聚肌胞苷酸诱导急性肺损伤湿热证小鼠模型的建立及评价  

Establishment and Evaluation of Mouse Model Induced by Poly(I:C)Acute Lung Injury with Damp-Heat Syndrome

作　　者：杨上松 张庭瑞 周泽 温伟波 王志红 李钦 YANG Shang-song;ZHANG Ting-rui;ZHOU Ze;WEN Wei-bo;WANG Zhi-hong;LI Qin(School of Basic Medical Sciences,Yunnan University of Chinese Medicine,Kunming 650500;Yunnan Key Laboratory of Integrated,Traditional Chinese and Western Medicine for Chronic Disease in Prevention and Treatment,Kunming 650500)

机构地区：[1]云南中医药大学基础医学院,昆明650500 [2]云南省中西医结合慢病防治重点实验室,昆明650500

出　　处：《中国中西医结合杂志》2025年第2期220-228,共9页Chinese Journal of Integrated Traditional and Western Medicine

基　　金：国家自然科学基金地区基金项目(No.82260930,No.82060864);国家中医药管理局新型冠状病毒感染中医药应急专项课题(No.2023ZYLCYJ02-24);国家中医药多学科交叉创新团队项目(No.ZYYCXTD-D-202201);云南省科学技术厅中医联合专项项目(No.202101AZ070001-020);云南省科学技术厅重点研发计划项目(No.202103AC100005);云南省教育厅科学研究项目(No.2023Y0440);国家级大学生创新训练计划项目(No.202310680002)。

摘　　要：目的 建立并评价病证结合急性肺损伤(ALI)湿热证小鼠模型。方法 将50只C57BL/6J雄性小鼠随机分为正常组、湿热模型组、聚肌胞苷酸[Poly(I:C)]组、Poly(I:C)^(+)湿热组。以Poly(I:C)为生物致病因子,结合“外湿热环境^(+)高脂饮食14天”的复合因素构建动物模型。动物模型评价;在气管滴注Poly (I:C) 72 h后,取材测量小鼠肺指数,自动细胞计数仪检测支气管肺泡灌洗液(BALF)沉淀物中总细胞数、BCA蛋白测定法检测BALF上清液总蛋白浓度。直接测定法检测小鼠血清低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)、总胆固醇(TC)含量。ELISA法测定血清中胃动素(MTL)、P物质(SP)、胃泌素(GAS)含量。微量法测定肝组织Na^(+)-K^(+)-ATP酶活性。免疫组化法测定肺组织中水通道蛋白1(AQP1)、水通道蛋白5(AQP5)蛋白表达。HE染色法观察小鼠肺组织、舌组织病理改变。结果 与正常组比较,湿热模型组体重、HDL-C、Na^(+)-K^(+)-ATP酶活力降低(P<0.05,P<0.01),TC、LDL-C、MTL、SP、总细胞数水平升高(P<0.01)。与湿热模型组比较,Poly (I:C)组TC、LDL-C、MTL、SP降低(P<0.01),体重、Na^(+)-K^(+)-ATP酶活力、HDL-C升高(P<0.05,P<0.01);Poly(I:C)组、Poly(I:C)^(+)湿热组总细胞数、总蛋白浓度、肺指数、白细胞介素(IL)-17、IL-23水平、肺损伤病理评分升高(P<0.05,P<0.01),AQP1,AQP5表达降低(P<0.01)。与Poly(I:C)组比较,Poly(I:C)^(+)湿热组肛温、TC、LDL-C、MTL、SP、总细胞数、IL-17水平升高(P<0.01);体重、HDL-C、Na^(+)-K^(+)-ATP酶活力水平降低(P<0.05,P<0.01),舌组织黏膜层增厚(P<0.01)。结论 Poly(I:C)叠加外湿热环境和高脂饮食可成功构建ALI湿热证病证结合模型。Objective To establish and evaluate a mouse model of acute lung injury(ALI)with dampheat syndrome. Methods A total of 50 C57BL/6J male mice were randomly divided into normal group, dampheat model group, polyinosinic-polycytidylic acid [Poly(I:C)] group, and Poly(I:C)^(+) damp-heat group. The mouse model was constructed using Poly(I:C) as a biological pathogenic factor, along with the compound factors of external damp-heat environment and high-fat diet for 14 days. Evaluate animal models. After 72 h of intratracheal Poly(I:C) instillation, the total number of cells in the bronch oalveolar lavage fluid(BALF) sediment and total protein concentration in BALF supernatant were calculated using an automatic cell counter and a BCA protein assay, respectively. In addition, the lung index was calculated. The contents of low-density lipoprotein cholesterol(LDL-C), high-density lipoprotein cholesterol(HDL-C), and total cholesterol(TC)and those of motilin(MTL), substance P(SP), and gastrin(GAS)in mouse serum were determined via direct measurements and enzyme-linked immunosorbent assay, respectively. Na^(+)-K^(+)-ATPase activity in mouse liver was determined using the microtiter method, and the expression levels of aquaporin 1(AQP1)and aquaporin 5(AQP5)in the lung tissue were determined via immunohistochemistry. Finally, pathological changes in lung and tongue tissues were characterized using HE staining. Results Compared with the normal group, the contents of weight, HDL-C, Na^(+)-K^(+)-ATPase activity decreased(P<0.05,P<0.01), the levels of TC, LDL-C, MTL, SP, total cell number increased(P<0.05,P<0.01)in the damp-heat model group. Compared with the damp-heat model group, the Poly(I:C) group showed the levels of TC,LDL-C,MTL,SP decreased(P<0.01),the contents of weigh,Na^(+)-K^(+)-ATPase, HDL-C increased(P<0.05,P<0.01). In addition, the Poly(I:C) and Poly(I:C)^(+)damp-heat groups showed the total cell number, total protein concentration, lung index, IL-17, IL-23 levels and the pathological score of lung injury increased(P<0.05

关 键 词：急性肺损伤 湿热证 急性肺损伤湿热证 模型评价 聚肌胞苷酸 

分 类 号：R28[医药卫生—中药学]