| Co-Investigator(Kenkyū-buntansha) |
ISHIKAWA Tetsuya Kawasaki Medical School, Biochemistry, Assistant, 医学部, 助手 (90221754)
HIDAKA Kazuo Kawasaki Medical School, Biochemistry, Lecturer, 医学部, 講師 (00069064)
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| Research Abstract |
First, we studied the effect of nitric oxide (NO) and glutathione (GSH) levels on PQ toxicity in A549 cells and rat organs exposed to PQ.Our in vitro models demonstrated that NO showed both beneficial and deleterious actions, depending on the concentrations produced and model system used. We detected, in vivo models, a remarkable increase in iNOS mRNA level and deposit of nitrotyrosine in liver tissues, whereas gene expressions for cytokines increased in lung tissues, suggesting that the mechanisms of PQ poisoning are different among organs exposed to PQ.We found a specific increase in GSH levels, the activities and the expression levels of the rate-limiting enzymes in GSH synthesis ; γ-glutamylcysteine synthetase (γ-GCS) and γ-glutamyl transpeptidase (γ-GT) in lungs exposed to PQ.Also, the GSH levels in A549 cells slightly increased by incubation with PQ.Next, by analyzing cholesterol-derived hydroperoxide, we found 7 α-OOH and 7 β-OOH accumulations in rat liver and kidney exposed to low dose of PQ, especially they reflected greater oxidative stress in the pathology of kidney. Finally, to identify genes that are differentially expressed in response to oxidative stress due to PQ, we carried out RT-PCR based differential display experiments using cDNAs from rat lungs that were treated for 3 hrs with 20 mg/kg PQ.Numerous cDNA bands were identified that were up-regulated and down-regulated ; a few clone were homologous to known genes and the remainders were unknown. Unknown six different cDNA clones were isolated and sequenced. The differential mRNA expression of these clones could be verified using RT-PCR method. These results may prove useful in elucidating the molecular changes during pulmonary fibrosis evoked by PQ intake.
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