Expression data from the cells of rat bronchoalveolar lavage fluid

Acute lung injury (ALI) refers to a clinical syndrome characterized by bilateral lung injury, severe lung diffuse failure and hypoxemia caused by non-cardiogenic pulmonary edema.Sepsis is the leading etiology of ALI and a common admission to the intensive care unit, which induces pulmonary inflammation leading disruption of endothelial-epithelial barriers by surge release of pro-inflammatory cytokines that increases the permeability of the alveolar-capillary membrane, pulmonary infiltration, and edema.Ultimately, gas exchange across the alveolar-capillary membrane is severely impaired and acute respiratory failure and hypoxia occur. ALI patients may suffer from pulmonary inflammation and hypoxia simultaneously or sequentially, those two pathophysiological processes may interact mutually and contribute together to the development of ALI. LPS is the most important biological mediator of sepsis induce secretion of inflammatory cytokines including TNF-α, IL-1, and IL-6 from many cell types in response to bacterial toxins. Thus LPS has been commonly used to establish inflammatory ALI models of rats and mice. Clinically, hypoxia commonly coexists with sepsis; however, the role of hypoxia on the development of inflammatory ALI is unclear. The understanding of interaction of hypoxia and inflammation in ALI is of the importance for the treatment of ALI. Briefly, 8-week-old male Sprague-Dawley (SD) rats, weighing 200 ± 20 g, were housed in microisolator cages with specific pathogen-free (SPF) condition and free access to water and food. Before experimentation, rats were housed for 3 days to adapt them to the environment. The rats were randomly divided into four experimental groups: (i) CTL group, naïve control rats with normoxia, (ii) LPS group, with LPS treatment with normoxia, (iii) HPO group, with exposure of acute hypobaric hypoxia, (iv) COMB group, with combined exposures of acute hypobaric hypoxia and LPS. For LPS administration, the animals were subjected to a tail vein injection of a 0.5 mg/kg LPS in saline at an injection volume of 0.2 ± 0.02 mL (LPS: 1 mg / ml). For exposure of hypobaric hypoxia, the animals were subjected to stay in the decompression chamber for 6 hours with an ambient air pressure of 405.35 mmHg (approx. 0.53atm, or equivalent of 5000 m altitude, or the equivalent of 11.2% O2). The animals in COMB groups who received double-stimulus were first subjected to LPS administration, then stay in the decompression chamber, the time used about twenty minutes when it achieved the ambient air pressure of 5000 m altitude from the plains. The rats in CTL group were subjected to a tail vein injection of 0.2 ± 0.02 mL of saline (0.9% NaCl water). All animals were fasted and banned water during the experiment. Per treatment received by the animal, the operation of collecting specimens were done in (normal or hypoxic) condition. Before specimen collection rats were subjected to anesthesia with a single dose of chloral hydrate (100 mg/kg, i.p.) and their lungs were removed. Bronchoalveolar lavage fluid (BALF) was obtained from the right lung as follows. First, the left bronchus was ligated with sutures and the right bronchus was cannulated. Then a 3 ml saline was introduced into the right lungs, which were then gently manipulated, followed by the withdrawal of lavage fluid. This procedure was repeated 3 times for each rat. The collected BALF was centrifuged. The cells fraction was suspended in PBS for the evaluation of gene expression profiling. For microarray analysis, we pooled the total RNA from three animals in the same treatment group as a pooled RNA sample, and a total of 3 pooled RNA samples underwent microarray analysis in each group. We used cluster analysis to analyze the total 12 RNA samples to find the common genes identified by paired t-test. The gene expression profiles were determined using Gene Chip Rat Genome 230 2.0 Array (Affymetrix).