Lipopolysaccharide-Induced Acute Lung Injury in Mice for Exosome and Immune Analysis

Abstract

Source: Yuan, Z., et al. Bronchoalveolar Lavage Exosomes in Lipopolysaccharide-induced Septic Lung Injury. J. Vis. Exp. (2018)

This video demonstrates an in vivo mouse model for inducing and analyzing acute lung injury (ALI) using intraperitoneal administration of bacterial lipopolysaccharide. The inflammatory cascade triggered by alveolar macrophages and neutrophils leads to epithelial and endothelial barrier disruption, allowing fluid and immune cells to accumulate in the alveoli. Alveolar fluid is then collected post-mortem by tracheal lavage for downstream analysis of immune components and exosomes.

Protocol

1. Mouse Acute Septic Lung Injury Model

1. Use 6-8 weeks-old male wild-type C57BL/6J mice (weight 20-22 g) for the animal model, and perform all procedures using aseptic techniques and instruments. Autoclave all surgical instruments, including forceps and scissors for 20 min at 121 °C.
2. Randomly distribute mice into experimental and control groups. Restrain mice on a surgical board. Treat mice with Escherichia coli lipopolysaccharide (LPS, 15 mg/kg) by intraperitoneal route. According to the individual body weight, dilute appropriate LPS dosages to 0.1 mL phosphate- buffered saline (PBS) per mouse, then inject LPS solution using a 1 mL syringe with 27 G needle and inject an equivalent volume of PBS to control animals.
3. Keep mice in the animal BSL2 facility, which consists of clean cages at appropriate room temperature.
4. 24 h later, euthanize mice with CO₂ inhalation.
5. Select the surgical area in the center of the mouse neck. Make a small incision (approximately 5 mm), and gently separate muscles for the access to the trachea using blunt forceps until the trachea is exposed.
6. Withdraw mouse broncho-alveolar lavage fluid (BALF) with 10 mL sterile syringe with a 27 G needle. First, gently, insert the needle into the trachea and inject 1 mL of sterile PBS slowly into the trachea, then draw BALF into a 3 mL disposable syringe with a 27 G needle. Upon completion of the procedure, dispose of syringes and needles in biohazard sharps containers.
7. Treat surgical instruments with chlorine dioxide, and then sterilize all surgical instruments by autoclaving for 20 min at 121 °C.

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Lipopolysaccharide	Sigma-Aldrich	Escherichia coli 055:B5
PBS pH7.2(1X)	Life technologies	20012-027