Full Text:   <2289>

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CLC number: R392.11

On-line Access: 2017-09-05

Received: 2016-05-03

Revision Accepted: 2016-10-07

Crosschecked: 2017-08-25

Cited: 0

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Citations:  Bibtex RefMan EndNote GB/T7714

 ORCID:

Yan-ting Gu

http://orcid.org/0000-0002-9129-3835

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Journal of Zhejiang University SCIENCE B 2017 Vol.18 No.9 P.770-777

http://doi.org/10.1631/jzus.B1600123


Protective effect of dihydropteridine reductase against oxidative stress is abolished with A278C mutation


Author(s):  Yan-ting Gu, Yan-chun Wang, Hao-jun Zhang, Ting-ting Zhao, Si-fan Sun, Hua Wang, Bin Zhu, Ping Li

Affiliation(s):  Beijing Key Lab for Immune-Mediated Inflammatory Diseases, Institute of Clinical Medical Sciences, China-Japan Friendship Hospital, Beijing 100029, China; more

Corresponding email(s):   lp8675@163.com

Key Words:  Dihydropteridine reductase, Transforming growth factor β, 1 (TGF-β, 1), Nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4), Superoxide dismutase 1 (SOD1), Glutathione peroxidase 3 (GPX3), Oxidative stress


Yan-ting Gu, Yan-chun Wang, Hao-jun Zhang, Ting-ting Zhao, Si-fan Sun, Hua Wang, Bin Zhu, Ping Li. Protective effect of dihydropteridine reductase against oxidative stress is abolished with A278C mutation[J]. Journal of Zhejiang University Science B, 2017, 18(9): 770-777.

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author="Yan-ting Gu, Yan-chun Wang, Hao-jun Zhang, Ting-ting Zhao, Si-fan Sun, Hua Wang, Bin Zhu, Ping Li",
journal="Journal of Zhejiang University Science B",
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pages="770-777",
year="2017",
publisher="Zhejiang University Press & Springer",
doi="10.1631/jzus.B1600123"
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%T Protective effect of dihydropteridine reductase against oxidative stress is abolished with A278C mutation
%A Yan-ting Gu
%A Yan-chun Wang
%A Hao-jun Zhang
%A Ting-ting Zhao
%A Si-fan Sun
%A Hua Wang
%A Bin Zhu
%A Ping Li
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A1 - Yan-ting Gu
A1 - Yan-chun Wang
A1 - Hao-jun Zhang
A1 - Ting-ting Zhao
A1 - Si-fan Sun
A1 - Hua Wang
A1 - Bin Zhu
A1 - Ping Li
J0 - Journal of Zhejiang University Science B
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DOI - 10.1631/jzus.B1600123


Abstract: 
Objective: To evaluate the antioxidation of dihydrobiopterin reductase and to explore the effect of A278C mutation of the quinoid dihydropteridine reductase (QDPR) gene on its antioxidant activity. Methods: First, plasmids with different genes (wild and mutant QDPR) were constructed. After gene sequencing, they were transfected into human kidney cells (HEK293T). Then, the intracellular production of reactive oxygen species (ROS) and tetrahydrobiopterin (BH4) was detected after cells were harvested. Activations of nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4), glutathione peroxidase 3 (GPX3), and superoxide dismutase 1 (SOD1) were analyzed to observe the oxidative stress after transfection. The expression of the neuronal nitric oxide synthase (nNOS) gene was analyzed by semiquantitative reverse-transcription polymerase chain reaction (RT-PCR). We also detected the activation of transforming growth factor β;1 (TGF-β;1) by enzyme-linked immunosorbent assay (ELISA) to observe the connection of TGF-β1 and oxidative stress. Results: The exogenous wild-type QDPR significantly decreased the expression of nNOS, NOX4, and TGF-β1 and induced the expression of SOD1 and GPX3, but the mutated QDPR lost this function and resulted in excessive ROS production. Our data also suggested that the influence on the level of BH4 had no significant difference between mutated and the wild-type QDPR transfection. Conclusions: Wild-type QDPR played an important role in protecting against oxidative stress, but mutant QDPR failed to have these beneficial effects.

A278C位点突变减弱了二氢生物蝶呤还原酶的抗氧化作用

目的:评估二氢生物蝶呤还原酶(QDPR)的抗氧化作用,并初步探讨QDPR基因A278C位点突变对其抗氧化作用的影响。
创新点:首次在体外实验中发现QDPR有抗氧化作用,且此作用在A278C位点突变后减弱。
方法:我们构建了野生型和突变型QDPR质粒,且分别转染至人胚肾293细胞中(HEK293T)。实验可分为以下三组:空白质粒对照组、野生型QDPR组和突变型QDPR组。三天后收集细胞观察活性氧(ROS)和四氢生物蝶呤(BH4)的表达量,使用免疫印迹的方法检测烟酰胺腺嘌呤二核苷酸磷酸氧化酶4(NOX4)、谷胱甘肽过氧化物酶3(GPX3)和超氧化物歧化酶1(SOD1)的蛋白表达水平。用半定量逆转录-聚合酶链反应(RT-PCR)方法分析神经型一氧化氮合成酶(nNOS)基因的表达。用酶联免疫吸附测定(ELISA)试剂盒检测转化生长因子-β1(TGF-β1)的活性。
结论:本实验中野生型QDPR可以显著降低nNOS、NOX4和TGF-β1的水平,同时提高SOD1和GPX3表达。但当QDPR发生位点突变后没有观察到上述现象,并且突变型会导致ROS过量产生。我们的数据还表明,野生型和突变型QDPR对BH4含量的影响无显著差异。综上所述,QDPR有抗氧化作用,但A278C位点突变后会影响QDPR的抗氧化功能。

关键词:二氢生物蝶呤还原酶(QDPR);转化生长因子-β1(TGF-β1);烟酰胺腺嘌呤二核苷酸磷酸氧化酶4(NOX4);超氧化物歧化酶1(SOD1);谷胱甘肽过氧化物酶3(GPX3);氧化应激

Darkslateblue:Affiliate; Royal Blue:Author; Turquoise:Article

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