低聚壳聚糖硒抵抗玉米赤霉烯酮诱导猪肠上皮细胞氧化损伤和内质网应激的作用研究

Effects of Oligomeric Chitosan Selenium on Oxidative Damage and Endoplasmic Reticulum Stress in Zearalenon-Induced Pig Intestinal Epithelial Cells

本试验旨在探究低聚壳聚糖硒拮抗玉米赤霉烯酮(ZEN)对猪肠上皮细胞(IPEC-J2细胞)损伤的影响。试验设定为:对照组(C组)、ZEN组(30μg/mL ZEN)、ZEN+0.5 Se组(30μg/mL ZEN+0.5μmol/L低聚壳聚糖硒,以Se计)、ZEN+1.5 Se组(30μg/mL ZEN+1.5μmol/L低聚壳聚糖硒,以Se计)、ZEN+3.0 Se组(30μg/mL ZEN+3.0μmol/L低聚壳聚糖硒,以Se计)。测定细胞存活率、乳酸脱氢酶(LDH)活性、活性氧(ROS)含量、线粒体膜电位变化、氧化应激和内质网应激(ERS)相关蛋白的表达。结果表明:与对照组相比,添加ZEN显著增加LDH活性(P<0.05),极显著增加ROS含量(P<0.01),极显著降低线粒体膜电位(P<0.01),显著降低细胞存活率(P<0.05);添加低聚壳聚糖硒可显著降低LDH活性(P<0.05),极显著降低ROS含量(P<0.01),显著增加线粒体膜电位(P<0.05),极显著增加细胞存活率(P<0.01);相比于对照组,添加ZEN显著降低转录因子E2相关因子(Nrf2)蛋白表达量(P<0.05),添加低聚壳聚糖硒极显著降低Nrf2蛋白表达量(P<0.01);相比于对照组,ZEN和ZEN+0.5 Se组血红素加氧酶-1(HO-1)蛋白表达量显著降低(P<0.05),ZEN+0.5 Se、ZEN+1.5 Se、ZEN+3.0 Se组与ZEN组无显著差异(P>0.05),但ZEN+1.5 Se、ZEN+3.0 Se组与对照组也无显著差异(P>0.05);相比于对照组,添加ZEN显著或极显著增加葡萄糖调节蛋白78(GRP78)(P<0.05)、磷酸化蛋白激酶R样内质网激酶/蛋白激酶R样内质网激酶(P-PERK/PERK)(P<0.05)、转录激活因子4(ATF4)(P<0.05)、C/EBP同源蛋白(CHOP)(P<0.01)蛋白表达量;添加低聚壳聚糖硒可显著或极显著降低GRP78(P<0.05)、P-PERK/PERK(P<0.01)、ATF4(P<0.01)、CHOP(P<0.05)蛋白表达量。综上所述,低聚壳聚糖硒可缓解ZEN引起的IPEC-J2细胞氧化损伤和内质网应激。

This study was to investigate the antagonistic effects of oligomeric chitosan selenium on zearalenon(ZEN)on injury of pig intestinal epithelial cells(IPEC-J2 cells).The experiment set the control group(C group),ZEN group(30μg/mL),ZEN+0.5 Se group(30μg/mL ZEN+0.5μmol/L oligomeric chitosan selenium,calculated as Se),ZEN+1.5 Se group(30μg/mL ZEN+1.5μmol/L oligomeric chitosan selenium,calculated as Se),ZEN+3 Se group(30μg/mL ZEN+3μmol/L oligomeric chitosan selenium,calculated as Se).The survival rate,lactate dehydrogenase(LDH)activity,reactive oxygen species(ROS)content,changes in mitochondrial membrane potential,protein expression related to oxidative stress and endoplasmic reticulum stress were measured.The results showed that compared with control group,adding ZEN significantly increased LDH activity(P<0.05)and extremely significantly increased ROS content(P<0.01),and extremely significantly decreased mitochondrial membrane potential(P<0.01)and significantly decreased survival rate(P<0.05);the addition of oligomeric chitosan selenium were significantly reduced LDH activity(P<0.05),extremely significantly reduced ROS content(P<0.01),and significantly increased mitochondrial membrane potential(P<0.05)and extremely significantly increased survival rate(P<0.01).Compared with the control group,adding ZEN significantly reduced transcription factor E2 related factors(Nrf2)protein expression(P<0.05),Nrf2 protein expression was significantly alleviated by the addition of oligomeric chitosan selenium(P<0.01);compared with the control group,haem oxygenase-1(HO-1)protein expression in the ZEN group and ZEN+0.5 Se group were significantly lower(P<0.05),there was no significant difference among ZEN+0.5 Se,ZEN+1.5 Se,ZEN+3.0 Se and ZEN groups(P>0.05),however,there was no significant difference among ZEN+1.5 Se,ZEN+3.0 Se and control group(P>0.05).Compared with the control group,adding ZEN significantly increased glucose-regulatory protein(GRP78)(P<0.05),endoplasmic reticulum regulated kinase(P-PERK/PERK)(P<0.05),transcriptional activator 4(ATF4)(P<0.05),C/EBP homologous protein(CHOP)(P<0.01)protein expression;the supplementation of oligomeric chitosan selenium significantly decreased GRP78(P<0.05),P-PERK/PERK(P<0.01),ATF4(P<0.01),CHOP(P<0.05)protein expression.In conclusion,oligomeric chitosan selenium can alleviate oxidative damage and endoplasmic reticulum stress of IPEC-J2 cells induced by ZEN.