细粒棘球蚴囊液致敏过程中IL-1β受体阻滞剂对肺损伤的治疗作用

doi:10.12140/j.issn.1000-7423.2024.02.010

中国寄生虫学与寄生虫病杂志 ›› 2024, Vol. 42 ›› Issue (2): 199-203.doi: 10.12140/j.issn.1000-7423.2024.02.010

细粒棘球蚴囊液致敏过程中IL-1β受体阻滞剂对肺损伤的治疗作用

王春生¹(), 乌尔格力¹, 西利扎提·库来西¹, 李玉倩¹, 先依旦·阿布拉¹, 苏比·泰来提¹, 蒲雪莉¹, 王佳玲¹, 李孟², 房志远³, 叶建荣¹^,^*()

1 新疆医科大学第一附属医院麻醉科，乌鲁木齐 830000
2 昌吉市人民医院麻醉手术部，新疆昌吉 831100
3 广东医科大学附属东莞第一医院麻醉科，东莞 523710

收稿日期:2023-07-17 修回日期:2023-09-11 出版日期:2024-04-30 发布日期:2024-04-30
通讯作者: * 叶建荣（1978—），女，博士，主任医师，从事围术期器官保护研究。E-mail：616227972@qq.com
作者简介:王春生（1989—）男，硕士，住院医师，从事围术期器官保护研究。E-mail：18024058@qq.com
基金资助:
国家自然科学基金(82060581);新疆围术期器官保护实验室(XJDX1411);自治区重点实验室开放课题(2020D04025)

Therapeutic effects of IL-1β receptor blocker on lung injury during the sensitization process of Echinococcus granulosus cyst fluid

WANG Chunsheng¹(), WU Ergeli¹, XILIZATI Kulaixi¹, LI Yuqian¹, XIANYIDAN Abula¹, SUBI Tailaiti¹, PU Xueli¹, WANG Jialing¹, LI Meng², FANG Zhiyuan³, YE Jianrong¹^,^*()

1 Department of Anesthesiology, the First Affiliated Hospital of Xinjiang Medical University, Urumqi 830000, China
2 Department of Anesthesia and Surgery, Changji People’s Hospital, Changji 831100, Xinjiang, China
3 Department of Anesthesiology, Dongguan First Hospital Affiliated to Guangdong Medical University, Dongguan 523710, China

Received:2023-07-17 Revised:2023-09-11 Online:2024-04-30 Published:2024-04-30
Contact: * E-mail: 616227972@qq.com
Supported by:
National Natural Science Foundation of China(82060581);Xinjiang Perioperative Organ Protection Laboratory(XJDX1411);Autonomous Region Key Laboratory Open Project(2020D04025)

摘要/Abstract

摘要：

目的探究细粒棘球蚴囊液外溢致敏过程中白细胞介素-1β（IL-1β）受体阻滞剂对肺损伤的治疗作用机制。方法细粒棘球蚴包囊采集自新疆乌鲁木齐华凌屠宰场感染细粒棘球蚴的新鲜羊肝，收集包囊囊液，消化沉淀获得原头节，上清经内毒素处理获得致敏囊液。将24只小鼠随机分为对照组、致敏组、阻滞剂治疗组和阻滞剂预防组，每组6只。致敏组、阻滞剂治疗组和阻滞剂预防组小鼠分别腹腔注射约2 000个细粒棘球蚴原头节，对照组小鼠腹腔注射等体积生理盐水（约1 ml）。3个月后致敏组小鼠腹腔注射致敏囊液（0.1 ml/g体质量），阻滞剂治疗组小鼠先腹腔注射致敏囊液、15 min后尾静脉注射IL-1β受体阻滞剂（4 μg/g体质量），阻滞剂预防组小鼠先尾静脉注射阻滞剂、15 min后腹腔注射致敏囊液，对照组小鼠腹腔注射等体积生理盐水。取小鼠肺组织，制备石蜡切片后苏木精-伊红（HE）染色，显微镜下观察肺组织的炎症损伤情况。提取小鼠肺组织总RNA，qRT-PCR检测肿瘤坏死因子α（TNF-α）、IL-6、半胱氨酸天冬氨酸蛋白酶-8（caspase-8）、磷脂酰肌醇3激酶（PI3K）、蛋白激酶B（Akt）、核因子-κB（NF-κB）的mRNA相对转录水平。提取小鼠肺组织蛋白，以β肌动蛋白（β-actin）为内参，蛋白质免疫印迹（Western blotting）检测PI3K、Akt、NF-κB的蛋白相对表达水平。应用GraphPad Prism软件对数据进行统计学分析，组间比较采用单因子方差分析。结果 HE染色结果显示，致敏组小鼠肺组织局部充血，充血部位周围炎症细胞游出、淋巴细胞聚集；阻滞剂治疗组、阻滞剂预防组和对照组肺组织均未见明显炎症反应。qRT-PCR结果显示，对照组小鼠肺组织中IL-6、TNF-α、caspase-8、PI3K、Akt和NF-κB的mRNA相对转录水平分别为1.057 ± 0.363、1.020 ± 0.217、1.004 ± 0.097、1.000 ± 0.031、1.035 ± 0.312、1.029 ± 0.304，致敏组分别为2.013 ± 0.514、2.189 ± 0.194、6.433 ± 0.340、1.594 ± 0.117、2.902 ± 0.181、1.342 ± 0.146，阻滞剂治疗组分别为1.243 ± 0.279、1.268 ± 0.225、0.869 ± 0.172、1.103 ± 0.180、1.371 ± 0.199、1.008 ± 0.202，阻滞剂预防组分别为1.223 ± 0.358、0.970 ± 0.303、0.932 ± 0.298、0.825 ± 0.404、1.421 ± 0.137、1.083 ± 0.222；致敏组均高于对照组（t = 3.481、2.759、37.640、2.237、12.670、2.274，均P < 0.05），阻滞剂治疗组和阻滞剂预防组均低于致敏组（t = 3.221、7.593、35.240、5.610、13.920、3.287，3.088、8.299、28.610、4.475、15.930、2.390，均P < 0.05）。Western blotting结果显示，对照组小鼠肺组织中PI3K、Akt和NF-κB的蛋白相对表达水平分别为0.516 ± 0.075、0.638 ± 0.103、0.198 ± 0.086，致敏组分别为0.831 ± 0.061、0.917 ± 0.069、0.784 ± 0.120，阻滞剂治疗组分别为0.535 ± 0.108、0.612 ± 0.206、0.247 ± 0.145，阻滞剂预防组分别为0.526 ± 0.117、0.565 ± 0.087、0.154 ± 0.031；致敏组均高于对照组（t = 5.650、3.901、6.871，均P < 0.05），阻滞剂治疗组和阻滞剂预防组均低于致敏组（t = 4.142、2.434、4.945，4.013、5.477、8.821，均P < 0.05）。结论 IL-1β受体阻滞剂在细粒棘球蚴囊液引起的过敏反应中能够通过抑制PI3K/Akt/NF-κB通路，降低肺部炎症反应，对肺损伤具有治疗作用。

关键词: 细粒棘球绦虫, 囊液致敏, IL-1β受体阻滞剂, 肺损伤治疗

Abstract:

Objective To investigate the therapeutic mechanism of interleukin-1β (IL-1β) receptor blocker on lung injury in the sensitization process of Echinococcus granulosus cyst extravasated fluid. Methods The cysts of E. granulosus were collected from fresh sheep livers infected with E. granulosus from Hualing Slaughterhouse of Urumqi, Xinjiang. The cyst fluid was collected, digested and sedimented to obtain protoscoleces, and the supernatant was treated with endotoxin to obtain allergenic cyst fluid. Twenty-four mice were randomly divided into control group, sensitized group, blocker treatment group and blocker prevention group, with 6 mice in each group. Mice in the sensitized group, the blocker treatment group and the blocker prevention group were intraperitoneally injected with about 2 000 protoscoleces, respectively, while mice in the control group were injected with about the same volume (1 ml) of normal saline. Three months later, mice in the sensitized group were intraperitoneally injected with allergenic cyst fluid (0.1 ml per g weight); mice in the blocker treatment group were injected with allergenic cyst fluid prior to intravenous injection with IL-1β blocker (4 μg per g weight) 15 minutes later; mice in the blocker prevention group were injected with IL-1β blocker prior to intraperitoneal injection with allergenic cyst fluid 15 minutes later; mice in the control group were injected with the same volume of normal saline. The paraffin sections of mice lung tissues were stained with hematoxylin-eosin (HE) to observe the inflammatory damage under microscope. Total RNA was extracted from lung tissue and the mRNA relative transcription levels of tumor necrosis factor α (TNF-α), IL-6, cysteinyl aspartate specific proteinase-8, phosphatidylinositol 3 kinase (PI3K), protein kinase B (Akt) and nuclear factor kappa-B (NF-κB) were detected by qRT-PCR. Western blotting was used to detect the protein relative expression levels of PI3K, Akt and NF-κB, with β-actin as an internal reference. Data was analyzed by GraphPad Prism software. One-way analysis of variance was used for comparison between groups. Results HE staining showed that the lung tissues of the sensitized group were congested, with inflammatory cells swimming out and lymphocytes aggregating around the congested area. There was no obvious inflammatory reaction in the lung tissues of the blocker treatment group, blocker prevention group and control group. qRT-PCR results showed that the mRNA relative transcription levels of IL-6, TNF-α, caspase-8, PI3K, Akt and NF-κB were 1.057 ± 0.363, 1.020 ± 0.217, 1.004 ± 0.097, 1.000 ± 0.031, 1.035 ± 0.312 and 1.029 ± 0.304 in the control group, 2.013 ± 0.514, 2.189 ± 0.194, 6.433 ± 0.340, 1.594 ± 0.117, 2.902 ± 0.181 and 1.342 ± 0.146 in sensitized group, 1.243 ± 0.279, 1.268 ± 0.225, 0.869 ± 0.172, 1.103 ± 0.180, 1.371 ± 0.199 and 1.008 ± 0.202 in the blocker treatment group and 1.223 ± 0.358, 0.970 ± 0.303, 0.932 ± 0.298, 0.825 ± 0.404, 1.421 ± 0.137 and 1.083 ± 0.222 in the blocker prevention group, respectively. The mRNA relative transcription levels in the sensitized group were higher than the control group (t = 3.481, 2.759, 37.640, 2.237, 12.670, 2.274, all P < 0.05). The mRNA relative transcription levels in the blocker treatment group and blocker prevention group were lower than those in the sensitized group (t = 3.221, 7.593, 35.240, 5.610, 13.920, 3.287; 3.088, 8.299, 28.610, 4.475, 15.930, 2.390, all P < 0.05). Western blotting results showed that the protein relative expression levels of PI3K, Akt and NF-κB were 0.516 ± 0.075, 0.638 ± 0.103 and 0.198 ± 0.086 in the control group, 0.831 ± 0.061, 0.917 ± 0.069 and 0.784 ± 0.120 in sensitized group, 0.535 ± 0.108, 0.612 ± 0.206 and 0.247 ± 0.145 in the blocker treatment group and 0.526 ± 0.117, 0.565 ± 0.087 and 0.154 ± 0.031 in the blocker prevention group, respectively. The protein relative expression levels in the sensitized group were higher than the control group (t = 5.650, 3.901, 6.871, all P < 0.05). The protein relative expression levels in the blocker treatment group and blocker prevention group were lower than those in the sensitized group (t = 4.142, 2.434, 4.945; 4.013, 5.477, 8.821, all P < 0.05). Conclusion IL-1β receptor blocker may reduce the pulmonary inflammatory response through inhibiting PI3K/Akt/NF-κB pathway in the allergic reaction induced by E. granulosus cyst fluid, implying its therapeutic effects on lung injury.

Key words: Echinococcus granulosus, Cyst fluid sensitization, IL-1β receptor blocker, Treatment of lung injury

中图分类号:

R532.32

王春生, 乌尔格力, 西利扎提·库来西, 李玉倩, 先依旦·阿布拉, 苏比·泰来提, 蒲雪莉, 王佳玲, 李孟, 房志远, 叶建荣. 细粒棘球蚴囊液致敏过程中IL-1β受体阻滞剂对肺损伤的治疗作用[J]. 中国寄生虫学与寄生虫病杂志, 2024, 42(2): 199-203.

WANG Chunsheng, WU Ergeli, XILIZATI Kulaixi, LI Yuqian, XIANYIDAN Abula, SUBI Tailaiti, PU Xueli, WANG Jialing, LI Meng, FANG Zhiyuan, YE Jianrong. Therapeutic effects of IL-1β receptor blocker on lung injury during the sensitization process of Echinococcus granulosus cyst fluid[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2024, 42(2): 199-203.

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基因 Gene	引物序列（5′→3′） Primer sequence (5′→3′)
TNF-α	F：CGACGTGGAACTGGCAGAAGAG
	R：GCAGGAATGAGAAGAGGCTGAGAC
IL-6	F：CTTCTTGGGACTGATGCTGGTGAC
	R：TCTGTTGGGAGTGGTATCCTCTGTG
caspase-8	F：CATCCTGACTGGCGTGAACTATGAC
	R：GGGCTGTGGCATCTGCTTTCC
NF-κB	F：GACACGACAGAATCCTCAGCATCC
	R：CCACCAGCAGCAGCAGACATG
PI3K	F：GGGAAGCGAGACGGCACTTTC
	R：TCTACCACTACGGAGCAGGCATAG
Akt	F：TCAGGATGTGGATCAGCGAGAGTC
	R：AGGCAGCGGATGATAAAGGTGTTG
GAPDH	F：GGTGAAGGTCGGTGTGAACG
	R：CTCGCTCCTGGAAGATGGTG

细粒棘球蚴囊液致敏过程中IL-1β受体阻滞剂对肺损伤的治疗作用

Therapeutic effects of IL-1β receptor blocker on lung injury during the sensitization process of Echinococcus granulosus cyst fluid

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