目录
摩羯座守护星
1 非特异性免疫疫苗通过调节Th1/Th2/Treg/Th17细胞平衡对
小鼠急性哮喘的作用 (3)
1.1 中文摘要 (3)
马诗李贺
1.2 英文摘要 (5)
1.3 前言 (7)
1.4 1.5 1.6 材料与方法 (10)
结果 (23)
讨论 (34)
炒牛蛙1.7 结论 (41)
1.8 参考文献 (42)
1.9 英汉缩略词对照表 (50)
2 致谢 (52)w2个世界
3 TIM-3及其在哮喘中的研究进展（综述） (53)
非特异性免疫疫苗通过调节Th1/Th2/Treg/Th17细胞平衡对小鼠
急性哮喘的作用
摘要孕检查
目的：哮喘是常见的呼吸系统疾病，对患者身心健康和生活质量造成严重损害，研究哮喘的发病机制和治疗手段具有重要的意义。异常的免疫反应在哮喘中发挥重要作用，Th1/Th2/Treg/Th17失衡是哮喘发生的重要机制，对免疫应答环节的人工干预有望成为哮喘治疗的有效途径。热休克蛋白HSP70和免疫刺激分子CD80能调节机体免疫平衡，在细胞和体液免疫过程中发挥重要作用。本实验运用HSP70和CD80构建非特异性免疫疫苗（HSP70/CD80 DNA疫苗）治疗急性哮喘小鼠，并研究该过程中Th1/Th2/Treg/Th17再平衡的作用机制，希望为哮喘的治疗提供新的方向。方法：本实验室以质粒pV AX1为载体构建HSP70/CD80 DNA疫苗。将40只BALB/c小鼠随机分为4组，即对照组、模型组、载体组和治疗组，每组10只动物。用鸡卵清蛋白（OV A）致敏构建急性小鼠哮喘模型，肌肉注射DN
A疫苗进行治疗。末次OV A激发后，测定雾化吸入乙酰甲胆碱的气道反应性；ELISA检测血清中IgE的含量；HE和AB-PAS染色检测肺组织的病理学改变；ELISA检测支气管肺泡灌洗液中细胞因子IFN-γ、IL-4、TGF-β和IL-17的表达；Real-time PCR检测肺组织中转录因子T-bet、GATA-3、t Foxp3和RORγ的表达。结果：我们发现：模型组相较对照组(1) 气道反应性明显升高（约1~2倍，P<0.05），(2) 血清IgE的浓度升高（约30%，P<0.05），(3) 肺组织中大量嗜酸性粒细胞和单核细胞浸润，支气管上皮杯状细胞增生肥大，管腔内大量黏液分泌。而治疗组相较模型组(1)
小鼠气道反应性明显下降（约1~1.5倍，接近实验对照组水平，P<0.05），(2)血清IgE浓度下降（约20%，P<0.05），(3) 肺组织炎症反应轻微，无明显杯状细胞增生，管腔黏液分泌较少。进一步研究该疫苗的作用机制检测发现细胞因子IFN-γ、IL-4、TGF-β和IL-17以及转录因子T-bet、GATA-3、Foxp3和RORγt的表达水平在激发哮喘和治疗哮喘过程中发生变化：模型组IFN-γ/IL-4和TGF-β/IL-17比值下降（相较对照组分别下降约50%和70%，P<0.05)，而疫苗治疗组IFN-γ/IL-4和TGF-β/IL-17比值回升（相较模型组分别升高约80%和250%，P<0.05)；模型组T-bet/GATA-3和Foxp3 /RORγt比值下降（分别下降约40%和60%，P<0.05)，而治疗组T-bet/GATA-3和Foxp3 /RORγt比值回升（分别均升高约3倍，P<0.05）。结论：本实验室构建的HSP70/CD80 DNA疫苗通过上调细胞因子IFN-γ、TGF-β和转录因子T-bet、Foxp3的表达，导致IFN-γ/IL-4、TGF-β/IL-17、T-bet/GATA-3、Foxp3/RORγt比值改变，实现Th1/Th2/Treg/Th17再平衡，从而发挥哮喘治疗的作用。
关键词：哮喘；疫苗；T细胞；转录因子；细胞因子
Non-specificity Immune Vaccine Regulates Asthma via Th1/Th2/Treg/Th17 Cell Balance in an Acute Mou Model一件什么的事
Abstract
Objective：Asthma is a common respiratory dia which impairs patients physically and mentally. Immune deregulation plays an important role in asthma, within which the Th1/Th2/Treg/Th17 imbalance contributes significantly to the pathogenesis of asthma. Intervention against abnormal immunity shows an effective path to regulate asthma. Heat shock protein 70 (Hsp70) and immune stimulator CD80 regulate cellular and humoral immune balances. Here we utilize a non-specificity immune vaccine(Hsp70/CD80 DNA vaccine) to cure mou acute asthma, and explore the mechanism of Th1/Th2/Treg/Th17 rebalancing during the treatment with this novel vaccine. Methods: pVXA1 plasmid vector was ud to construct a recombinant Hsp70/CD80 fusion gene vaccine. 40 BALB/c mice were delivered into four groups (for control, asthma-model, pVXA1-vector and vaccine treatment), each group contained 10 animals. OV A-induced acute asthma mice were treated with vaccine by intramscular injection. After latest OV A stimulation, the airway hyperresponsi
veness (AHR) was evaluated by inhaling methacholine; rum IgE was measured by ELISA; pathological changes was evaluated by HE and PAS staining; expression of cytokines (IFN-γ,IL-4, TGF-βand IL-17) in BALF was examined by ELISA; expression of transcription factors (T-bet, GATA-3, Foxp3 and RORγt) in lung tissue was assayed by Real-time PCR. Results: Compared with control group the asthma-model group changed as follows: (1) AHR incread about 1~2 folds, P<0.05; (2) rum IgE incread about 30%, P<0.05; (3) infiltration of eosinophils and monocytes incread in lung tissue, proliferation and hypertrophy of goblet cells incread, bronchial mucus deposited. Compared with asthma-model group, vaccine group changed as follows: (1) AHR decread about 1~1.5folds, rembling with the control
group, P<0.05; (2) rum IgE decread about 20%, P<0.05; (3) inflammatory changes decread in lung tissues, proliferation and hypertrophy of goblet cells and bronchial mucus were represd. The findings indicated that we successfully builded OV A-induced acute asthma-model, and the Hsp70/CD80 DNA vaccine contributed in the treatment of acute asthma. Furthermore, we explored the molecule mechanism of this vaccine treatment, focusing on the Th1/Th2/Treg/Th17 rebalancing. We found expression of cytokines (IFN-γ, IL-4, TGF-βand IL-17) and transcription factors (T-bet, GATA-3, Foxp3 and RORγt) changed through the vaccine treatment: IFN-γ/IL-4 and TGF-β/IL-17 dec
read about 50% and 70% individually in asthma group compared with control group,P<0.05; and incread about 80% and 250% in vaccine group compared with asthma-model group, P<0.05. T-bet/GATA-3 and Foxp3/ROR γt decread about 40% and 60% individually in asthma group compared with control group,P<0.05; and incread about three folds in vaccine group compared with asthma-model group, P<0.05. The findings suggested the Hsp70/CD80 DNA vaccine regulated asthma, depending on cytokines (IFN-γand TGF-β) and transcription factors (T-bet, Foxp3). Conclusion: The Hsp70/CD80 DNA vaccine was effective to asthma, which up-regulated the expression level of cytokines (IFN-γand TGF-β) and transcription factors (T-bet and Foxp3), resulted changes of the ratio (IFN-γ/IL-4, TGF-β/IL-17, T-bet/GATA-3, Foxp3/RORγt) to restore Th1/Th2/Treg/Th17 cell balancing. Key Words: asthma; vaccine; T cell; transcription factor; cytokine咏梅的诗句
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