T-bet基因转染对哮喘小鼠脾脏Th1/Th2平衡的影响  

Effect of Intranasal T-bet Plasmid Transfer on Th1/Th2 Balance in Spleen MNC in a Established Murine Allergic Asthmatic Model

作　　者：蓝丹[1] 檀卫平[1] 夏焱[1] 吴葆菁[1] 麦贤弟[1] 李晓圆[1] 黄花荣[1] 

机构地区：[1]中山大学附属第二医院儿科,广东广州510120

出　　处：《中山大学学报（医学科学版）》2010年第1期74-78,共5页Journal of Sun Yat-Sen University:Medical Sciences

基　　金：广东省自然科学基金(031668A;06021301A)

摘　　要：【目的】观察气道内T-bet基因转染对哮喘小鼠脾脏淋巴细胞Th1/Th2平衡、T-bet及GATA-3mRNA表达的影响。【方法】将C57BL/6小鼠32只随机分为4组,每组8只,分别为正常对照组(A组)、哮喘模型组(B组)、空质粒干预组(C组)和T-bet质粒干预组(D组)。卵白蛋白(OVA)抗原溶液腹腔注射致敏,滴鼻造模。空质粒组和T-bet质粒干预组OVA激发24h前,分别经鼻滴入50μg空质粒、重组T-bet质粒,对照组用生理盐水代替OVA。最后一次滴鼻激发48h后处死小鼠,流式细胞仪检测各组实验小鼠的脾脏淋巴细胞的Th1/Th2,RT-PCR检测小鼠脾脏淋巴细胞的T-bet及GATA-3mRNA表达。【结果】哮喘小鼠气道转染pcDNA3-T-bet质粒后:①流式细胞仪检测发现,脾脏Th1百分率较哮喘模型组显著升高[(2.29±1.55)%vs.(1.93±1.14)%,P﹤0.05],Th2百分率显著降低[(0.93±0.64)%vs.(1.63±0.59)%];②RT-PCR检测发现脾脏淋巴细胞转录因子T-bet mRNA表达水平显著增加(0.53±0.027vs.0.28±0.035,P﹤0.05),GATA-3mRNA表达水平显著降低(0.24±0.022vs.0.58±0.038,P﹤0.05)。而pcDNA3空质粒干预后与哮喘模型组比较无显著差异。【结论】气道内转导pcDNA3-T-bet质粒后,可显著上调哮喘小鼠体内T-betmRNA表达,下调GATA-3mRNA表达,有效改善哮喘小鼠体内Th1/Th2比例失衡,从而抑制哮喘小鼠的气道炎症,为哮喘的T-bet基因治疗提供依据。【Objective】 This study was designed to determine Th1,Th2 cell numbers and investigate T-bet mRNA,GATA-3 mRNA expression of spleen MNC in a murine asthmatic model which intended to understand effect of airway T-bet plasmid gene transfer on Th differentiation.【Methods】 A mouse asthmatic model was established by sensitization with ovalbumin （OVA）.Thirty-two C57BL / 6 mice were divided into four groups （8 mice in each group）： the normal control group （group A）,the asthmatic model group （group B）,the pcDNA3 plasmid group （group C）,the pcDNA3-T-bet group （group D）.All animals were sensitized and challenged with OVA,except group A normal saline was applied.The group C was intranasally administered 50 μg pcDNA3 plasmid at 24 h before intranasal challenges,and the 50 μg pcDNA3-T-bet plasmid for the mice of group D.We investigated Th1 and Th2 cell numbers by FACS and T-bet,GATA-3mRNA expression of spleen mononuclear cells （MNC） by semi-quantitative PCR in the four groups.【Result】 Th1 percent in spleen MNC of pcDNA3-T-bet treated mice was significantly increased （[2.29 ± 1.55]% vs.[1.93 ± 1.14]%,P﹤0.05）,while Th2 percent was significantly decreased （[0.93 ± 0.64]% vs.[1.63 ± 0.59]%）,compared with that of the asthmatic control group mice by FACS.Spleen MNC was detected a high level of T- bet mRNA expression （0.53 ± 0.027 vs.0.28 ± 0.035,P﹤0.05） and a low level of GATA-3 mRNA expression （0.24 ± 0.022 vs.0.58 ± 0.038,P﹤0.05） after pcDNA3-T-bet treatment by RT-PCR.There was no significant change between the pcDNA3 plasmid group and the asthmatic model group.【Conclusion】 The intranasal transfer of pcDNA3-T-bet plasmid was effective in modulating the imbalance of Th1 / Th2 in mice asthma model,which provides a novel therapeutic strategy for transferring transcriptional factor in allergic asthma.

关 键 词：哮喘 T-BET GATA-3 Th1 TH2 

分 类 号：R725.6[医药卫生—儿科]