白细胞介素1受体拮抗剂对二硝基氯苯诱导的奶牛乳腺上皮细胞氧化损伤的保护作用

郭咏梅; 闫素梅; 史彬林

doi:10.3969/j.issn.1006-267x.2019.10.030

动物营养学报 >

2019 , Vol. 31 >Issue 10: 4659 - 4666

DOI: https://doi.org/10.3969/j.issn.1006-267x.2019.10.030

分子营养 Molecular Nutrition

白细胞介素1受体拮抗剂对二硝基氯苯诱导的奶牛乳腺上皮细胞氧化损伤的保护作用

郭咏梅 ,
闫素梅 ,
史彬林

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内蒙古农业大学动物科学学院, 内蒙古自治区高校动物营养与饲料科学重点实验室, 呼和浩特 010018

郭咏梅(1989-),女,内蒙古呼和浩特人,博士研究生,从事动物矿物质与维生素营养研究。E-mail:ymguo2015@163.com

收稿日期: 2019-03-29

网络出版日期: 2019-10-17

基金资助

国家自然科学基金项目（31560650）

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Protective Effect of Interleukin-1 Receptor Antagonist on Oxidative Damage Induced by 2,4-Dinitrochlorobenzene in Bovine Mammary Epithelial Cells

GUO Yongmei ,
YAN Sumei ,
SHI Binlin

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Inner Mongolia Key Laboratory of Animal Nutrition and Feed Science, College of Animal Science, Inner Mongolia Agricultural University, Hohhot 010018, China

Received date: 2019-03-29

Online published: 2019-10-17

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摘要

本试验旨在探究白细胞介素-1受体拮抗剂（IL-1Ra）对二硝基氯苯（DNCB）诱导的奶牛乳腺上皮细胞（BMEC）氧化损伤的保护作用，并确定抑制白细胞介素-1（IL-1）活性的最佳IL-1Ra浓度。采用完全随机试验设计，将第3代BMEC随机分为12个组，每组6个重复。1组为对照组，利用不含DNCB及IL-1Ra的培养液培养6 h；2~6组是不同剂量的IL-1Ra处理组，分别利用含有1、5、10、20、50 μg/L的IL-1Ra培养液培养6 h；7组为DNCB损伤组，利用含有300 μmol/L的DNCB培养液培养6 h；8~12组是不同剂量的IL-1Ra保护组，分别利用含有1、5、10、20、50 μg/L的IL-1Ra与300 μmol/L DNCB培养液培养6 h。结果表明：与对照组相比，经300 μmol/L的DNCB作用6 h后，BMEC的相对增殖率（RGR）以及含硒蛋白[硫氧还蛋白还原酶（TrxR）、谷胱甘肽过氧化物酶（GPx）]、抗氧化酶[超氧化物歧化酶（SOD）、过氧化氢酶（CAT）]活性与总抗氧化能力（T-AOC）均显著降低（P<0.05）；同时，诱导型一氧化氮合酶（iNOS）与活性氧（ROS）活性以及一氧化氮（NO）、IL-1、白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）、丙二醛（MDA）含量则呈现相反的变化规律，均显著升高（P<0.05）。300 μmol/L DNCB与不同浓度的IL-1Ra同时作用6 h后，BMEC的RGR及抗氧化酶活性随IL-1Ra浓度的增加均呈现升高趋势，其中5、10、20、50 μg/L IL-1Ra保护组的上述指标均显著高于DNCB损伤组（P<0.05），并且RGR及CAT、GPx、TrxR活性均以IL-1Ra浓度为10 μg/L时最高；同时，IL-1、IL-6、TNF-α与NO含量及iNOS活性则随IL-1Ra浓度增加呈现先降低后升高的趋势，其中5、10、20、50 μg/L IL-1Ra保护组的上述指标均显著低于DNCB损伤组（P<0.05），以IL-1Ra浓度为5、10、20 μg/L时效果较好。然而，正常的BMEC经过不同浓度的IL-1Ra处理后，上述指标（5 μg/L IL-1Ra处理组的TNF-α含量除外）均未发生显著变化（P>0.05）。综上所述，IL-1Ra竞争性与IL-1R结合，抑制了IL-1的活性，进而抑制了DNCB引起的BMEC氧化损伤及炎症反应，本试验条件下以10 μg/L为适宜浓度。

关键词： 白细胞介素-1受体拮抗剂; 奶牛乳腺上皮细胞; 炎症因子; 氧化损伤

本文引用格式

郭咏梅 , 闫素梅 , 史彬林 . 白细胞介素1受体拮抗剂对二硝基氯苯诱导的奶牛乳腺上皮细胞氧化损伤的保护作用[J]. 动物营养学报, 2019 , 31(10) : 4659 -4666 . DOI: 10.3969/j.issn.1006-267x.2019.10.030

Abstract

This experiment was conducted to investigate the protective effect of interleukin-1 receptor antagonist (IL-1Ra) on oxidative damage induced by 2,4-dinitrochlorobenzene (DNCB) in bovine mammary epithelial cells (BMEC), and determine to the suitable concentration of IL-1Ra inhibiting interleukin-1 (IL-1) activity. A completely randomized trial design was used in this experiment. The third-generation BMEC was randomly divided into 12 groups with 6 replicates per group. BMEC in group 1 (control group) was treated by culture solution with DNCB and IL-1Ra for 6 h; BMEC in groups 2 to 6 (IL-1Ra treatment groups) was treated by culture solution with 1, 5, 10, 20 and 50 μg/L IL-1Ra for 6 h, respectively; BMEC in group 7 (DNCB damage group) was treated by culture solution with 300 μmol/L DNCB for 6 h; BMEC in groups 8 to 12 (IL-1Ra protection groups) was treated by culture solution with 1, 5, 10, 20 and 50 μg/L IL-1Ra+300 μmol/L DNCB for 6 h, respectively. The results showed that compared with the control group, the relative growth rate (RGR) and the activities of selenoprotein[thioredoxin reductase (TrxR) and glutathione peroxidase (GPx)] plus antioxidase[superoxide dismutase (SOD), catalase (CAT)] and total antioxidant capacity (T-AOC)] were significantly reduced after BMEC was exposed to 300 μmol/L DNCB for 6 h (P<0.05). However, the activities of nitric oxide synthase (iNOS) and reactive oxygen species (ROS) as well as the contents of nitric oxide (NO), IL-1, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and malondialdehyde (MDA) showed opposite changes, they were significantly increased (P<0.05). BMEC was simultaneously exposed by 300 μmol/L DNCB and different concentrations of IL-1Ra for 6 h, the RGR plus antioxidase activities of BMEC were enhanced with the increase of IL-1Ra concentration, and the above indexes in 5, 10, 20 and 50 μg/L IL-1Ra protection groups were significantly higher than those in DNCB damage group (P<0.05), with the highest values of RGR as well as the activities of CAT, GPx and TrxR were found at 10 μg/L IL-Ra. Oppositely, IL-1, IL-6, TNF-α and NO contents and iNOS activity were rose up first and then went down with the increase of IL-1Ra concentration, and the above indexes in 5, 10, 20 and 50 μg/L IL-1Ra protection groups were significantly lower than those in DNCB damage group (P<0.05), with better protective effects of IL-1Ra at 5, 10 and 20 μg/L. BMEC was alone exposed by different concentrations of IL-1Ra had no significant effects on the indexes mentioned above with an exception of TNF-α content in 5 μg/L IL-1Ra treatment group (P>0.05). In summary, IL-1Ra can competitively bind with IL-1R to inhabit the activity of IL-1, and then the oxidative damage and inflammatory response of DNCB were suppressed. Under this experimental condition, the suitable concentration of IL-1Ra inhibiting IL-1 activity is 10 μg/L.

Key words： interleukin-1 receptor antagonist; bovine mammary epithelial cells; inflammation parameter; oxidative damage

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