In vivo endothelial transcriptomic responses to diesel engine exhaust are potentiated by hypercholesterolemia

By means of Operon V3 microarrrays, the in vivo, aortic endothelial transcriptomic responses to chronic (30 day) whole body exposure to diesel exhaust were assessed in wild type and Apo E (-/-) mice. The in vitro response of cultured Svec 4-10 cells exposed to a soluble extract extract of diesel engine particulate were similarly assessed. Mice of the Tie 2 GFP strain as well as a stable hybrid strain derived by cross of Tie 2 GFP with the Apo E (-/-) strain were bred in suffficient quantities for these experiments. Male mice at 4 months of age at the initiation of the exposure to 30 days of dilute diesel engine 94 hours/day, 5 days/week) at either 0 (control), 300 or 1000 ug/ cubic meter exhaust particulate. Following exposure, three pooled aortae of each experimental and ceach c age-matched control group were dissected from the aortic arch to the iliac bifurcation, minced and collagenolytically digested prior to FACS into Trizol. Purification yielded aortic endothelial RNA subjected to amplification with Nugen isothermal linear amplification to produce amplified cDNA. Microarrays were performed with a single dye reversal to compare the transcriptome of exposed mice to age-matched control mice. In addition, identical cultures of Svec 4-10 cells were exposed for 24 hours to a methanol extract of forklift diesel engine particulate at 50 ug/ml. Following extraction of RNA and amplification to cDNA as described above, duplicate arrays with a single dye reversal were performed.