Suppressive effect of 1-nitropyrene on benzo[a]pyrene-induced CYP1A1 protein expression in HepG2 cells

Shur Hueih Cherng, Shih Lan Hsu, Jia Ling Yang, Chang Tze Ricky Yu, Huei Lee

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

The genotoxicity of polycyclic aromatic hydrocarbons (PAHs) and nitrated PAHs may be influenced by the interaction of the compounds. In this study, our data showed that benzo[a]pyrene (BaP)-DNA adduct levels were decreased in a dose-dependent manner when the human hepatoma cell line HepG2 simultaneously treated with BaP and 1-nitropyrene (1-NP). To further investigate the molecular mechanism by which 1-NP interferes with the covalent binding of BaP to DNA, we conducted experiments to analyze the mRNA level and protein stability of cytochrome P450 1A1 (CYP1A1), which is engaged in the activation of BaP, leading to the generation of BaP-DNA adducts. Northern blot analysis presented that 1-NP attenuated BaP-induced CYP1A1 mRNA expression by 30.4-39.6% (p <0.05). Western blot analysis revealed that the co-treatment with BaP and 1-NP resulted in a significant inhibition of BaP-induced CYP1A1 protein expression (70.7-88.2%, p <0.05). However, the decrease in CYP1A1 protein levels was significantly larger than that in CYP1A1 mRNA levels. To confirm the effect of 1-NP on the CYP1A1 protein expression, in vitro proteolysis of CYP1A1 protein was evaluated. The results demonstrated that the addition of 1-NP enhanced CYP1A1 protein degradation and the proteolysis of CYP1A1 protein was inhibited by the addition of an antioxidant, dithiothreitol. In addition, the relative levels of reactive oxygen species (ROS) were elevated in HepG2 cells co-treated with BaP and 1-NP, indicating that the decrease of CYP1A1 protein level was probably attributed to the production of ROS generated by binary mixture. Taken together, these findings suggested that the transcriptional suppression and posttranslational mechanism may be involved in loss of CYP1A1 protein, causing the decrease of BaP-DNA adduct levels in the presence of binary mixtures of 1-NP and BaP.

Original languageEnglish
Pages (from-to)236-243
Number of pages8
JournalToxicology Letters
Volume161
Issue number3
DOIs
Publication statusPublished - Mar 1 2006
Externally publishedYes

Fingerprint

1-nitropyrene
Benzo(a)pyrene
Hep G2 Cells
Cytochrome P-450 Enzyme System
Proteins
Proteolysis
Polycyclic Aromatic Hydrocarbons
Binary mixtures
Messenger RNA
Reactive Oxygen Species

Keywords

  • 1-Nitropyrene
  • Benzo[a]pyrene
  • Chemical interaction
  • CYP1A1
  • Proteolysis
  • Transcriptional suppression

ASJC Scopus subject areas

  • Toxicology

Cite this

Suppressive effect of 1-nitropyrene on benzo[a]pyrene-induced CYP1A1 protein expression in HepG2 cells. / Cherng, Shur Hueih; Hsu, Shih Lan; Yang, Jia Ling; Yu, Chang Tze Ricky; Lee, Huei.

In: Toxicology Letters, Vol. 161, No. 3, 01.03.2006, p. 236-243.

Research output: Contribution to journalArticle

Cherng, Shur Hueih ; Hsu, Shih Lan ; Yang, Jia Ling ; Yu, Chang Tze Ricky ; Lee, Huei. / Suppressive effect of 1-nitropyrene on benzo[a]pyrene-induced CYP1A1 protein expression in HepG2 cells. In: Toxicology Letters. 2006 ; Vol. 161, No. 3. pp. 236-243.
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abstract = "The genotoxicity of polycyclic aromatic hydrocarbons (PAHs) and nitrated PAHs may be influenced by the interaction of the compounds. In this study, our data showed that benzo[a]pyrene (BaP)-DNA adduct levels were decreased in a dose-dependent manner when the human hepatoma cell line HepG2 simultaneously treated with BaP and 1-nitropyrene (1-NP). To further investigate the molecular mechanism by which 1-NP interferes with the covalent binding of BaP to DNA, we conducted experiments to analyze the mRNA level and protein stability of cytochrome P450 1A1 (CYP1A1), which is engaged in the activation of BaP, leading to the generation of BaP-DNA adducts. Northern blot analysis presented that 1-NP attenuated BaP-induced CYP1A1 mRNA expression by 30.4-39.6{\%} (p <0.05). Western blot analysis revealed that the co-treatment with BaP and 1-NP resulted in a significant inhibition of BaP-induced CYP1A1 protein expression (70.7-88.2{\%}, p <0.05). However, the decrease in CYP1A1 protein levels was significantly larger than that in CYP1A1 mRNA levels. To confirm the effect of 1-NP on the CYP1A1 protein expression, in vitro proteolysis of CYP1A1 protein was evaluated. The results demonstrated that the addition of 1-NP enhanced CYP1A1 protein degradation and the proteolysis of CYP1A1 protein was inhibited by the addition of an antioxidant, dithiothreitol. In addition, the relative levels of reactive oxygen species (ROS) were elevated in HepG2 cells co-treated with BaP and 1-NP, indicating that the decrease of CYP1A1 protein level was probably attributed to the production of ROS generated by binary mixture. Taken together, these findings suggested that the transcriptional suppression and posttranslational mechanism may be involved in loss of CYP1A1 protein, causing the decrease of BaP-DNA adduct levels in the presence of binary mixtures of 1-NP and BaP.",
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AU - Yu, Chang Tze Ricky

AU - Lee, Huei

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N2 - The genotoxicity of polycyclic aromatic hydrocarbons (PAHs) and nitrated PAHs may be influenced by the interaction of the compounds. In this study, our data showed that benzo[a]pyrene (BaP)-DNA adduct levels were decreased in a dose-dependent manner when the human hepatoma cell line HepG2 simultaneously treated with BaP and 1-nitropyrene (1-NP). To further investigate the molecular mechanism by which 1-NP interferes with the covalent binding of BaP to DNA, we conducted experiments to analyze the mRNA level and protein stability of cytochrome P450 1A1 (CYP1A1), which is engaged in the activation of BaP, leading to the generation of BaP-DNA adducts. Northern blot analysis presented that 1-NP attenuated BaP-induced CYP1A1 mRNA expression by 30.4-39.6% (p <0.05). Western blot analysis revealed that the co-treatment with BaP and 1-NP resulted in a significant inhibition of BaP-induced CYP1A1 protein expression (70.7-88.2%, p <0.05). However, the decrease in CYP1A1 protein levels was significantly larger than that in CYP1A1 mRNA levels. To confirm the effect of 1-NP on the CYP1A1 protein expression, in vitro proteolysis of CYP1A1 protein was evaluated. The results demonstrated that the addition of 1-NP enhanced CYP1A1 protein degradation and the proteolysis of CYP1A1 protein was inhibited by the addition of an antioxidant, dithiothreitol. In addition, the relative levels of reactive oxygen species (ROS) were elevated in HepG2 cells co-treated with BaP and 1-NP, indicating that the decrease of CYP1A1 protein level was probably attributed to the production of ROS generated by binary mixture. Taken together, these findings suggested that the transcriptional suppression and posttranslational mechanism may be involved in loss of CYP1A1 protein, causing the decrease of BaP-DNA adduct levels in the presence of binary mixtures of 1-NP and BaP.

AB - The genotoxicity of polycyclic aromatic hydrocarbons (PAHs) and nitrated PAHs may be influenced by the interaction of the compounds. In this study, our data showed that benzo[a]pyrene (BaP)-DNA adduct levels were decreased in a dose-dependent manner when the human hepatoma cell line HepG2 simultaneously treated with BaP and 1-nitropyrene (1-NP). To further investigate the molecular mechanism by which 1-NP interferes with the covalent binding of BaP to DNA, we conducted experiments to analyze the mRNA level and protein stability of cytochrome P450 1A1 (CYP1A1), which is engaged in the activation of BaP, leading to the generation of BaP-DNA adducts. Northern blot analysis presented that 1-NP attenuated BaP-induced CYP1A1 mRNA expression by 30.4-39.6% (p <0.05). Western blot analysis revealed that the co-treatment with BaP and 1-NP resulted in a significant inhibition of BaP-induced CYP1A1 protein expression (70.7-88.2%, p <0.05). However, the decrease in CYP1A1 protein levels was significantly larger than that in CYP1A1 mRNA levels. To confirm the effect of 1-NP on the CYP1A1 protein expression, in vitro proteolysis of CYP1A1 protein was evaluated. The results demonstrated that the addition of 1-NP enhanced CYP1A1 protein degradation and the proteolysis of CYP1A1 protein was inhibited by the addition of an antioxidant, dithiothreitol. In addition, the relative levels of reactive oxygen species (ROS) were elevated in HepG2 cells co-treated with BaP and 1-NP, indicating that the decrease of CYP1A1 protein level was probably attributed to the production of ROS generated by binary mixture. Taken together, these findings suggested that the transcriptional suppression and posttranslational mechanism may be involved in loss of CYP1A1 protein, causing the decrease of BaP-DNA adduct levels in the presence of binary mixtures of 1-NP and BaP.

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