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【目的】 研究硒蛋白谷胱甘肽过氧化物酶4（glutathione peroxidases 4,GPX4）失活如何参与调控脂多糖（lipopolysaccharide,LPS）诱导的RAW264.7巨噬细胞炎症反应及其潜在的分子机制。【方法】 体外培养RAW264.7巨噬细胞,以DMSO为对照,使用0.1~5.0 μmol/L GPX4抑制剂FIN56处理,通过CCK-8法检测细胞活力和Western blotting检测GPX4蛋白表达水平,确定抑制剂最适浓度。将RAW264.7巨噬细胞分为4组：对照组,添加DMSO培养24 h;FIN56（GPX4抑制剂）组,添加0.5 μmol/L FIN56培养24 h;DMSO-LPS组,DMSO培养24 h后使用LPS （100 ng/mL）刺激3 h;FIN56-LPS组,FIN56培养24 h后使用LPS刺激3 h。各组细胞经培养后,利用荧光探针2',7'-二氯二氢荧光素二乙酸酯（2',7'-dichlorodi-hydrofluorescein diacetate,H2DCFDA）检测细胞内活性氧（reactive oxygen species,ROS）水平,使用试剂盒法检测细胞内丙二醛（malondialdehyde,MDA）水平,分别使用ELISA和荧光定量PCR检测促炎细胞因子白细胞介素1β（interleukin-1β,IL-1β）、白细胞介素6（interleukin-6,IL-6）、肿瘤坏死因子α（tumor necrosis factor-α,TNF-α）的分泌水平和基因表达量,使用Western blotting法检测Toll样受体4（toll like receptor 4,TLR4）信号通路相关蛋白表达水平。【结果】 与DMSO处理相比,0.5 μmol/L FIN56处理巨噬细胞24 h对细胞活性无显著影响（P>0.05）,且显著降低GPX4蛋白表达水平（P<0.05）,故选用0.5 μmol/L FIN56用于后续实验。与对照组相比,FIN56和LPS均显著增加了ROS积累（P<0.05）,而与DMSO-LPS组相比,FIN56-LPS组ROS水平显著升高（P<0.05）。与对照组相比,LPS可显著增加小鼠巨噬细胞IL-1β、IL-6和TNF-α的mRNA表达量和IL-6含量,以及c-Jun氨基末端蛋白激酶（c-Jun N-terminal protein kinase,JNK）和c-Jun磷酸化水平（P<0.05）,而FIN56可显著下调LPS诱导的IL-6的mRNA表达量和含量及JNK和c-Jun磷酸化水平（P<0.05）,但对p38蛋白（p38 MAPK,p38）、细胞外调节蛋白激酶（phosphorylate extracellular regulated protein kinases1/2,Erk1/2）蛋白表达水平无显著影响（P>0.05）。【结论】 GPX4失活可有效阻断JNK和c-Jun磷酸化,从而特异性抑制LPS诱导的巨噬细胞的炎症反应。该结果可为以GPX4为靶点研发抗炎治疗策略提供理论依据。 相似文献
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低氧胁迫对河川沙塘鳢抗氧化酶及ATP酶活性的影响 总被引：1，自引：0，他引：1
研究了在急性低氧1.5 h、5 h和慢性低氧3 d下,河川沙塘鳢(Odontobutis potamophila)5种组织(心、脑、肝、鳃和肾)的超氧化物歧化酶(SOD)、过氧化氢酶(CAT)及谷胱甘肽过氧化物酶(GPX)3种抗氧化酶和ATP酶活性的变化规律。结果显示:在急性低氧暴露1.5 h时,河川沙塘鳢SOD和GPX的活力在各组织中与对照组相比均无显著差异,CAT活力在心、鳃和肝3种组织呈现显著升高(P0.05),ATP酶活力在心和肝组织极显著升高(P0.01);在急性低氧暴露5 h时,除肝组织SOD酶活性显著降低外(P0.05),其它4种组织的CAT、GPX和ATP酶均不同程度显著升高(P0.05);在慢性低氧处理3 d时,心、脑组织的抗氧化酶已基本恢复至与对照组无显著差异的水平,但鳃、肝和肾中酶活力仍较高(P0.05)。研究表明,河川沙塘鳢能通过自身调节抗氧化酶及ATP酶活性,改变代谢底物,提高机体适应低氧环境的能力。 相似文献
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[Objective] Using molecular biotechnology to clone the sus scrofa GPX2 gene.[Method] Using total RNA of sus scrofa duodenum as template,degenerated primer pairs were designed according to the homology alignment analysis of GPX2 gene of human,rat,mouse,dog and cattle.A sus scrofa GPX2 gene sequence of 330 bp was obtained by RT-PCR application method.Primes were designed respectively according to the known sequence,sus scrofa GPX2 gene was isolated and cloned by 3-RACE and 5-RACE method and analyzed the gene sequence.[Result] A mRNA sequence of 924 bp was successfully cloned and isolated in this research.This sequence contained complete 3' end and had higher sequence homology with human,mouse,cattle and dog GPX2 gene,and there was codon called TGA which encoding Sec on the position of No.114-116 gene.[Conclusion] Sequence alignment analysis showed that the cloned gene was sus scrofa GPX2 gene(NCBI GenBank database,the sequence number was DQ98982). 相似文献
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[目的]检测太湖猪核心保种群体中雌激素受体基因(ESR)、促卵泡素β亚基基因(FSHβ)和谷胱甘肽过氧化物5基因(GPX5)的多态性。[方法]利用PCR和PCR-RFLP方法对太湖猪核心保种群体中ESR、FSHβ、GPX5基因进行多态性检测。[结果]ESR和GPX5基因在太湖猪群体中均检测到AA、AB和BB 3种基因型,FSHβ亚基基因只检测到BB和AB 2种基因型。ESR基因以BB基因型为优势基因型,等位基因B为优势基因。FSHβ基因中等位基因B的频率远大于等位基因A。GPX5基因的基因型频率呈ABBBAA趋势。ESR和FSHβ基因为低度多态,GPX5基因处于中度多态。[结论]经χ2适合性检验表明,ESR、FSHβ、GPX5基因均处于Hardy-Weinberg平衡状态(P0.05)。 相似文献
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This study was conducted to investigate the effect of vitamin A and E supplementation on the antioxidant defences of broiler chickens against carbon tetrachloride (CCl(4))-induced oxidative stress at 4 weeks of age. Superoxide dismutase (SOD) and glutathione peroxidase (GPX) activities as well as total antioxidant (TAO) level were analysed before and after CCl(4) challenge. Day-old Lohman broiler chickens (n = 144) were randomly assigned to six factorially arranged dietary treatments consisting of vitamin A [1.35 (control) or +20 mg/kg] and vitamin E [20 (control), +40 or +60 mg/kg]. The background of vitamins A and E in the basal diet was 4500 IU (1.35 mg) and 30 IU (20 mg) respectively. At 4 weeks of age, eight chickens from each treatment were bled before interperitoneal injection with 1 ml of CCl(4) (mixed with olive oil in a ratio of 1:1) and bled again 24 h post-injection. Vitamin E supplementation decreased (p < 0.05) the activity of both SOD and GPX and showed a tendency (p = 0.07) for TAO reduction. CCl(4) attenuated SOD and GPX activities as well as TAO level. The decrease was profound (p < 0.05) in chickens fed the basal diet as well as those fed basal diet supplemented with 20 mg vitamin A. TAO levels behaved similarly when chickens were challenged with CCl(4). After CCl(4) injection, SOD activities of all experimental groups were equivalent. The presence of vitamin A decreased (p < 0.05) plasma GPX activity in chickens fed the basal diet supplemented with 40 mg/kg of vitamin E. Results of this experiment suggested that vitamin E supplementation elevated antioxidant enzyme activities while vitamin A supplementation attenuated this effect. Vitamin E supplementation improved the total reducing power by maintaining comparable levels of TAO upon CCl(4) challenge. Further experiments need to be carried out to investigate the role of vitamin A in oxidative stress and to evaluate the lipid peroxidation products. 相似文献
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奶牛胎衣不下与血浆GPX、CAT活性的相关研究 总被引：1，自引：0，他引：1
为了阐明奶牛分娩前抗氧化系统变化规律与胎衣不下的关系,以荷斯坦奶牛为研究对象,筛选165头妊娠期各项指标正常的奶牛,于分娩前1周静脉采血及测定血浆中谷胱甘肽过氧化物酶（GPX）、过氧化氢酶（CAT）含量,并按照胎衣不下的标准（产后胎衣12h内不能正常排出）进行分组对比分析。结果表明：筛选的165头奶牛有19头在产后表现为胎衣不下,发病率为11.5%;胎衣不下奶牛血浆GPX活性（104.00±80.19U/mL）显著低于正常奶牛（134.82±71.34U/mL,P〈0.05）,CAT活性（1.44±1.08U/mL）也低于正常奶牛（1.63±1.74U/mL,P〉0.05）。说明奶牛血浆中GPX、CAT活性与产后胎衣不下存在必然联系。 相似文献
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猪GPX5,FUT1和PRLR基因的遗传多态性及遗传效应分析 总被引：2，自引：1，他引：1
为了分析谷胱甘肽过氧化物酶5基因(Glutathion Peroxidase 5,GPX5),岩藻糖转移酶1基因(α1-Fucosytransferase,FUT1)和催乳素受体基因(Prolactin receptor,PRLR)在野猪与大白猪杂交的F1代群体中的遗传多态性及遗传效应。采用PCR-RFLP方法检测了GPX5,FUT1和PRLR基因的多态性,分析其对初生重(LiB)和30日龄体重(Li30)的遗传效应。结果表明,除了PRLR基因外,其余2个基因的基因频率与大白猪已有研究结果基本一致;PRLR基因的AA型个体在Li30上比AB和BB型个体可显著增重1.42～1.47 kg(P0.05);GPX5和FUT1基因对这2个性状的效应均不显著。结果提示,野猪血统的引入没有显著改变现有猪群的遗传基础,PRLR基因可在生产中考虑应用。 相似文献