天津医药

天津医药 ›› 2019, Vol. 47 ›› Issue (11): 1145-1150.doi: 10.11958/20191532

• 实验研究 • 上一篇    下一篇

热打击通过活化NLRP3炎性小体增加肺毛细血管的通透性

庄小垒，李俊岭，李茜汝，丁洪光   

  1. 1东莞市第三人民医院急诊科（邮编 523000），2血液科；3广东省人民医院（广东省医学科学院）急危重症医学部
  • 收稿日期:2019-05-23 修回日期:2019-09-24 出版日期:2019-11-15 发布日期:2019-12-17
  • 通讯作者: 李茜汝 E-mail:125231823@qq.com

Heat stress increases pulmonary capillary permeability via activating NLRP3 inflammasome

ZHUANG Xiao-lei,LI Jun-ling,LI Qian-ru,DING Hong-guang   

  1. 1 Emergency Department, 2 Hematology Department, Dongguan Third People's Hospital, Dongguan 523000, China; 3 Department of Emergency & Critical Care Medicine, Guangdong Provincial People's Hospital and Guangdong Academy of Medical Sciences
  • Received:2019-05-23 Revised:2019-09-24 Published:2019-11-15 Online:2019-12-17

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摘要： 目的 验证热打击通过活化 NLRP3炎性小体增加肺毛细血管通透性的机制。方法 C57BL/6小鼠和肺微血管内皮细胞，用 42 ℃热打击。体内实验分组包括：对照组和热打击组，每组 12只小鼠；体外实验分组包括：对照组、热打击组、热打击+2,2,6,6-四甲基哌啶氧化物（TEMPO）组、热打击+半胱氨酸的天冬氨酸蛋白水解酶（caspase）抑制剂（Z-VAD-FMK）组、热打击+白细胞介素-1受体拮抗剂（IL-1Ra）组，每组 4例。检测肺组织和肺微血管内皮细胞活性氧（ROS）表达；Western blot 和（或）免疫荧光检测 caspase-1、白细胞介素-1β（IL-1β）、紧密连接蛋白 ZO-1、occludin、claudin-5的表达；用伊文氏蓝检测小鼠肺毛细血管的通透性。结果 体内实验结果显示，与对照组比较，热打击组小鼠肺组织伊文氏蓝浓度明显升高、ROS表达上调、NLRP3炎性小体活化、IL-1β表达上调和紧密连接蛋白的表达下调（P＜0.01）。体外实验结果显示，用 TEMPO 清除 ROS 后，NLRP3炎性小体活化被抑制（P＜0.01）；用 ZVAD-FMK抑制 caspase-1作用后，IL-1β表达显著下调（P＜0.01）；用 IL-1Ra阻断 IL-1β作用后，紧密连接蛋白表达显著上调（P＜0.01）。结论 热打击可通过活化 NLRP3炎性小体促进 IL-1β的表达，进而下调紧密连接蛋白的表达，导致肺微血管通透性增加。

关键词: 中暑, 炎性体, 白细胞介素 1β, 紧密连接蛋白质类, NLR家族, 热蛋白结构域包含蛋白 3, 毛细血管通透性, 热打击

Abstract: Objective To explore whether heat stress would increase pulmonary capillary permeability via activating the NLRP3 (NLR family, pyrin domain-containing 3) inflammasome. Methods C57BL / 6 mice and pulmonary microvascular endothelial cells (PMVECs) were underwent 42 ℃ heat stress. The mice were randomly divided into two groups: control group and heat stress group, 12 mice for each group. The PMVECs were randomly divided into five groups:control group, heat stress group, heat stress + TEMPO group, heat stress + Z-VAD-FMK group and heat stress + interleukin-1 receptor antagonist (IL-1Ra) group, 4 cases for each group. Caspase-1, IL-1β and tight junction proteins (ZO-1, occludin,claudin-5) expressions were assessed by Western blot assay or double immunofluorescence. The permeability of blood-brain barrier was assessed by Evans blue staining. Results The concentrations of Evans blue in lung tissue were significantly increased in heat stress group compared with those of control group (P＜0.01). The expression levels of ROS, caspase-1 and IL-1β were significantly increased, and the expression levels of ZO-1, occludin and claudin-5 were significantly decreased,in heat stress group compared with those of control group (P＜0.01). Scavenging of ROS decreased the expression levels of caspase-1 (P＜0.01). The pharmacological (Z-VAD-FMK) inhibition of NLRP3 inflammasome activation decreased the expression levels of IL-1β (P＜0.01). The pharmacological (IL-1Ra) blocking of IL-1β receptor increased the expression levels of tight junction protein (ZO-1: P＜0.01, occludin: P＜0.01, claudin-5: P＜0.01). Conclusion Heat stress may increase pulmonary capillary permeability via the activation of NLRP3 inflammasome and the reducing of tight junction protein expression.

Key words: heat stroke, inflammasomes, interleukin-1beta, tight junction proteins, NLR family, pyrin domaincontaining 3 protein, capillary permeability, heat stress