病毒性肺炎湿热证小鼠模型的建立及评价*

作者：张庭瑞1,2,3，杨上松1,2，向 纯1,3，温伟波1,2，李 钦1,2

单位：1.云南中医药大学，云南 昆明 650000； 2.云南省中西医结合慢病防治重点实验室，云南 昆明 650000； 3.仙桃市中医医院，湖北 仙桃 433099

引用：引用：张庭瑞，杨上松，向纯，温伟波，李钦.病毒性肺炎湿热证小鼠模型的建立及评价[J].中医药导报,2025,31(3):51-59.

DOI：10.13862/j.cn43-1446/r.2025.03.009

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摘要：

目的：构建并评价病毒性肺炎湿热证小鼠模型。方法：将32只SPF级雄性C57/BL6小鼠随机分为正常组、湿热组、病毒模拟物组、湿热+病毒模拟物组，每组8只。湿热组采用高脂饮食和气候箱湿热暴露14 d的方法构建小鼠湿热证模型，病毒模拟物组采用气管内滴注Poly(I:C)的方法构建小鼠病毒性肺炎肺损伤模型，湿热+病毒模拟物组采用高脂饮食和气候箱湿热暴露14 d叠加气管内滴注Poly(I:C)复合造模法，建立病毒性肺炎湿热证小鼠模型。观测小鼠体质量、食量、饮水量及证候评分变化，分别用酶联免疫法、酶联免疫组织化学染色法、血气分析仪、透射电镜观察血脂代谢（血清HDL-C及LDL-C水平）、胃肠功能（血清MTL、SP水平）、能量代谢（肝组织Na+-K+-ATP酶活性）、水液转运（肺组织AQP1、AQP5、Na+-K+-ATP水平，肺组织细胞线粒体结构）、肺泡-毛细血管屏障（肺组织W/D，BALF中总细胞、总蛋白水平），炎症反应（BALF中TNF-α、IL-1β、IL-6水平）和肺生理功能（动脉血PaO2、PaCO2、FiO2水平），HE染色观察肺组织病理学变化，16SrRNA测序技术观察小鼠肠道菌群。结果：与正常组比较，湿热+病毒模拟物组小鼠的食量、饮水量、体质量下降，湿热证候积分，血清LDL-C、MTL、SP水平，肺组织AQP1、AQP5表达，肺组织W/D、BALF中总细胞数、总蛋白浓度，BALF中TNF-α、IL-1β、IL-6水平，动脉血PaCO2升高（P＜0.05或P＜0.01）；肝、肺组织Na+-K+-ATP酶活性，动脉血PaO2、PaO2/FiO2水平下降（P＜0.05或P＜0.01）；肺组织结构，细胞线粒体结构破坏，肺组织病理损伤评分升高。肠道菌群发生了改变，在属水平主要是拟杆菌、副拟杆菌、屎豆属菌、毛螺菌、NK4A136菌、罗氏菌、NK4A214菌、埃希氏菌相对丰度存在差异，肠道菌群的功能则在需氧化能异养，硝酸盐还原，动物寄生生物或共生体，人类病原体，发酵等方面表现出差异。结论：高脂饮食和气候箱湿热暴露14 d叠加气管滴注Poly(I:C)复合造模法可构建病毒性肺炎湿热证模型，造模方法安全易行，模型稳定可靠。

关键词：病毒性肺炎；湿热证；病证结合；模型评价；Poly(I:C)

Abstract：

Objective: To construct and evaluate a mouse model of simulated viral pneumonia with damp heat syndrome. Method: Totally 32 SPF grade male C57/BL6 mice were randomly divided into normal group, damp heat group, virus mimic group and damp heat virus mimic group, with 8 mice in each group. The damp heat group constructed a mouse model of damp heat syndrome using a combination of rich and sweet diet and 14 days exposure to humid heat in a climate box. The virus mimetic group constructed a mouse model of simulated viral pneumonia using tracheal instillation of Poly (I∶C). The damp heat virus mimetic group established a mouse model of simulated viral pneumonia damp heat syndrome using a combination of rich and sweet diet and 14 days exposure to humid heat in a climate box through a superimposed gas tube instillation of Poly (I∶C). The changes in body weight, food intake, dietary intake and syndrome scores were observed. Enzyme-linked immunosorbent assay, enzyme-linked immunohistochemistry staining, blood gas analyzer, and transmission electron microscopy were applied to observe lipid metabolism (serum HDL-C and LDL-C levels), gastrointestinal function (serum MTL and SP level), energy metabolism (Na+-K+-ATPase activity in liver tissue), water liquid transport (AQP1, AQP5 and Na+-K+-ATPase levels in lung tissue, mitochondrial structure of lung tissue cells), alveolar capillary barrier (lung tissue W/D, total cell and protein levels in BALF), inflammatory response (TNF-α, IL-1β and IL-6 in BALF), and pulmonary physiological function (arterial blood PaO2, PaCO2 and FiO2 levels). HE staining was used to observe pathological changes in lung tissue, and 16s RNA sequencing technology was used to observe the gut microbiota of mice. Results: Compared with the normal group, the food intake, water intake and weight decreased in damp heat virus mimic group. Damp heat syndrome score, serum LDL-C, MTL and SP levels, lung tissue AQP1 and AQP5 expression, lung tissue W/D, total cell count and total protein concentration in BALF, TNF-α, IL-1β and IL-6 levels in BALF, and elevated arterial PaCO2 levels increased in damp heat virus mimic group (P＜0.05 or P＜0.01); Na+-K+-ATPase activity in liver and lung tissues, arterial blood PaO2, and PaO2/FiO2 levels decrease in damp heat virus mimic group (P<0.05 or P<0.01); The structure of lung tissue and mitochondrial structure of cells are disrupted, and the pathological damage score of lung tissue was increased in damp heat virus mimic group. The gut microbiota has undergone changed in damp heat virus mimic group, mainly at the genus level including Bacteroides, Parabacteroides, Faecalitalea, Lachnospiraceae, NK4A136 group, Roseburia, NK4A214 group, and Escherichia-Shigella, while the functions of gut microbiota was varied in areas such as aerobic chemoheterotrophy, nitrate reduction, animal parasites or symbionts, human pathogens, and fermentation. Conclusion: The compound modeling method of adding Poly (I∶C) through a stacked gas tube drip after 14 days of exposure to humid heat in a fat and sweet diet and climate box can construct a viral pneumonia damp heat syndrome model. The modeling method is safe and easy to implement, and the model is stable and reliable.

Key words：viral pneumonia; damp heat syndrome; combination of disease and syndrome; model evaluation; Poly(I:C)

发布时间：2025-12-14

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