Epithelial cells lining the airways are thought to play a prominent role in respiratory diseases. We utilized cDNA representational difference analysis to identify the genes in which expression is induced by the proinflammatory cytokines tumor necrosis factor-alpha and interleukin-1beta in primary human bronchial epithelial cells and hence are relevant to airway inflammation. Hybridization of the subtraction product to arrayed cDNAs indicated that known tumor necrosis factor-alpha- and interleukin-1beta-inducible genes such as B94, Zfp36, and regulated on activation normal T cell expressed and secreted were represented, confirming the success of the subtraction experiment. A 1,152-clone library potentially representing genes with higher transcript levels in cytokine-treated human bronchial epithelial cells was generated and sequenced. Sequence similarity searches indicated that these clones represented 57 genes of known function, 1 gene of unknown function, 6 expressed sequence tags, and 2 novel sequences. The expression of 19 of these clones was studied by a combination of Northern blotting and RT-PCR analyses and confirmation of differential expression for 10 known genes, 2 expressed sequence tags, and a novel sequence not represented in any of the public databases was obtained. Thus cDNA representational difference analysis was utilized to isolate known and novel differentially expressed genes, which putatively play a role in airway inflammation.
|Journal||American journal of physiology. Lung cellular and molecular physiology|
|Publication status||Published - 1 May 2001|