二陈汤加味通过抑制NLRP3通路对慢性阻塞性肺疾病的防治作用

尚立芝; 季书; 王国强; 陈晓辉; 王红伟; 石龙涛; 张光远; 李耀洋; 徐莉莉

doi:10.13422/j.cnki.syfjx.20192204

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二陈汤加味通过抑制NLRP3通路对慢性阻塞性肺疾病的防治作用

二陈汤加味对慢性阻塞性肺疾病作用的研究专题 | 更新时间：2021-02-09

- 二陈汤加味通过抑制NLRP3通路对慢性阻塞性肺疾病的防治作用
- Effect of Modified Erchentang on NLRP3 Inflammasome Expression in Peripheral Blood Mononuclear Cells (PBMCs) of Rats with Chronic Obstructive Pulmonary Disease
- 中国实验方剂学杂志 2019年25卷第23期页码：56-64
- 作者机构：
  
  河南中医药大学，郑州　 450046
- 作者简介：
  
  尚立芝，硕士，教授，从事中医药作用机制研究，Tel：0371-86253082，E-mail：lzshang2014@163.com
  王国强，博士，助理研究员，从事中医药对免疫性疾病作用机制研究，Tel：0371-86253082，E-mail：biowgq@126.com；
  王红伟，副教授，从事中医药治疗肺系疾病临床研究，E-mail: 156119506@qq.com
- 基金信息：
  
  国家自然科学基金面上项目(81573881);河南省高等学校重点科研项目(15A360030);国家级大学生创新创业训练计划项目(S201810471013;S201910471010);河南中医药大学生创新学习项目(CXXM[2017]0252;CXXM[2019]0001);河南省科重点科技攻关项目(152102310337)
- DOI：10.13422/j.cnki.syfjx.20192204
  中图分类号： R2-0; R22; R285.5; R289
- 收稿日期：2019-05-31，
  
  网络出版日期：2019-08-02，
  
  纸质出版日期：2019-12-05
- 稿件说明：
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尚立芝, 季书, 王国强, 等. 二陈汤加味通过抑制NLRP3通路对慢性阻塞性肺疾病的防治作用[J]. 中国实验方剂学杂志, 2019,25(23):56-64.

Li-zhi SHANG, Shu JI, Guo-qiang WANG, et al. Effect of Modified Erchentang on NLRP3 Inflammasome Expression in Peripheral Blood Mononuclear Cells (PBMCs) of Rats with Chronic Obstructive Pulmonary Disease[J]. Chinese journal of experimental traditional medical formulae, 2019, 25(23): 56-64.
尚立芝, 季书, 王国强, 等. 二陈汤加味通过抑制NLRP3通路对慢性阻塞性肺疾病的防治作用[J]. 中国实验方剂学杂志, 2019,25(23):56-64. DOI： 10.13422/j.cnki.syfjx.20192204.

Li-zhi SHANG, Shu JI, Guo-qiang WANG, et al. Effect of Modified Erchentang on NLRP3 Inflammasome Expression in Peripheral Blood Mononuclear Cells (PBMCs) of Rats with Chronic Obstructive Pulmonary Disease[J]. Chinese journal of experimental traditional medical formulae, 2019, 25(23): 56-64. DOI： 10.13422/j.cnki.syfjx.20192204.

摘要

目的：

观测二陈汤加味对慢性阻塞性肺疾病(COPD)大鼠外周血单个核细胞(PBMCs)中Nod样受体热蛋白结构域相关蛋白3(NLRP3)炎症小体相关基因表达的影响，探讨二陈汤加味对COPD抗炎的分子机制。

方法：

采用脂多糖(LPS)联合香烟烟雾方法制备COPD大鼠模型。40只大鼠随机平均分为正常组，模型组，二陈汤加味组，MCC950(NLRP3抑制剂)组。造模期间(从实验第1～30天)，MCC950组于实验第1天单次腹腔注射60 mg·kg

－1

；二陈汤加味组以10 g·kg

－1

剂量灌胃，1次/2 d。造模成功后，从实验第31～45天，MCC950组腹腔注射3 mg·kg

－1

，1次/2 d；二陈汤加味组以10 g·kg

－1

剂量灌胃，2次/d；正常组、模型组灌胃生理盐水10 g·kg

－1

。酶联免疫吸附测定法(ELISA)测定大鼠肺组织匀浆中白细胞介素-1

(IL-1

)，白细胞介素-18(IL-18)和趋化因子8(CXCL8)含量；实时荧光定量聚合酶链式反应(Real-time PCR)检测大鼠外周血PBMCs中NLRP3，凋亡相关斑点样蛋白(ASC)和半胱氨酸天冬氨酸蛋白水解酶-1(Caspase-1)mRNA表达；蛋白免疫印迹法(Western blot)检测PBMCs中NLRP3，ASC，Caspase-1蛋白表达；光镜观察肺组织结构变化，免疫组化检测NLRP3，ASC和Caspase-1在肺组织中的定位表达。

结果：

与正常组比较，模型组大鼠肺组织匀浆中IL-1

，IL-18和CXCL8含量均明显升高(

0.05，

0.01)；PBMCs中NLRP3，ASC，Caspase-1 mRNA及蛋白表达水平均明显增高(

0.01)；与模型组比较，二陈汤加味组PBMCs中NLRP3，ASC，Caspase-1 mRNA和蛋白表达水平均明显降低(

0.05，

0.01)，肺匀浆中IL-1

，IL-18和CXCL8含量均明显下降(

0.05，

0.01)，肺组织结构明显改善。

结论：

二陈汤加味对COPD有抗炎作用。其机制可能与抑制PBMCs中NLRP3，ASC，Caspase-1 mRNA表达，减少IL-1

，IL-18和CXCL8的释放有关。

Abstract

Objective:

To observe the effect of modified Erchentang on the expressions of NLRP3 inflammasome genes in peripheral blood mononuclear cells (PBMCs) and the levels of interleukin-1

(IL-1

)and interleukin-18(IL-18)and chemokine8 (CXCL8) in lung tissue of rats with chronic obstructive pulmonary disease (COPD)

in order to explore the molecular mechanism of modified Erchentang against inflammation of COPD.

Method:

Forty SD rats were randomly divided into normal control group

model group

MCC950 (NLRP3 inhibitor) group and modified Erchentang group. The COPD model of rats was prepared by using cigarette smoke and dripping with lipopolysaccharide (LPS). During the modeling period (from the 1

to the 30

day)

the MCC950 group received a single intraperitoneal injection with 60 mg·kg

-1

on the first day of the experiment

and the modified Erchentang group was given intragastric administration with 10 g·kg

-1

once every 2 days. From the 31

to the 45

day

the MCC950 group was intraperitoneally injected with 3 mg·kg

-1

once every 2 days

the modified Erchentang group was given intragastric administration with 10 g·kg

-1

twice a day

and the normal group and the model group received normal saline (NS) with 10 g·kg

-1

twice a day. The levels of interleukin-1

(IL-1

)

interleukin-18(IL-18) and chemokine8 (CXCL8) in rats lung tissue homogenate were detected by enzyme-linked immunosorbent assay (ELISA). The expressions of NLRP3

apoptosis-associated speck-like protein (ASC) and cysteinyl aspartate specific proteinase-1 (Caspase-1) mRNA in PBMCs were measured by Real-time fluorescence quantitative PCR (Real-time PCR). Western blot was used to detect the levels of NLRP3

ASC and Caspase-1 proteins in PBMCs. Immunohistochemical(IHC)method was used to detect the expressions of NLRP3

ASC and Caspase-1 proteins in lung tissues.

Result:

The expressions of NLRP3

ASC and Caspase-1 mRNA and protein were increased significantly (

0.01) in model group compared with normal group. Compared with model group

the expressions of NLRP3

ASC and Caspase-1 mRNA and protein were decreased significantly in PBMCs (

0.05

0.01) of modified Erchentang group. The levels of IL-18

IL-1

and CXCL8 in lung tissue homogenate in model group were significantly higher than those in the control group. However

compared with model group

the levels of IL-18

IL-1

and CXCL8 were decreased significantly (

0.05

0.01) in modified Erchentang group.

Conclusion:

NLRP3 inflammasome is involved in the inflammatory response in COPD rats. Modified Erchentang may inhibit the inflammatory response of COPD effectively. The mechanism may be correlated with the reduction of NLRP3

ASC and Caspase-1 gene expressions

and the inhibition of the release of IL-18

IL-1

and CXCL8.

关键词

Keywords

references

Global Strategy for the Diagnosis, Management and Prevention of COPD, Global Initiative for Chronic Obstructive Lung Disease (GOLD) . updated 2016 . http://goldcopd.org/.1 .

Martinon F , Burns K , Tschopp J . The inflammasome: a molecular platform triggering activation of inflammatory caspases and processing of pro IL-β [J]. Mol Cell , 2002 , 10 ( 2 ): 417 - 426 .

Howrylak J A , Nakahira K . Inflammasomes: key mediators of lung immunity [J]. Annu Rev Physiol , 2017 , 79 ( 1 ): 471 - 494 .

Shrivastava G , León-Juárez M , García-Cordero J , et al . Inflammasomes and its importance in viral infections [J]. Immunol Res , 2016 , 64 ( 5-6 ): 1101 - 1117 .

SHI J , GAO W , SHAO F . Pyroptosis: gasdermin-mediated programmed necrotic cell death [J]. Trends Biochem Sci , 2016 , 42 ( 4 ): 245 - 254 .

HE Y , ZENG M Y , YANG D , et al . NEK7 is an essential mediator of NLRP3 activation downstream of potassium efflux [J]. Nature , 2016 , 530 ( 7590 ): 354 - 357 .

Giavridis T , van der Stegen S J C , Eyquem J , et al . CAR T cell-induced cytokine release syndrome is mediated by macrophages and abated by IL-1 blockade [J]. Nat Med , 2018 , 24 : 731 - 738 .

Norelli M , Camisa B , Barbiera G , et al . Monocyte-derived IL-1 and IL-6 are differentially required for cytokine-release syndrome and neurotoxicity due to CART cells [J]. Nat Med , 2018 , 24 ( 6 ): 739 - 748 .

Misawa T , Takahama M , Kozaki T , et al . Microtubule-driven spatial arrangement of mitochondria promotes activation of the NLRP3 inflammasome [J]. Nat Immunol , 2013 , 14 ( 5 ): 454 - 460 .

Iyer S S , HE Q , Janczy J R , et al . Mitochondrial cardiolipin is required for NLRP3 inflammasome activation [J]. Immunity , 2013 , 39 ( 2 ): 311 - 323 .

刘力兴，贾钦尧．支气管哮喘患者外周血单个核细胞NLRP3 mRNA和血清IL-18、IL-1β的表达和研究 [J]．临床和实验医学杂志， 2016 ， 15 ( 4 )： 345 - 347 ．

张维康，潘灵辉． NOD样受体蛋白3炎症小体在呼吸机相关性肺损伤中的作用机制研究 [J]．中华危重病急救医学， 2015 ， 27 ( 10 )： 821 - 825 ．

谢文英，司春婴，尚立芝．爱罗咳喘宁对老年COPD急性加重期痰湿阻肺证患者血清中细胞因子及肺功能的影响 [J]．中国实验方剂学杂志， 2015 ， 21 ( 12 )： 143 - 146 ．

尚立芝，谢文英，张良芝，等．爱罗咳喘宁对稳定期和急性加重COPD抗炎作用及机制研究 [J]．中国实验方剂学杂志， 2015 ， 21 ( 1 )： 134 - 139 ．

尚立芝，谢文英，张良芝，等．爱罗咳喘宁对COPD大鼠肺组织炎症因子及氧化应激的影响 [J]．中国实验方剂学杂志， 2014 ， 20 ( 24 )： 168 - 171 ．

尚立芝，季书，谢文英，等．二陈汤加味对COPD急性期患者CCl6，SP-D及HAT/HDAC的影响 [J]．中国实验方剂学杂志， 2017 ， 23 ( 10 )： 163 - 170 ．

陈四清，谢文英，尚立芝，等．二陈汤加味对慢性阻塞性肺疾病急性加重期老年患者免疫功能及CCL18、CC16、IL-8、sICAM-1的影响 [J]．中国实验方剂学杂志， 2017 ， 23 ( 10 )： 171 - 177 ．

谢文英，季书，尚立芝．二陈汤加味对COPD患者氧化应激、缺氧诱导因子1α及沉默信息调节因子1(Sirt1)的影响 [J]．中国实验方剂学杂志， 2017 ， 23 ( 10 )： 155 - 162 ．

Yildirim E , Kormi I , Baolu K , et al . Periodontal health and serum, saliva matrix metalloproteinases in patients with mild chronic obstructive pulmonary disease [J]. J Periodontal Res , 2013 , 48 ( 3 ): 269 - 275 .

Mahadeva R , Shapiro S D . Chronic obstructive pulmonary disease: experimental animalmodels of ulmonary emphysema [J]. Thorax , 2002 , 57 ( 10 ): 908 - 914 .

齐先梅，王蕾，张瑞恒，等． MCC950在野百合碱诱导大鼠肺动脉高压模型中的治疗作用及其机制研究 [J]．首都医科大学学报， 2018 ， 39 ( 6 )： 871 - 875 ．

Coll R C , Robertson A A , Chae J J , et al . A small-molecule inhibitor of the NLRP3 inflammasome for the treatment of inflammatory diseases [J]. Nat Med , 2015 , 21 ( 3 ): 248 - 255 .

吴珂，尚立芝，谢文英，等．二陈汤加味对慢性阻塞性肺疾病细支气管壁细胞外基质重塑的影响 [J]．中国实验方剂学杂志， 2018 ， 24 ( 14 )： 122 - 127 ．

尚立芝，季书，刘坦，等．二陈汤加味对慢性阻塞性肺疾病大鼠 Smad3，4，6，7基因表达的影响 [J]．中国实验方剂学杂志， 2017 ， 23 ( 10 )： 139 - 146 ．

何子凡，高岩，王晓玲，等． NLRP3炎性小体在慢性阻塞性肺疾病患者机体炎症反应中作用的研究 [J]．中国呼吸与危重监护杂志， 2018 ， 17 ( 2 )： 119 - 123 ．

ZHU J , WU S , HU S , et al . NLRP3 inflammasome expression in peripheral blood monocytes of coronary heart disease patients and its modulation by rosuvastatin [J]. Mol Med Rep , 2019 , 20 ( 2 ): 1826 - 1836 .

Riteau N , Gasse P , FAUConnier L , et al . Extracellular ATP is a danger signal activating P2X7 receptor in lung inflammation and fibrosis [J]. Am J Respir Crit Care Med , 2010 , 182 ( 6 ): 774 - 783 .

Kanneganti T D , Body-Malapel M , Amer A , et al . Critical role for Cryopyrin/Nalp3 in activation of Caspase-1 in response to viral infection and double-stranded RNA [J]. J Biol Chem , 2006 , 281 ( 48 ): 36560 - 36568 .

Kanneganti T D , Ozören N , Body-Malapel M , et al . Bacterial RNA and small antiviral compounds activate Caspase-1 through cryopyrin/Nalp3 [J]. Nature , 2006 , 440 ( 7081 ): 233 - 236 .

Martinon F , Agostini L , Meylan E , et al . Identification of bacterial muramyl dipeptide as activator of the NALP3/cryopyrin inflammasome [J]. Curr Biol , 2004 , 14 ( 21 ): 1929 - 1934 .

GUO H , Callaway J B , TING J P . Inflammasomes: mechanism of action, role in disease, and therapeutics [J]. Nat Med , 2015 , 21 ( 7 ): 677 - 687 .

Schroder K , Tschopp J . The inflammasomes [J]. Cell , 2010 , 140 ( 6 ): 821 - 883 .

De Nardo D , Latz E . NLRP3 inflammasomes link inflammation and metabolic disease [J]. Trends Immunol , 2011 , 32 ( 8 ): 373 - 379 .

Gross O , Thomas C J , Guarda G , et al . The inflammasome: an integrated view [J]. Immunol Rev , 2011 , 243 ( 1 ): 136 - 151 .

Franklin B S , Bossaller L , De Nardo D , et al . The adaptor ASC has extracellular and prionoid activities that propagate inflammation [J]. Nat Immunol , 2014 , 15 ( 8 ): 727 - 737 .

Uh S T , Koo S M , Kim Y , et al . The activation of NLRP3-inflammsome by stimulation of diesel exhaust particles in lung tissues from emphysema model and RAW 264.7 cell line [J]. Korean J Intern Med , 2017 , 32 ( 5 ): 865 - 874 .

LI C , HUANG Z H , LI W L , et al . The nucleotide-binding oligomerization domain-Like receptor family pyrin domain-containing inflammasome regulates bronchial epithelial cell injury and proapoptosis after exposure to biomass fuel smoke [J]. Am J Respir Cell Mol Biol , 2016 , 55 ( 6 ): 815 - 824 .

Pinkerton J W , Kim R Y , Robertson A A B , et al . Inflammasomes in the lung [J]. Mol Immunol , 2017 , 86 : 44 - 55 .

Goldberg E L , Asher J L , Molony R D , et al . β-Hydroxybu-tyrate deactivates neutrophil NLRP3 inflammasome to relieve gout flares [J]. Cell Rep , 2017 , 18 ( 9 ): 2077 - 2087 .

TIAN R , ZHU Y , YAO J Y , et al . NLRP3 participates in the regulation of EMT in bleomycin-induced pulmonary fibrosis [J]. Exp Cell Res , 2017 , 357 ( 2 ): 328 - 334 .

Cocco M , Pellegrini C , Martinez-Banaclocha H , et al . Development of an acrylate derivative targeting the NLRP3 inflammasome for the treatment of inflammatory bowel disease [J]. J Med Chem , 2017 , 60 ( 9 ): 3656 - 3671 .

Tsai Y M , Chiang K H , Hung J Y , et al . Der f1 induces pyroptosis in human bronchial epithelia via the NLRP3 inflammasome [J]. Int J Mol Med , 2018 , 41 ( 2 ): 757 - 764 .

彭菲菲，邵强，赵宁，等． MCC950对线粒体损伤相关分子模式诱导大鼠肺损伤的保护作用 [J]．医学研究生学报， 2017 ， 30 ( 11 )： 1166 - 1171 ．

Dima E , Koltsida O , Katsaounou P , et al . Implication of Interleukin (IL)-18 in the pathogenesis of chronic obstructive pulmonary disease (COPD) [J]. Cytokine , 2015 , 74 ( 2 ): 313 - 317 .

Jordan J A , GUO R F , YUN E C , et al . Role of IL-18 in acute lung inflammation [J]. J Immunol , 2001 , 167 ( 12 ): 7060 - 7068 .

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