摘要
本试验旨在研究白藜芦醇对脂多糖(LPS)诱导的猪小肠上皮细胞(IPEC-J2细胞)炎性损伤与屏障功能障碍的缓解作用。将IPEC-J2细胞在含有不同浓度的LPS和白藜芦醇的培养基中分别培养18 h后,MTT法检测细胞活性确定LPS造模浓度(10.0 mg/L)和白藜芦醇预处理浓度(5.0、20.0、35.0μmol/L)。分别用上述确定的预处理浓度(5.0、20.0、35.0μmol/L)的白藜芦醇预处理IPEC-J2细胞后进行LPS(10.0 mg/L)诱导,同时设置对照组(不添加LPS与白藜芦醇)与LPS单独刺激组(添加10.0 mg/L LPS),测定细胞活性、细胞迁移率、炎症因子分泌量、Toll样受体4(TLR4)/核转录因子-κB(NF-κB)信号通路及紧密连接相关基因mRNA相对表达量。结果显示:相较于LPS单独刺激组,20.0μmol/L白藜芦醇预处理可以显著缓解由LPS刺激引起的IPEC-J2细胞活性的降低(P<0.05);5.0和20.0μmol/L白藜芦醇预处理组IPEC-J2细胞迁移率较LPS单独刺激组显著提高(P<0.05);5.0、20.0和35.0μmol/L白藜芦醇预处理组细胞上清液中白细胞介素-8(IL-8)、肿瘤坏死因子-α(TNF-α)含量显著低于LPS单独刺激组(P<0.05),白细胞介素-10(IL-10)含量显著高于LPS单独刺激组(P<0.05);5.0和20.0μmol/L白藜芦醇预处理显著缓解了LPS刺激引起的细胞中IL-8、TNF-α、白细胞介素-1β(IL-1β)、TLR4、髓样分化因子88(MyD88)的mRNA相对表达量的升高(P<0.05)以及IL-10与NF-κB抑制蛋白α(IκBα)mRNA相对表达量的降低(P<0.05);5.0和20.0μmol/L白藜芦醇预处理后细胞中封闭蛋白-1(claudin-1)、闭锁蛋白(occludin)和闭锁小带蛋白-1(ZO-1)mRNA相对表达量显著提高(P<0.05),且35.0μmol/L白藜芦醇预处理后细胞中ZO-1 mRNA相对表达量显著提高(P<0.05)。综上所述,白藜芦醇通过提高细胞迁移率、调节炎症细胞因子分泌、抑制TLR4/NF-κB信号通路过度激活,提高紧密连接相关基因表达,缓解LPS诱导的IPEC-J2细胞的炎性损伤和屏障功能障碍。
The aim of this experiment was to investigate the alleviating effect of resveratrol(RES)on lipopolysaccharide(LPS)-induced inflammatory damage and barrier dysfunction in porcine small intestinal epithelial cells(IPEC-J2 cells).IPEC-J2 cells were cultured for 18 h in medium supplemented with different concentrations of LPS and RES,respectively,and the cell activity was determined by MTT assay to determine the LPS modeling concentration(10.0 mg/L)and RES pretreatment concentrations(5.0,20.0 and 35.0μmol/L).The cells were pretreated with the above concentrations(5.0,20.0 and 35.0μmol/L)of RES and then subjected to LPS(10.0 mg/L)induction.A control group(no addition of LPS and RES)and an LPS alone stimulation group(addition of 10.0 mg/L LPS alone)were set up to determine cell activity,cell migration rate,inflammatory cytokine secretion,and the mRNA relative expression levels of TLR4/NF-κB signaling pathway and tight junction related genes.The results showed that,compared with LPS alone stimulation group,20.0μmol/L RES representment significantly alleviated the decrease of IPEC-J2 cell activity induced by LPS stimulation(P<0.05);the migration rate of IPEC-J2 cells was significantly increased in the 5.0 and 20.0μmol/L RES pretreatment groups compared with the LPS alone stimulation group(P<0.05).The interleukin-8(IL-8)and tumor necrosis factor-α(TNF-α)contents were significantly lower and the interleukin-10(IL-10)content was significantly higher in the supernatant of cells in the 5.0,20.0 and 35.0μmol/L RES pretreatment groups than those in the LPS alone stimulation group(P<0.05).Compared with LPS alone stimulation group,5.0 and 20.0μmol/L RSE pretreatment significantly alleviated the increase of the mRNA relative expression levels of IL-8,TNF-α,interleukin-1β(IL-1β),TLR4 and myeloid differentiation factor 88(MyD88)and the decrease of the mRNA relative expression levels of IL-10 and NF-κB inhibitor proteinα(IκBα)in cells induced by LPS stimulation(P<0.05);the mRNA relative expression levels of claudin-1,ocludin and zona occludens-1(ZO-1)were significantly increased after 5.0 and 20.0μmol/L RES pretreatment(P<0.05),and the mRNA relative expression level of ZO-1 significantly increased after 35.0μmol/L RES pretreatment(P<0.05).In conclusion,RES alleviated LPS-induced inflammatory damage and barrier dysfunction in IPEC-J2 cells by increasing cell migration rate,regulating inflammatory factor secretion,curbing excessive activation of TLR4/NF-κB signaling pathway and increasing expression of tight junction-related genes.
作者
张磊正
冯志华
龚建刚
郝艳霜
刘观忠
刘彦慈
马可为
ZHANG Leizheng;FENG Zhihua;GONG Jiangang;HAO Yanshuang;LIU Guanzhong;LIU Yanci;MA Kewei(College of Animal Science and Technology,Hebei Agricultural University,Baoding 071000,China;College of Food Science and Technology,Hebei Agricultural University,Baoding 071000,China;Baoding Vocational and Technical College,Baoding 071000,China)
出处
《动物营养学报》
CAS
CSCD
北大核心
2023年第11期7344-7354,共11页
CHINESE JOURNAL OF ANIMAL NUTRITION
基金
河北省自然科学基金资助项目(C2021204175)
河北省现代农业产业技术体系蛋鸡肉鸡(家禽)营养与饲料(HBCT2023210202)。