Dynamic Changes in Lung Function and Blood Flow During Lung Adenocarcinoma Progression in Mice by Whole-body Plethysmography and Doppler Ultrasound

Summary

This protocol uses mouse pulmonary function and Doppler ultrasound as investigative techniques, offering a non-invasive, simple, and efficient method for studying lung adenocarcinoma.

Abstract

Lung adenocarcinoma (LUAD) is a common malignant pulmonary tumor, and its progression leads to alterations in ventilation and blood flow, highlighting the distinct functional characteristics of the lung under pathological conditions. The non-invasive whole-body plethysmography (WBP) system enables real-time monitoring of lung function and airway responsiveness in awake mice, avoiding interference from tracheotomy and anesthesia. Small animal Doppler ultrasound (DU), utilizing high-frequency imaging technology, non-invasively captures microvascular structures and changes in pulmonary blood flow velocity, offering crucial support for investigating ventilation and perfusion abnormalities associated with LUAD. In this study, a LUAD model was established in A/J mice using 100 mg/kg of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). WBP combined with DU was employed to monitor pulmonary function and hemodynamic parameters in mice. The results revealed a significant decline (p < 0.05) in key metrics, including 50% expiratory flow (EF50), minute ventilation (MV), peak expiratory flow (PEF), peak inspiratory flow (PIF), enhanced pause (Penh), and tidal volume (TV), as well as a marked reduction in pulmonary arterial blood flow velocity (p < 0.05) in the model group during tumor progression. This combined approach effectively captures the functional disruptions occurring during the pathological development of LUAD in mice, providing a non-invasive and efficient method for advancing research into respiratory diseases.

Introduction

Lung adenocarcinoma (LUAD) is the most common subtype of non-small cell lung cancer¹. With increasing environmental pollution, tobacco use, and aging populations, the global prevalence of LUAD has been rising annually^2,3,4. Current studies indicate that patients experience progressive declines in pulmonary function and hemodynamic disturbances as LUAD advances^5,6,7,8. Developing non-invasive, efficient, and cost-effective met....

Protocol

All animal experiments described in this study were approved by the Ethics Committee on Animal Welfare of Chengdu University of Traditional Chinese Medicine and were conducted in compliance with relevant laws and standards for animal research ethics (20240616IV). Female inbred A/JGpt mice (7-8 weeks old) were maintained at 20-24 °C and a relative humidity of 40%-70%. Throughout the 12 h light-dark cycle, the mice were provided with standard animal feed and purified water. The animals were acclimated to this environment 7 days before beginning the experiments.

1. Creation and preparation of the animal model

1. ....

Results

To validate the feasibility, accuracy, and reproducibility of the pulmonary function and pulmonary arterial blood flow assessment methodology, we plotted the minute-by-minute mean curves of PIF, PEF, and expiratory flow at 50% of vital capacity (EF50, 10x magnified) during the 4th-week monitoring period in the blank group (Figure 1A). As shown in this panel, the three corresponding curves exhibit a high degree of conformity in shape. Despite the minimal absolute changes, particularly in EF50.......

Discussion

In this study, NNK combined with A/J mice was selected to make a lung adenocarcinoma model according to previous reports. Relevant studies have shown that NNK combined with A/J mice can start to have minimal tumor formation in the 10^th week, and stable tumor formation in the 16^th to 20^th week^18,19. Therefore, we selected the test at the 4^th week after injection to observe the changes of lung function and pulmonary arter.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
A/J mice	GemPharmatech LLC.	N000018
0.5ml EDTA tubes	Labshark	130201070
1-Butanone,4-(methylnitrosoamino)-1-(3-pyridinyl)	Gu Shi Gong Yuan Medical Equipment Co.	N589770
75% ethanol	ChengDu Chron Chemicals Co,.Ltd	2023052901
Anesthesia induction chamber	Shanghai Tawans Intelligent Technology Co., Ltd.	MZ-MR
Depilatory cream	Shanghai Yuyan Instruments Co., Ltd.	20230623
Depilatory device	Shanghai Yuyan Instruments Co., Ltd.	3180
Isoflurane	Shanghai Duma Biotechnology Co., Ltd.	DM-M9462
Medical tape	Beijing Lanbokangs Technology Co., Ltd.	19-5023
medical tricorder	MedChemexpress	69652
PBS buffer solution	Shanghai Fusheng Co., Ltd.	A-RY5256
Saline (medicine)	Beijing Biolabs Technology Co.	GL1736‌
Ultrasound coupling agent	Qisheng (Shanghai) Medical Equipment Co., Ltd.	20212060273