文章摘要
郑继生,钱景莉,姜娜,等.芪冬活血饮对急性肺损伤小鼠肺泡巨噬细胞iRhom2/TACE信号通路的影响[J].浙江中医药大学学报,2021,(5):530-536.
芪冬活血饮对急性肺损伤小鼠肺泡巨噬细胞iRhom2/TACE信号通路的影响
Effect of Qidong Huoxue Decoction on iRhom2/TACE Signaling Pathway in Alveolar Macrophages of Acute Lung Injury Mice
投稿时间:2020-05-02  
DOI:10.16466/j.issn1005-5509.2021.05.018
中文关键词: 芪冬活血饮  肠缺血/再灌注  急性肺损伤  肺泡巨噬细胞  iRhom2/TACE通路  TNF-α  IL-6  凋亡
英文关键词: Qidong Huoxue decoction  intestinal ischemia/reperfusion  acute lung injury  alveolar macrophages  iRhom2/TACE pathway  TNF-α  IL-6  apoptosis
基金项目:浙江省中医药科技计划项目(2019ZA027);国家自然科学基金项目(81774220、81603545)
作者单位E-mail
郑继生 浙江省立同德医院 杭州 310012  
钱景莉 浙江中医药大学附属第二医院  
姜娜 浙江省立同德医院 杭州 310012  
魏毅 浙江省立同德医院 杭州 310012  
何海栋 浙江省立同德医院 杭州 310012  
马春芳 浙江省人民医院  
蔡宛如 浙江中医药大学附属第二医院  
柴秀娟 浙江省立同德医院 杭州 310012 chaixiujuan0214@163.com 
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中文摘要:
      [目的]探索芪冬活血饮对肠缺血/再灌注诱导的急性肺损伤小鼠肺泡巨噬细胞未活化的菱形蛋白2(inactive rhomboid protein 2,iRhom2)/肿瘤坏死因子-α转化酶(tumor necrosis factor-α convertase,TACE)信号通路的影响。[方法]通过肠缺血/再灌注方法构建急性肺损伤小鼠模型。24只无特定病原体(specific pathogen free,SPF)级雌性C57BL/6小鼠被随机分为6组,每组4只,即正常组,假手术组,模型组,芪冬活血饮低(2mL·kg-1)、中(4mL·kg-1)、高(8mL·kg-1)剂量组。芪冬活血饮低、中、高剂量组小鼠在造模前24、12h以及造模后2、14h分别用2、4、8mL·kg-1芪冬活血饮灌胃,2次/d,共4d;模型组予等量的0.9%氯化钠溶液灌胃;正常组和假手术组不给药。末次给药24h后,分离并培养肺泡巨噬细胞。以酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)检测巨噬细胞分泌肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)和白介素-6(interleukin-6,IL-6)的水平。以末端标记法(terminal-deoxynucleoitidyl transferase mediated nick end labeling,TUNEL)染色分析肺上皮细胞凋亡情况。实时荧光定量聚合酶链反应(Real-time-quantitative polymerase chain reaction,RT-qPCR)和Western blot检测肺泡巨噬细胞中iRhom2 和TACE的mRNA和蛋白表达水平。[结果]与假手术组比较,模型组小鼠肺泡巨噬细胞TNF-α和IL-6分泌水平显著增加(P<0.0001);经芪冬活血饮治疗后,TNF-α和IL-6水平以浓度依赖的方式显著降低(P<0.01,P<0.001,P<0.0001)。TUNEL结果显示,芪冬活血饮显著抑制急性肺损伤模型小鼠的肺上皮细胞凋亡(P<0.05,P<0.01,P<0.001)。与假手术组比较,模型组小鼠巨噬细胞中iRhom2和TACE mRNA和蛋白表达被显著激活(P<0.001,P<0.0001,P<0.01);经芪冬活血饮治疗后,巨噬细胞中iRhom2和TACE表达以浓度依赖的方式显著降低(P<0.05,P<0.001,P<0.0001)。[结论]芪冬活血饮能够有效改善肠缺血/再灌注诱导的急性肺损伤,其机制可能与抑制小鼠肺泡巨噬细胞中iRhom2/TACE信号通路有关。
英文摘要:
      [Objective]To explore the effect of Qidong Huoxue decoction on inactive rhomboid protein 2/tumor necrosis factor-α convertase(iRhom2/TACE) signaling pathway of alveolar macrophages in mice with acute lung injury induced by intestinal ischemia/reperfusion.[Methods] In this study, a mouse model of acute lung injury induced by intestinal ischemia/reperfusion was constructed by clamping the superior mesenteric artery for 1 hour and reperfusion for 3 hours. Twenty-four female C57BL/6 mice were randomly divided into six groups, including normal group, sham operation group, model group, Qidong Huoxue decoction low-dose group(2mL·kg-1), Qidong Huoxue decoction medium-dose group(4mL·kg-1) and Qidong Huoxue decoction high-dose group(8mL·kg-1), with 4 mice in each group. Mice in Qidong Huoxue decoction low, medium, and high-dose groups were given 2, 4 and 8mL·kg-1 Qidong Huoxue decoction 24, 12h before modeling and 2, 14h after modeling, respectively.After that, the drug was administered twice a day for a total of 4 days. Model group was given the same amount of 0.9% sodium chloride aqueous solution. No treatment was given in normal group and sham operation group. Twenty-four hours after the last administration, the alveolar macrophages were isolated and cultured. The level of tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) secreted by macrophages was detected by enzyme-linked immunosorbent assay(ELISA). Terminal-deoxynucleoitidyl transferase mediated nick end labeling(TUNEL) test was used to analyze the apoptosis of lung epithelial cells. The mRNA and protein levels of iRhom2 and TACE in alveolar macrophages were detected by Real-time-quantitative polymerase chain reaction(RT-qPCR) and Western blot, respectively.[Results] Compared with sham operation group, the levels of TNF-α and IL-6 secreted by the alveolar macrophages in model group were significantly increased(P<0.0001). Qidong Huoxue decoction distinctly inhibited the levels of TNF-α and IL-6 in a concentration-dependent manner(P<0.01, P<0.001, P<0.0001). TUNEL results showed that Qidong Huoxue decoction significantly inhibited the apoptosis of lung epithelial cells in mice with acute lung injury in a concentration-dependent manner(P<0.05,P<0.01, P<0.001). Compared with sham operation group, both mRNA and protein expression of iRhom2 and TACE was significantly activated in the macrophages of model group(P<0.001, P<0.0001, P<0.01). However, after treatment with Qidong Huoxue decoction,the expression of iRhom2 and TACE in macrophages was significantly suppressed in a concentration-dependent manner(P<0.05, P<0.001, P<0.0001).[Conclusion] Qidong Huoxue decoction can improve acute lung injury induced by intestinal ischemia/reperfusion, the mechanism may be related to the Rhibition of iRhom2/TACE signaling pathway in mouse alveolar macrophages.
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