猴头菇多糖对氧化应激的IPEC-J2细胞抗氧化能力及紧密连接蛋白ZO-1的影响

发布时间：2018-04-18 17:46

本文选题：猴头菇多糖 + 过氧化氢　；参考：《畜牧兽医学报》2017年09期

【摘要】：旨在探究在H_2O_2诱导的氧化应激损伤状态下,猴头菇多糖(HEPs)对IPEC-J2细胞的保护作用。将IPEC-J2细胞分为5组:空白组、模型组、HEPs低剂量组、HEPs中剂量组、HEPs高剂量组。采用MTT法分别选取试验用的H_2O_2和HEPs的最佳剂量;利用DCFH-DA染色观察并测定细胞内活性氧簇(ROS)的含量;通过比色法检测培养细胞上清液中丙二醛(MDA)的含量,细胞中超氧化物歧化酶(SOD)和过氧化氢酶(CAT)的活力以及乳酸脱氢酶(LDH)的释放率;应用实时荧光定量PCR(qRT-PCR)检测紧密连接蛋白ZO-1基因的转录水平。结果显示,(1)构建IPEC-J2细胞氧化应激模型的H_2O_2浓度为0.4mmol·L-1。(2)H_2O_2能够极显著地增加细胞内的ROS含量,诱导细胞凋亡,同时极显著地降低了SOD和CAT的活力,极显著地升高MDA含量与LDH释放率。但是,与模型组相比,经过HEPs预处理后,细胞内ROS的含量极显著地降低(P0.01);同时极显著地提高了SOD和CAT的活力(P0.01),极显著地降低了MDA含量与LDH释放率(P0.01)。(3)实时荧光定量PCR结果显示,H_2O_2能够极显著地降低ZO-1基因的转录量(P0.01),与模型组比较,HEPs能够极显著的提高ZO-1基因的转录量(P0.01);Western blot结果显示ZO-1蛋白的表达量与ZO-1基因类似。可见,猴头菇多糖对H_2O_2诱导氧化损伤状态下的IPEC-J2细胞具有保护作用。
[Abstract]:To investigate the protective effect of hericium Erinaceus polysaccharides (Hep) on IPEC-J2 cells under oxidative stress induced by H_2O_2.IPEC-J2 cells were divided into five groups: blank group, model group, low dose group, middle dose group and high dose group.MTT method was used to select the best dosage of H_2O_2 and HEPs, DCFH-DA staining was used to observe and measure the content of reactive oxygen species (Ros) in cells, and the content of malondialdehyde (MDA) in supernatant of cultured cells was detected by colorimetric method.The activity of superoxide dismutase (SOD) and catalase (catalase) and the release rate of lactate dehydrogenase (LDH) were determined by real-time fluorescence quantitative analysis of ZO-1 gene transcription level.The results showed that the H_2O_2 concentration of 0.4mmol L-1.(2)H_2O_2 in the oxidative stress model of IPEC-J2 cells was 0.4mmol L-1.(2)H_2O_2, which could significantly increase the ROS content, induce apoptosis, decrease the activity of SOD and CAT, and significantly increase the content of MDA and the release rate of LDH.However, compared with the model group, after HEPs pretreatment,The results of real-time fluorescence quantitative PCR analysis showed that H2O-2 could significantly decrease the activity of ZO-1 gene, and increase the activity of SOD and CAT, and decrease the content of MDA and the release rate of LDH.Compared with the model group, Hep could significantly increase the transcription quantity of ZO-1 gene. The results of Western blot showed that the expression of ZO-1 protein was similar to that of ZO-1 gene.Therefore, hericium Erinaceus polysaccharides can protect IPEC-J2 cells from oxidative damage induced by H_2O_2.
【作者单位】：福建农林大学福建省兽医中药与动物保健重点实验室;中西兽医结合与动物保健福建省高等学校重点实验室;
【基金】：国家自然科学基金(31502110)
【分类号】：S853.7

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