhibition of Rho A expression of rat hepatic stellate cells. Methods Rat hepatic stellate cell line HSC-T6 was divided into the siRNA group and ASODN group,respectively,and then each group was separately divided into subgroups A,B,C,and D. Cells in the subgroup A of the siRNA group and ASODN group were cultured normally,subgroup B was transfected with HK-A negative control plasmid,subgroup C with Rat1,and subgroup D with Rat2. After transfection for 48 h,the mRNA expression of Rho A and ColⅠin HSC-T6 cells of each group was detected by reverse-transcription polymerase chain reaction（RT-PCR）,respectively; the content of hyaluronic acid（HA） and laminin（LN） in culture serum was measured by specific ELISA. Results The expression of Rho A mRNA and Col I mRNA in the subgroup C of siRNA group and ASODN group was the lowest,and significant difference was found between subgroup A and subgroup C（both P〈0. 05）,but no significant difference was found between the rest of subgroups（all P〉0. 05）. The mRNA expression of Rho A and Col Ⅰ in the subgroup C of the siRNA group was lower than that in the ASODN group（P〈0. 05）.There was no significant difference in the subgroups A,B,and D between the siRNA group and ASODN group（all P〉0. 05）. The levels of HA and LN in the supernate of subgroup C of the siRNA group and ASODN group were the lowest,and significant difference was found between subgroup A and subgroup C（both P〈0. 05）,but no significant difference was found between the rest of subgroups（all P〉0. 05）. The levels of HA and LN in the supernate of subgroup C of the siRNA group were lower than those in the ASODN group（both P〈0. 05）. There was no significant difference in the subgroups A,B,and D between the siRNA group and ASODN group（all P〉0. 05）. Conclusion Both siRNA and ASODN can inhibit Rho A expression of rat HSC-T6 cells,and RNA interference mediated

关 键 词：小干扰RNA 反义寡核苷酸 RHOA 透明质酸 Ⅰ型胶原 层粘连蛋白 肝星状细胞 

分 类 号：R365[医药卫生—病理学] Q522[医药卫生—基础医学]