靶向DEK适配体DTA-64可通过阻断TGF-β1信号通路抑制支气管哮喘小鼠气道上皮细胞的上皮间质转化  

DEK-targeting aptamer DTA-64 inhibits epithelial-mesenchymal transition of airway epithelial cells in bronchial asthmatic mice via blocking TGF-β1 signaling pathway

作　　者：宋艺兰 王知广 姜京植 朴艺花 李莉[1,2] 徐畅 朴红梅 李良昌 延光海[1,2] SONG Yilan;WANG Zhiguang;JIANG Jingzhi;PIAO Yihua;LI Li;XU Chang;PIAO Hongmei;LI Liangchang;YAN Guanghai(Jilin Key Laboratory of Immune and Targeting Research on Common Allergic Diseases,Yanbian University,Yanji 133002;Department of Anatomy,Histology and Embryology,Medical College,Yanbian University,Yanji 133002;Postdoctoral Programme,Research Center,Affiliated Hospital of Yanbian University,Yanji 133000;Department of Respiratory Medicine,Affiliated Hospital of Yanbian University,Yanji 133000;Department of Intensive Care Unit,Affiliated Hospital of Yanbian University,Yanji 133000,China)

机构地区：[1]吉林省过敏性常见疾病免疫与靶向研究重点实验室,吉林延吉133002 [2]延边大学医学院解剖学教研室,吉林延吉133002 [3]延边大学附属医院博士后工作站,吉林延吉133000 [4]延边大学附属医院呼吸内科,吉林延吉133000 [5]延边大学附属医院重症医学科,吉林延吉133000

出　　处：《细胞与分子免疫学杂志》2022年第2期103-109,共7页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(81970018,81860729);中央引导地方科技发展资金吉林省基础研究专项(202002020JC);吉林省博士后基金(2020498)

摘　　要：目的 本研究旨在探讨靶向DEK适配体64(DTA-64)对卵清蛋白(OVA)诱导的哮喘小鼠气道炎症和EMT的抑制作用。方法 选取雌性BALB/c小鼠32只(8周龄),随机分为PBS组、 OVA模型组、 DTA-64组(1μg/只)和对照适配体组,每组8只。肺组织HE染色检测气道周围炎性细胞浸润,免疫组织化学染色检测气道周围DEK表达;ELISA检测血清IgE以及支气管肺泡灌洗液(BALF)的2型辅助T(Th2)细胞因子白细胞介素4(IL-4)、 IL-5、 IL-13以及Th1细胞因子γ干扰素(IFN-γ)水平。Western blot法检测上皮间质转化(EMT)相关蛋白α平滑肌肌动蛋白(α-SMA)、兔抗锌指转录因子(Snail+Slug)、波形蛋白(vimentin)和上皮钙黏素(E-cadherin)相关蛋白以及转化生长因子β1/Smad(TGF-β1/Smad)、丝裂原活化蛋白激酶(MAPK)、磷脂酰肌醇3激酶(PI3K)、蛋白激酶B(AKT)和哺乳动物雷帕霉素靶蛋白(mTOR)的蛋白表达;流式细胞术检测肺单细胞悬液中α-SMA,探讨DTA-64对哮喘EMT和气道重塑中的作用机制。结果 DEK蛋白在哮喘模型小鼠肺组织高表达,在DTA-64组中低表达;DTA-64能够降低气道周围嗜酸性粒细胞和中性粒细胞的浸润,下调血清OVA特异性IgE和BALF中IL-4、 IL-5、 IL-13水平,上调IFN-γ水平;DTA-64还能降低肺组织vimentin、α-SMA、 Snail+Slug表达,上调上皮标志物E-cadherin表达;DTA-64能抑制TGF-β1及其下游经典通路Smad2/3、 Smad4表达,以及非经典通路ERK1/2、 p38 MAPK、 JNK和PI3K/AKT/mTOR的磷酸化水平。结论 DTA-64可能通过阻断TGF-β1/Smad、 MAPK、 PI3K信号通路抑制OVA诱导的哮喘小鼠气道炎症和EMT的进程,从而抑制哮喘气道重塑。Objective To investigate the inhibitory effect of DEK targeting aptamer 64(DTA-64) on airway inflammation and epithelial to mesenchymal transition(EMT) induced by ovalbumin(OVA) in asthmatic mice. Methods Thirty-two female BALB/c mice(8 weeks old) were randomly divided into PBS group, OVA model group, DTA-64 group(1 μg/mouse), and control aptamer group, with 8 in each. HE staining of lung tissues was used to detect inflammatory cell infiltration around the airways;immunohistochemical staining was used to detect DEK expression around the airways;ELISA was used to detect serum IgE, and Th2-type cytokines IL-4, IL-5, IL-13 and Th1-type cytokine IFN-γ in bronchoalveolar lavage fluid(BALF);Western blot was applied to detect the EMT-related proteins α-SMA, Snail+Slug, vimentin, and E-cadherin, and TGF-β1/Smad, MAPK, PI3K, AKT, as well as mTOR in lung;and flow cytometry was used to observe the α-SMA expression in the lung single cell suspensions. Results DEK protein was highly expressed in the lung tissues of OVA group mice and decreased in the DTA-64 group mice;DTA-64 reduced the infiltration of eosinophils and neutrophils around the airways, down-regulated serum OVA-specific IgE and IL-4, IL-5, IL-13 in BALF, and up-regulated IFN-γ;DTA-64 also reduced the expressions of vimentin, α-SMA, Snail+Slug in the lung tissue, and up-regulated epithelial marker E-cadherin. DTA-64 inhibited the expressions of TGF-β1 and its downstream canonical pathways Smad2/3 and Smad4, as well as the phosphorylation of non-canonical TGF-β1 pathways ERK1/2, p38 MAPK, JNK and PI3K/AKT/mTOR. Conclusion DTA-64 may inhibit the airway inflammation and EMT induced by OVA in asthmatic mice via blocking TGF-β1/Smad, MAPK and PI3K signaling pathways, thereby alleviating airway remodeling in asthma.

关 键 词：DEK 上皮间质转化 气道重塑 支气管哮喘 卵清蛋白(OVA) 

分 类 号：R392.8[医药卫生—免疫学] R562.25[医药卫生—基础医学]