IL-4/IL-13诱导小鼠支气管上皮细胞中IL-13Rα2基因转录的调节  

Regulation of IL-13Rα2 transcription in mouse bronchial epithelial cells induced by IL-4/IL-13

作　　者：杨青[1] 况九龙[1] 饶纬华[2] 

机构地区：[1]江西医学院第二附属医院呼吸内科,江西南昌330006 [2]江西医学院第二附属医院分子医学研究中心

出　　处：《中国呼吸与危重监护杂志》2004年第1期15-18,共4页Chinese Journal of Respiratory and Critical Care Medicine

摘　　要：目的　了解支气管哮喘发病中白细胞介素 13受体α2 (IL 13Rα2 )基因转录的调节作用。方法　采用PCR扩增hIL 13Rα2基因启动子区不同长度的基因片段F1～F6 ,其中F1和F2含有信号转导子和转录活化子 6 (STAT6 )的结合区域。将F1～F6连入载体pGEM Teasy中 ,转化 ,获得菌落 ,经鉴定后将pGEM Teasy构建体的插入部分F1～F6连入增强载体pGL3中 ,转化 ,获得菌落 ,再以pGL3构建体 (F1～F6 ,mock)转染入小鼠支气管上皮细胞TGMBE 0 2 3。转染 2 4h后 ,给予小鼠IL 4 (mIL 4 ,10 μg/L)和小鼠IL 13(mIL 13,5 0 μg/L)处理 ,2 4h后进行荧光分析。 结果　 (1)基因组DNA(编号BCI0 0 1)IL 13Rα2启动子区的碱基序列与GeneBank中登录的序列完全一致。 (2 )各构建体转染后 ,对照组与mIL 4或mIL 13刺激组的相对荧光活性无显著差异 (P均 >0 .0 5 )。结论　IL 4、IL 13的刺激不影响hIL 13Rα2基因的转录活性。Objective To understand the regulation of interleukin-13 receptor α2 gene transcription in the pathogenesis of bronchial asthma.Methods Various lengths of gene segments(F1～F6)of promoter region in hIL-13Rα2 gene was amplified by PCR.F1 and F2 contained signal transducer and transcription activator 6 (STAT6) binding domain.The amplified DNA fragments were TA-cloned into pGEM-Teasy vector.After fluorescence and PCR screenings,suitable clones were selected to extract plasmid DNA and restriction enzyme digestion analysis were carried out.To exclude clones of mismatching during PCR amplification,the sequences of all clones were determined.The inserts of TA-clone were subcloned into pGL3 enhancer vector.Plasmids were extracted from these constructs and then were confirmed through restriction enzyme digestion analysis.Proper clones were selected for extracting plasmid on a large scale.pGL3 vector constructs (F1～F6 and mock) were transfected into mouse bronchial epithelial cell line TGMBE-02-3.After 24 hours,mIL-4 (10 μg/L ) or mIL-13 (50 μg/L) was added.Luciferase assay was performed 24 hours after stimulation.Results (1)The sequence of hIL-13Rα2 promoter region from genomic DNA (BCI001) was identical to the sequence from Gene Bank.(2) After transfection of all constructs,the relative fluroscence activities had no significant difference between control group and mIL-4/mIL-13 stimulation group.Conclusion mIL-4 or mIL-13 stimulation did not affect the transcriptional activity of hIL-13Rα2 gene.

关 键 词：支气管哮喘 白细胞介素-13受体α2 基因转录 白细胞介素-4 白细胞介素-13 

分 类 号：R562.25[医药卫生—呼吸系统]