CAJ | 학술논문

目的通过siRNA的方法体内靶向抑制哮喘小鼠T细胞非受体酪氨酸蛋白激酶Lck的基因表达,研究Lck特异性siRNA对哮喘小鼠的治疗作用.方法化学法合成小鼠T细胞Lck基因特异的siRNA片段,以in vivo-jetPEITM作为转染试剂,将合成的RNA片段与转染试剂混合后通过尾静脉注射到哮喘小鼠体内,48 h后处死小鼠,ELISA法检测肺泡灌洗液(BALF)中细胞因子IL-4、IL-17的含量；HE染色观察肺组织病理改变情况；免疫组织化学法检测肺组织中Lck表达的变化；Western Blot法检测肺组织匀浆中Lck蛋白的含量.结果 SiRNA干扰组哮喘小鼠BALF中IL-4、IL-17[( 142.62±15.57) pg/ml,(74.18 ±5.36)pg/ml]的水平较哮喘组[(234.68±11.15) pg/ml,(96.76±8.28) pg/ml]明显下降(P＜0.01),肺组织炎症病理改变有所减轻,免疫组化提示肺组织中Lck蛋白表达下降,肺组织匀浆中Lck蛋白的表达下降(P＜0.05).结论 Lck特异性siRNA可以降低哮喘小鼠肺组织中炎症因子IL-4、IL-17的水平,减轻肺部炎症反应,具有一定的治疗作用.
목적 통과siRNA적방법체내파향억제효천소서T세포비수체락안산단백격매Lck적기인표체,연구Lck특이성siRNA대효천소서적치료작용.방법 화학법합성소서T세포Lck기인특이적siRNA편단,이in vivo-jetPEITM작위전염시제,장합성적RNA편단여전염시제혼합후통과미정맥주사도효천소서체내,48 h후처사소서,ELISA법검측폐포관세액(BALF)중세포인자IL-4、IL-17적함량；HE염색관찰폐조직병리개변정황；면역조직화학법검측폐조직중Lck표체적변화；Western Blot법검측폐조직균장중Lck단백적함량.결과 SiRNA간우조효천소서BALF중IL-4、IL-17[( 142.62±15.57) pg/ml,(74.18 ±5.36)pg/ml]적수평교효천조[(234.68±11.15) pg/ml,(96.76±8.28) pg/ml]명현하강(P＜0.01),폐조직염증병리개변유소감경,면역조화제시폐조직중Lck단백표체하강,폐조직균장중Lck단백적표체하강(P＜0.05).결론 Lck특이성siRNA가이강저효천소서폐조직중염증인자IL-4、IL-17적수평,감경폐부염증반응,구유일정적치료작용.
Objective Using the technology of siRNA to inhibit the gene expression of no-receptor　tyrosine protein kinase Lck in T cells of asthmatic mice,and to study the therapeutic effect of Lck specific　siRNA in asthmatic mice.Methods Receptor tyrosine protein kinase Lck specific siRNA fragments were　taken from chemosynthesis.In vivo-jetPEITM was used to transfect the siRNA into mice body through tail　vein injection.The mice were killed 48 hours later,and the levels of IL-4,IL-17 in bronchoalveolar lavage　fluid (BALF) were detected with respondent ELISA kits.The change of inflammatory histopathology in lung　was observed with H.E.staining.The expression of Lck in lung was detected with immunohistochemistry　(IHC),and the level of Lck in lung tissue homogenate was detected with Western Blot.Results Compared with asthmatic group[ (234.68 ± 11.15 ) pg/ml,( 96.76 ± 8.28 ) pg/ml],the levels of IL-4,IL-17　[ (234.68 ± 11.15)pg/ml,(96.76 ±8.28) pg/ml] in the BALF of siRNA interference group decreased,　and the inflammation in the lung relieved.IHC indicated that the expression of Lck in lung decreased and　the level of Lck in lung tissue homogenate decreased ( P ＜ 0.05 ).Conclusions Lck specific siRNA　could reduce the level of IL-4,IL-17 in the lung tissues of asthmatic mice,and relieve the inflammatory reaction in lung.