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The Expression of 5-lipoxygenase and the Activation and Cytotoxicity of Acrylonitrile in Human Bronchial Epithelial Cell

Author: YangFeng
Tutor: HuJianAn
School: Central South University
Course: Occupational and Environmental Health
Keywords: 5-lipoxygenase co-oxidation protein expression cytotoxicity DNA damage
CLC: R113
Type: Master's thesis
Year: 2010
Downloads: 21
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Abstract


[Objectives] The effects of acrylonitrile to 5-lipoxygenase protein expression and cytotoxicity and DNA damage were been investigated in HBE cells, with the experiment that can affirm the co-oxidation of acrylonitrile mediated by SLO in the vitro-enzymic system, in order to approach the metabolism of xenobiotic by 5-LOX and provide further proof that lipoxygenase is the alternative pathway for the oxidation of xenobiotics mediated by cytochrome P450.[Methods] Enzymic experiment:In vitro suitable reaction conditions soybean lipoxygenase (SLO), substrate (acrylonitrile) and other component react in the enzymic system, and followed by detecting the reaction product in the reaction medium by spectrophotometry. The oxidative rate of acrylonitrile catalyzed by SLO and the inhibition ratio of the GTP for the oxidation are calculated, respectively.Cellular experiment:The effect of acrylonitrile on cell survival rate were assessed by reductions of tetrazolium dye(MTT) in cultured HBE cells. After HBE cells exposure to different concentrations of acrylonitrile for 12 hours, the protein expressions of 5-lipoxygenase in the cells are tested by western-blot, and the DNA damages by the single cell gel electrophoresis. At last, the effect of the specific inhibitor of 5-lipoxygenase(AA-861) on 5-lipoxygenase protein expression and DNA damage in the cells are detected.[Results] SLO can catalyze the co-oxidation of acrylonitrile to generate cyanoethylene oxide in the presence of hydrogen peroxide. GTP can inhibit the oxidation of acrylonitrile by SLO, which appears to concentration-dependent manner in special range. Acrylonitrile can induce 5-lipoxygenase protein expression, but AA-861 can not in HBE. Acrylonitrile causes toxic action and DNA damage in HBE, which can [Conclusions] In the presence of hydrogen peroxide, SLO can mediate acrylonitrile co-oxidation, which can be inhibited by GTP. 5-LOX protein expression in HBE can be regulated by acrylonitrile, and specific inhibitor of 5-LOX (AA-861) has no obvious effect on the expression. The co-oxidation of acrylonitrile by 5-LOX turns into electrophiles that covalently bind to DNA and induce DNA damage, which can be significantly inhibited by AA-861.

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CLC: > Medicine, health > Preventive Medicine,Health > Health - basic science > Biochemistry of Health,Sanitary Chemistry
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