Mouse Model for Acute Respiratory Distress Syndrome (ARDS)

Mouse were anesthetized via enterocoelia and injected LPS (6mg/kg) into lung using the method of exposure tracheotomy dripping. Then revolve mouse vertically to diffuse LPS at lung evenly. Building the mouse model for endotoxic acute lung injury. Perhaps LPS given to mouse via caudal vein or enterocoelia, building the mouse ARDS model induced by systemic inflammatory response.
6 hours, 12 hours, 24 hours and 48 hours after LPS dripping, take 3 mouse respectively. Collect bronchoalveolar lavage fluid (BALF) and lung tissue.
Sample collection:
BALF: After blood collection, open its chest, separate mediastina, inject 3ml saline from trachea to the lung. Every time lavage thrice, collect the douche. Centrifuged it at 4℃, 3000r/min for 15 min, take the supernatant.
Lung tissue: Mouse were anesthetized via enterocoelia, the anticoagulant treatment were carried out with 1ml heparinized saline through the right ventricular cavity, and different lung tissues were taken from the chest immediately.

Diff-Quik staining and cell count:
Obtained BALF for Diff-Quik staining, observation and count of neutrophils, macrophages and lymphocytes.
Wet/dry weight ratio of lung:
Wet/dry weight ratio of lung directly reflect pneumonedema, and also reflect severity of lung injury. Kill mouse, cut their weasand, take complete lung and weigh wet. Then dry the lung in a 65 ℃ incubator, 24h later weigh weigh dry and count wet/dry weight ratio of lung.
Determination of lung permeability by Evans blue:
30min before the end of the experiment, intravenous injection of Evans blue (EBD, 30mg/kg), After the experiment, open chest and cut right chamber, wash lesser circulation, take double lung completely, cut off the trachea, clean and dry lung tissue. Homogenate after weighing, soaking lung tissue homogenate with 1ml formamide per 100mg lung tissue, incubated at 37℃ for 24h, centrifuge at 5000r/min, take supernatant to measure EBD concentration.

Histopathology observation of lung injury (HE staining)
Take the right lung, 4% poly formaldehyde solution fixed, embedded to make the paraffin section (5μm), HE staining.
Scoring methods: Bleeding and edema scores were obtained from 6 fields.

Detect the concentration of TNF-a, IL-6 and IL-1β in bronchoalveolar lavage fluid (BALF) by ELISA.

SPSS software is used for statistical analysis, measurement data to mean ± standard deviation (x ±ｓ), using t test and single factor analysis of variance for group comparison , P<0.05 indicates there was a significant difference, P<0.01 indicates there are very significant differences.