Nuclear factor-erythroid 2-related factor 2 (Nrf2) is a key transcription factor that plays a central role in regulating antioxidant gene expression. γ-Glutamyl cysteine synthetase (γ-GCS) is a major antioxidant gene in lung. However, it is presently unclear how Nrf2 regulates γ-GCS expression.Thus,the aim of the study is to investigate the mechanism by cigarette smoke extract(CSE) in the rat bronchial epithelial cells. Immunofluorescence showed Nrf2 protein mainly expressed in the nucleus after exposure to CSE for 1,3 and 6 hours.Immunocytochemistry and Western blot showed γ-GCS protein expression was significantly increased in the CSE 1， 3，6 hours groups than the control group(P <0.05).Nrf2 protein of nucleus strongly expressed in the CSE 1， 3， 6 hours groups.p-aPKCι/ζ protein expression was increased in the CSE 1，3 hours groups,and had significant difference as compared with the control group (P <0.05).RT-PCR showed that γ-GCS mRNA significantly enhanced in the CSE 1， 3， 6 hours groups (P <0.05). γ-GCS activity and GSH levels were significantly increased in the CSE 1， 3， 6 hours groups than the control group (P <0.05).Pretreated with aPKCι/ζ inhibitor RO318220,Nrf2 protein mainly expressed in the cytoplasm,GSH levels,p-aPKCι/ζ protein and γ-GCS mRNA,protein and activity were significantly decreased than CSE 3 hours group(P＜0.05). The correlation analysis demonstrated that there was a positive correlation between Nrf2 and γ-GCS, p-aPKCι/ζ,γ-GCS activity, and between p-aPKCι/ζ and Nrf2,γ-GCS protein, γ-GCS activity.Taken together,CSE might regulate the expression and activity of γ-GCS via aPKCι/ζ -Nrf2 signaling pathway in rat bronchial epithelial cells.
Key words
cigarette smoke extract(CSE) /
antypical protein kinase Cι/ζ /
nuclear factor-erythroid 2-related factor 2 (Nrf2) /
γ-glutamyl cysteine synthetase (γ-GCS)
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