筛选反义寡核苷酸抑制哮喘大鼠CD4^+T细胞IL-4表达的研究  

The selection of antisense oligonucleotides to inhibit IL-4 expression by rat CD4^+T cells with asthma

作　　者：曹勇[1] 熊维宁[1] 徐永健[1] 张珍祥[1] 高宝安[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院呼吸内科,武汉430030

出　　处：《中国免疫学杂志》2006年第4期371-374,共4页Chinese Journal of Immunology

基　　金：国家自然科学基金资助项目(30300144)

摘　　要：目的:探讨不同序列的反义寡核苷酸对哮喘大鼠CD4+T细胞IL4表达的抑制作用。方法:采用脂质体转染技术,将不同的反义寡核苷酸(AS1:反外显子1;AS2:反外显子2;AS3:反翻译终止部位)和空白对照分别转入经免疫磁珠阴性分离的哮喘大鼠CD4+T淋巴细胞,细胞培养28小时后,用ELISA和半定量RTPCR分别检测细胞培养上清IL-4和细胞内IL4mRNA的水平。结果:不同组RTPCR结果(IL4/βactin相对吸光度):AS1、AS2、AS3及空白组分别为0.2615±0.1476、0.2885±0.1411、1.1012±0.3641及1.2068±0.3836(F=22.597,P<0.01)。ELISA检测培养上清液IL-4结果:AS1、AS2、AS3及空白组分别为13.800±7.233、15.329±7.358、52.643±12.075及58.286±14.100(F=34.976,P<0.01)。经AS1、2干预后细胞内IL4-mRNA和细胞培养上清IL4的水平均较AS3和空白组干预后IL-4的水平低(P<0.01)。结论:IL4反义寡核苷酸能够抑制哮喘大鼠CD4+T淋巴细胞IL-4和IL-4mRNA表达;不同序列IL-4反义寡核苷酸抑制作用存在差异。Objective：To investigate the effects of IL-4 antisense oligonucleotides of different sequences on the expression of IL-4 and IL-4 mRNA in CD4^＋T lymphocytes of asthma rats. Methods ：The CD4^＋T lymphocytes from asthma rats were purified by negative selection with a magnetic cell sort. Through liposome, different antisense oligonucleotides （AS-1 to translation initiation, AS-2 to Exon-2, AS-3 to translation terminal） and blank control were transfected into the CD4^＋T lymphocytes respectively. After 28 hours, IL- 4 in supernatant of cells culture and IL-4 mRNA in T lymphocytes were detected by ELISA and semi-quantitative RT-PCR respectively. Results：RT-PCR showed IL-4/β-actin （relative ratio of absorbance）：0. 261 5 ± 0. 1476 for AS-1, 0. 288 5 ± 0. 141 1 for AS-2,± 101 2 ± 0. 364 1 for AS-3, and 1. 206 8 ± 0. 383 6 for blank group respectively （ F = 22. 597, P 〈 0. 01 ）. The production of IL-4 in culture supernatant showed：13. 800 ±7.233 for AS-1,15. 329 ±7. 358 for AS-2,52. 643 ± 12. 075 for AS-3,and 58. 286 ± 14. 100 for blank group respectively（ F = 34. 976 ,P 〈 0.01 ）. The expression of IL-4 was significantly lower in the groups treated by AS-1 and AS- 2 than that in the group treated by AS-3 and blank（ P 〈 0. 01 ）. Conclusion：IL-4 antisense oligonucleotides can inhibit the expression of IL-4 and IL-4 mRNA in CD4^＋T lymphocytes of asthma rats ; the inhibitory effects of three kinds of antisense oligonucleotides are different signifieantly.

关 键 词：哮喘 IL-4 反义寡核苷酸 

分 类 号：R562.25[医药卫生—呼吸系统]