基于NLRP3/caspase-1/GSDMD通路探讨扶正通络颗粒对肺纤维化大鼠模型巨噬细胞焦亡的影响  

Effect of Fuzheng Tongluo Granules on pyroptosis in rat model with pulmonary fibrosis based on NLRP3/caspase-1/GSDMD pathway

作　　者：陈凤 段乃凡 张兴[1] 张炜[1] CHEN Feng;DUAN Nai-fan;ZHANG Xing;ZHANG Wei(Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China)

机构地区：[1]上海中医药大学附属曙光医院,上海201203

出　　处：《中国中药杂志》2024年第23期6399-6406,共8页China Journal of Chinese Materia Medica

基　　金：上海市临床重点专科项目(shslczdzk05101);上海市自然科学基金项目(22ZR1460100);上海市市级科技重大专项(ZD2021CY001);上海市科委上海中医临床重点实验室项目(20DZ227220);医学技术博士学位点培育建设项目(A1-N24-204-04)。

摘　　要：探讨扶正通络颗粒对特发性肺纤维化(idiopathic pulmonary fibrosis,IPF)的治疗作用及其机制。将72只SD大鼠随机分为对照组,模型组,吡非尼酮组(162 mg·kg^(-1))以及扶正通络颗粒低、中、高剂量组(2.63、5.25、10.5 g·kg^(-1)),采用单次无创气管插管滴注博来霉素(bleomycin,BLM)诱导IPF大鼠模型。造模第2天后每日灌胃,并记录体质量。第28天取材,称量肺质量,计算肺系数,HE、Masson染色观察肺组织病理变化,碱水解法检测肺组织羟脯氨酸(hydroxyproline,HYP)含量,ELISA法检测肺组织肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β(interleukin-1β,IL-1β)、白细胞介素-18(interleukin-18,IL-18)含量,免疫组化观察α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)、Ⅰ型胶原蛋白(collagenⅠ)表达,Western blot检测肺组织NOD样蛋白受体3(NOD-,LRR-and pyrin domain-containing 3,NLRP3)、半胱氨酸天冬氨酸蛋白酶1(cysteine-requiring aspartate protease type 1,caspase-1)、消皮素D-N(gasdermin D-N,GSDMD-N)、凋亡相关斑点样蛋白(apoptosis-associated speck-like protein containing a CARD,ASC)蛋白表达,免疫荧光共定位观察GSDMD和CD68的表达。结果显示,与对照组比较,模型组大鼠肺系数、Ashcroft评分、Szapiel评分增加,肺组织HYP、TNF-α、IL-1β、IL-18含量增加,NLRP3、caspase-1、GSDMD-N、ASC蛋白表达升高,GSDMD、CD68表达增加,GSDMD与CD68存在高度共定位。与模型组比较,各给药组大鼠肺系数、Ashcroft评分、Szapiel评分降低,肺组织HYP、TNF-α、IL-1β、IL-18含量减少,NLRP3、caspase-1、GSDMD、ASC蛋白表达下降,GSDMD、CD68表达减少,GSDMD与CD68存在高度共定位。综上所述,扶正通络颗粒可有效改善IPF大鼠肺组织纤维化和炎症,其机制可能与下调NLRP3/caspase-1/GSDMD通路抑制巨噬细胞焦亡有关。To investigate the therapeutic effect of Fuzheng Tongluo Granules on idiopathic pulmonary fibrosis(IPF)and its mechanism.Seventy-two SD rats were randomly divided into the control group,model group,pirfenidone group(162 mg·kg^(-1)),and low-,medium-and high-dose of Fuzheng Tongluo Granules groups(2.63,5.25,10.5 g·kg^(-1)).Rat model of IPF was induced by a single non-invasive tracheal intubation drip of bleomycin(BLM).The corresponding drugs were given daily by gavage after the 2nd day of modeling,and body mass was recorded.On the 28th day,the samples were collected and weighed,and the lung coefficients were calculated.The pathological changes in the lung tissue were observed by HE and Masson staining,and the hydroxyproline(HYP)content of the lung tissue was detected by alkaline hydrolysis.The contents of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),and interleukin-18(IL-18)of the lung tissue were determined by ELISA.The expression of collagen type I(collagenⅠ)andα-smooth muscle actin(α-SMA)was observed by immunohistochemistry.The expression levels of NOD-,LRR-and pyrin domain-containing 3(NLRP3),cysteine-requiring aspartate protease type 1(caspase-1),gasdermin D-N(GSDMD-N),and apoptosis-associated speck-like protein containing a CARD(ASC)in the lung tissue were detected by Western blot.Immunofluorescence co-localization was used to observe the expression of GSDMD and CD68.The results show that compared with the control group,the model group showed increased lung coefficient,Ashcroft score,Szapiel score,HYP,TNF-α,IL-1β,and IL-18 content in the lung tissue and elevated protein expression levels of NLRP3,caspase-1,GSDMD-N,and ASC.The expression levels of GSDMD and CD68 were increased,and there was a high degree of co-localization between GSDMD and CD68.Compared with those in the model group,the lung coefficient,Ashcroft score,and Szapiel score decreased in all drug administration groups,and the content of HYP,TNF-α,IL-1β,and IL-18 decreased.The protein expression levels of NLRP3,caspase-1,GSDMD-

关 键 词：特发性肺纤维化 扶正通络颗粒 细胞焦亡 NLRP3/caspase-1/GSDMD通路 巨噬细胞 

分 类 号：R285.5[医药卫生—中药学]