Open Tracheostomy Gastric Acid Aspiration Murine Model of Acute Lung Injury Results in Maximal Acute Nonlethal Lung Injury

Summary

This protocol induces acute lung injury in a mouse that has close fidelity to the pathogenesis of acid pneumonitis observed in humans. We generate a maximal acute nonlethal low pH lung injury and account for differences in rodent-human anatomic respiratory structure using an open tracheostomy coupled with circumferential pressure release.

Abstract

Acid pneumonitis is a major cause of sterile acute lung injury (ALI) in humans. Acid pneumonitis spans the clinical spectrum from asymptomatic to acute respiratory distress syndrome (ARDS), characterized by neutrophilic alveolitis, and injury to both alveolar epithelium and vascular endothelium. Clinically, ARDS is defined by acute onset of hypoxemia, bilateral patchy pulmonary infiltrates and non-cardiogenic pulmonary edema. Human studies have provided us with valuable information about the physiological and inflammatory changes in the lung caused by ARDS, which has led to various hypotheses about the underling mechanisms. Unfortunately, difficulties determining the etiology of ARDS, as well as a wide range of pathophysiology have resulted in a lack of critical information that could be useful in developing therapeutic strategies.

Translational animal models are valuable when their pathogenesis and pathophysiology accurately reproduce a concept proven in both in vitro and clinical settings. Although large animal models (e.g., sheep) share characteristics of the anatomy of human trachea-bronchial tree, murine models provide a host of other advantages including: low cost; short reproductive cycle lending itself to greater data acquisition; a well understood immunologic system; and a well characterized genome leading to the availability of a variety of gene deletion and transgenic strains. A robust model of low pH induced ARDS requires a murine ALI that targets mainly the alveolar epithelium, secondarily the vascular endothelium, as well as the small airways leading to the alveoli. Furthermore, a reproducible injury with wide differences between different injurious and non-injurious insults is important.

The murine gastric acid aspiration model presented here using hydrochloric acid employs an open tracheostomy and recreates a pathogenic scenario that reproduces the low pH pneumonitis injury in humans. Additionally, this model can be used to examine interaction of a low pH insult with other pulmonary injurious entities (e.g., food particles, pathogenic bacteria).

Introduction

ARDS is characterized by widespread lung inflammation and is clinically seen as acute shortness of breath with hypoxemia. These symptoms often occur less than 24 h after an inciting event such as trauma, sepsis, blood transfusion related reactions or aspiration. It is characterized histopathologically by neutrophilic alveolitis (i.e., widespread inflammation) localized to alveolar epithelium and vascular endothelium leading to protein leakage and subsequently hyaline membrane formation. Aspiration is categorized as chemical pneumonitis or aspiration pneumonia.¹ The acidic component of the gastric aspiration contributes to both the pneu....

Protocol

All materials and equipment need to be gathered and adequately organized prior to procedure. The procedure is to be conducted with seamless transitions from one step to the next in order to provide consistent, reproducible data. This protocol follows institutional policies set by the Institutional Animal Care and Use Committee at the Buffalo Veterans Affairs Medical Center.
NOTE: Both male and female C57BL/6 mice were used for this protocol. There is no significant difference of albumin leakage between sexes.

1. Hydrochloric Acid Preparation

1. Make 56.2 mM HCl_sterile by adding 281 µL of 1 N HCl to 4.72 mL sterile normal saline (S....

Results

Pulmonary Histopathology of the Murine Model of Acid Aspiration Pneumonitis

Mice were injured as described above and lungs were removed 24 h post low pH insult, sectioned and H&E stained (Figure 1). Necrotic cells, loss of lung parenchymal architecture, cells and debris within airspaces and significant PMN infiltration are clearly observed. Similar to aspiration clinically, our model results in a heterogeneous .......

Discussion

The objective was to develop an ALI animal model using gastric acid aspiration that closely resembles the pathophysiology that occurs in humans during development of acid pneumonitis and subsequent ARDS. In developing a model, we chose an animal species that offers high throughput data acquisition due to its low cost, short reproductive cycle, and a well understood immunological system with an abundance of investigative tools (i.e., monoclonal antibodies, transgenic strains).

ALI foll.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
syringe, 1 cc	Becton Dickinson	309628
syringe, 5 cc	Becton Dickinson	309646
needle, 22 G x 1 1/2"	Becton Dickinson	305159
needle, 26 G x 1 1/2"	Becton Dickinson	305111
1-O Braided Silk Suture	Harvard Apparatus	517730
3" Curved tissue serrated forceps	Fine Science Tools	11065-07
3" Curved tissue "toothed" forceps, 1 x 2 teeth	Fine Science Tools	11067-07
4" curved micro dissecting scissors	Fine Science Tools	14061-10
bone cutting spring scissors	Fine Science Tools	16144-13
3 1/2" curved locking hemostat	Fine Science Tools	13021-12
Disposable Skin Stapler	3M	DS-25
tracheal cannula (20 gauge x 1/2" stainless steal tubing adapter)	Becton Dickinson	408210
60-degree Incline Dissection Board
0.5% Bupivacaine
Isoflurane
Betadine and "Q-tip" cotton applicator