Methods

Subjects enrolled in the Phase II Randomized Placebo-controlled Trial of Omega-3 Fatty Acids for the Treatment of Acute Lung Injury trial (6) conducted between 2006 and 2008 were included in this study. We performed genome-wide expression analysis of total RNA isolated from paired AM and PBM samples purified from BAL fluid and peripheral blood specimens, respectively, collected from patients within 48 hours of the diagnosis of ARDS. Negative selection for AMs and PBMs was achieved by incubating cells with antibody-labeled microbeads containing the following markers: CD3, CD15, CD19, CD235a, CD294, and CD326. We did not use antibodies for CD294 (eosinophils) and CD326 (epithelial cells) for the blood samples because mononuclear cells were isolated from whole blood before antibody incubation via polyester gel centrifugation. RNA extracted from isolated cells was assessed for purity, and then hybridized to an Illumina HumanRef-8 BeadChip that was inclusive of 18,415 unique genes.