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Gene-expression profiling using suppression-subtractive hybridization and cDNA microarray in rat mononuclear cells in response to welding-fume exposure

Center for Occupational Toxicology, Occupational Safety & Health Research Institute, KOSHA, Daejeon, Republic of Korea, Division of Molecular & Life Sciences, College of Science & Technology, Hanyang University, Ansan, Republic of Korea

Kun Koo Park

Pharmacogenechips Inc., Chuncheon, Republic of Korea

Jae Hyuck Sung

Center for Occupational Toxicology, Occupational Safety & Health Research Institute, KOSHA, Daejeon, Republic of Korea

Yong Hyun Chung

Center for Occupational Toxicology, Occupational Safety & Health Research Institute, KOSHA, Daejeon, Republic of Korea

Jeong Hee Han

Center for Occupational Toxicology, Occupational Safety & Health Research Institute, KOSHA, Daejeon, Republic of Korea

Key Seung Cho

Division of Molecular & Life Sciences, College of Science & Technology, Hanyang University, Ansan, Republic of Korea

Kwang Jong Kim

Center for Occupational Toxicology, Occupational Safety & Health Research Institute, KOSHA, Daejeon, Republic of Korea

Il Je Yu

Center for Occupational Toxicology, Occupational Safety & Health Research Institute, KOSHA, Daejeon, Republic of Korea, u1670916{at}chollian.net

Welders with radiographic pneumoconiosis abnormalities have shown a gradual clearing of the X-ray identified effects following removal from exposure. In some cases, the pulmonary fibrosis associated with welding fumes appears in a more severe form in welders. Accordingly, for the early detection of welding-fume-exposure-induced pulmonary fibrosis, the gene expression profiles of peripheral mononuclear cells from rats exposed to welding fumes were studied using suppression-subtractive hybridization (SSH) and a cDNA microarray. As such, Sprague-Dawley rats were exposed to a stainless steel arc welding fume for 2 h/day in an inhalation chamber with a 107.59 ^/ 2.6 mg/m³ concentration of total suspended particulate (TSP) for 30 days. Thereafter, the total RNA was extracted from the peripheral blood mononuclear cells, the cDNA synthesized from the total RNA using the SMART^TM PCR cDNA method, and SSH performed to select the welding-fume-exposure-regulated genes. The cDNAs identified by the SSH were then cloned into a plasmid miniprep, sequenced and the sequences analysed using the NCBI BLAST programme. In the SSH cloned cDNA microarray analysis, five genes were found to increase their expression by 1.9-fold or more, including Rgs 14, which plays an important function in cellular signal transduction pathways; meanwhile 36 genes remained the same and 30 genes decreased their expression by more than 59%, including genes associated with the immune response, transcription factors and tyrosine kinases. Among the 5200 genes analysed, 256 genes (5.1%) were found to increase their gene expression, while 742 genes (15%) decreased their gene expression in response to the welding-fume exposure when tested using a commercial 5.0k DNA microarray. Therefore, unlike exposure to other toxic substances, prolonged welding-fume exposure was found to substantially downregulate many genes.

Key Words: diagnosis • cDNA microarray • gene expression profile • lung fibrosis • suppression-subtractive hybridization • welding fume

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