Use of toxicogenomics for identifying genetic markers of pulmonary oedema.
This study was undertaken primarily to identify genetic markers of oedema and inflammation. Mild pulmonary injury was induced following the instillation of the oedema-producing agent, bleomycin (0.5 units). Oedema was then confirmed by conventional toxicology (lavage protein levels, free cell counts and lung/body weight ratios) and histology 3 days post-bleomycin instillation. The expression profile of 1176 mRNA species was determined for bleomycin-exposed lung (Clontech Atlas macroarray, n=9). To obtain pertinent results from these data, it was necessary to develop a simple, effective method for bioinformatic analysis of altered gene expression. Data were log10 transformed followed by global normalisation. Differential gene expression was accepted if: (a) genes were statistically significant (P < or = 0.05) from a two-tailed t test; (b) genes were consistently outside a two standard deviation (SD) range from control levels. A combination of these techniques identified 31 mRNA transcripts (approximately 3%) which were significantly altered in bleomycin treated tissue. Of these genes, 26 were down-regulated whilst only five were up-regulated. Two distinct clusters were identified, with 17 genes classified as encoding hormone receptors, and nine as encoding ion channels. Both these clusters were consistently down-regulated. The magnitude of the changes in gene expression were quantified and confirmed by Q-PCR (n = 6), validating the macroarray data and the bioinformatic analysis employed. In conclusion, this study has developed a suitable macroarray analysis procedure and provides the basis for a better understanding of the gene expression changes occurring during the early phase of drug-induced pulmonary oedema. This work has been presented orally, in part at the British Association for Lung Research Summer Meeting, University of Brighton, 3-5 September, 2003 and in full at the British Toxicology Society Annual Congress, Heriot Watt University, Edinburgh, 21-24 April 2004.
Balharry
Dominique
D
Cardiff School of Biosciences, Cardiff University, Museum Avenue, Cardiff, CF10 3US, United Kingdom. balharry@cf.ac.uk
Oreffo
Victor
V
Richards
Roy
R
D000818:Animals; D001761:Bleomycin / Q000008:administration & dosage / Q000009:adverse effects / Q000493:pharmacokinetics; D001992:Bronchoalveolar Lavage Fluid / Q000737:chemistry / Q000166:cytology; D015536:Down-Regulation; D020869:Gene Expression Profiling / Q000379:methods; D005819:Genetic Markers; D007249:Inflammation / Q000139:chemically induced / Q000175:diagnosis / Q000235:genetics; D018836:Inflammation Mediators; D008168:Lung / Q000187:drug effects / Q000473:pathology; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000473:pathology; D008297:Male; D005810:Multigene Family / Q000187:drug effects / Q000502:physiology; D009929:Organ Size / Q000187:drug effects; D011654:Pulmonary Edema / Q000139:chemically induced / Q000235:genetics* / Q000473:pathology; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013047:Specific Pathogen-Free Organisms; D013997:Time Factors; D043922:Toxicogenetics / Q000379:methods*; D014158:Transcription, Genetic; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology; D015430:Weight Gain / Q000187:drug effects
D005819:Genetic Markers; D018836:Inflammation Mediators; D001761:Bleomycin