Cigarette smoke extract regulating the expression of γ-glutamylcysteine synthetase through the pathway of PI3K/Akt-Nrf2 in the bronchial epithelial cells of rats / 中国医师杂志

ABSTRACT

Objective To observe the effect of PDK ( phosphoinositol -3-kinase, PI3K)/Akt-Nrf2 (Nuclear factor -E2 related factor) signal pathway on γ-glutamylcysteine synthetase (γ-GCS) in the bronchial epithelial cells of rats treated with cigarette smoke extract (CSE). Methods The bronchial epithelial cells were dealt with 10% concentration of CSE for different time and pretreated with PI3k inhibitor (LY294002). The expressions of Nrf2,p-Akt and γ-GCS proteins were examined by immunocytochemistry, flow cytometry, immunofluorescence and western blot. The expressions of γ-GCS mRNA were detected by reverse transcription-polymerase chain reaction (RT-PCR). Reduced glutathione(GSH) content and the lev-el of γ-CCS activi-ty were examined. Results GSH content in CSElh group was significantly decreased, but still higher in CSE3 and 6 groups compared to the control. Nrf2 protein mainly located in the cytoplasm. Nrf2 plasmosin mainly increased in the nucleus in control group, and Nrf2 nucleic protein significantly enhanced in CSE1, 3 and 6 groups. P-Akt protein was up-regulated at lh, reached its peak at 3h, declined slightly at 6h after exposure to CSE. The tendency of the percentage of p-Akt positive cells was as same as p-Akt protein. γ-GCS mRNA, protein and activity, gradually increased in CSE lh, CSE 3h,CSE 6h groups. Pretreated with LY294002, the expression of p-Akt protein was markedly decreases, while Nrf2 plasmosin expressed strongly, and γ-GCS mRNA, protein, activity and GSH content were significantly decreased compared to CSE3h group. Linear correlation analysis demonstrated that there were a positive correlation among Nrf2 and γ-GCS,γ-GCS activity, and among p-Akt and Nrf2,GSH,γ-GCS,γ-GCS activity. Conclusion P13K/Akt signal path might participate in Nrf2 nuclear translocation via regulating the expression of γ-GCS.