Methods for Detecting Cough and Airway Inflammation in Mice

Summary

Here, we describe the measurement of cough using a noninvasive and real-time whole-body plethysmography (WBP) system and the normative procedures for harvesting tissue samples of mice and introduce some methods to assess airway inflammation.

Abstract

Chronic cough, which lasts for more than 8 weeks, is one of the most common complaints requiring medical attention, and patients suffer from a huge socioeconomic burden and a marked decrement in quality of life. Animal models can mimic the complex pathophysiology of the cough and are important tools for cough research. The detection of cough sensitivity and airway inflammation is of great significance for studying the complex pathological mechanism of cough. This article describes the measurement of cough using a noninvasive and real-time whole-body plethysmography (WBP) system and the normative procedures for harvesting tissue samples (including blood, lung, spleen, and trachea) of mice. It introduces some methods to assess airway inflammation, including pathological changes in hematoxylin and eosin (HE)-stained lung and trachea sections, the total protein concentration, the uric acid concentration, and the lactate dehydrogenase (LDH) activity in the supernatant of bronchoalveolar lavage fluid (BALF), and the leukocytes and differential cell counts of BALF. These methods are reproducible and serve as valuable tools to study the complex pathophysiology of cough.

Introduction

Cough is an important defense behavior to maintain airway patency and protect the lungs from potentially harmful substances. However, when dysregulated, cough becomes a pathological condition¹. Chronic cough, usually defined as lasting eight or more weeks, is one of the most frequent symptoms requiring medical attention². Because chronic cough frequently persists for years, patients suffer from a huge socioeconomic burden and a marked decrement in quality of life^3,4,5. Chronic cough is widely considered a c....

Protocol

All the procedures were approved by the Animal Care and Use Committee of Guangzhou Medical University (20240248) and were performed in strict accordance with approved guidelines. Male specific pathogen-free C57BL/6 mice weighing 20-25 g were used in this study. All mice were housed under controlled temperature (22 ± 2 °C), humidity (50% ± 20%), and lighting (6:30 AM to 6:30 PM) in solid bottom cages with food and water available ad libitum. The time line of the protocol is shown in Fig.......

Discussion

Some chronic refractory and postinfectious coughs are common conditions associated with respiratory virus infection²⁷. In order to better evaluate cough sensitivity and airway inflammation, a mouse model of cough was established using the H1N1 virus in this study. Appropriate mouse cough models should be selected for other studies according to the purpose of the study. Most previous studies used the guinea pig as an animal model in mechanistic studies or new drug trials for cough^.......

Materials

Name	Company	Catalog Number	Comments
4% paraformaldehyde	Biosharp	BL539A
Buxco Small Animal Whole Body Plethysmography System	DSI	—
Calcium-free and magnesium-free Hank’s Balanced Salt Solution	Beyotime	C0219
Citric acid	Sigma-Aldrich	C2404
Hematoxylin-Eosin	BASO Biotechnology	BA-4098
Heparin sodium	Alfa Aesar	A16198
Influenza A/California/7/2009 (H1N1) virus	ATCC	VR-1894
Isoflurane	RWD	R510-22
Lactate dehydrogenase assay kit	Nanjing Jiancheng Bioengineering Institute	A020-2-2
Normal saline	Guangzhou Zhongbo Biotechnology	1234-1
Pasteur pipet	NEST	318415
Pentobarbital sodium	Merck	P3761
Phosphate buffered saline	Meilunbio	MA0015
Total protein assay kit	Nanjing Jiancheng Bioengineering Institute	A045-3
Uric acid assay kit	Thermo Fisher Scientific	A22181