丙酸钠对IPEC-J2细胞紧密连接及炎症细胞因子的影响

Effects of sodium propionate on the tight junction and inflammatory cytokines in IPEC-J2 cells

[目的]本文以猪肠道上皮IPEC-J2细胞为模型,探究不同水平丙酸钠对细胞紧密连接和炎症细胞因子的影响。[方法]分别用1和10 mmol·L^(-1)丙酸钠处理IPEC-J2细胞,以不加丙酸钠处理IPEC-J2细胞为对照组,分别处理12 h和24 h后测定各组细胞活力、跨膜电阻(TEER)值、紧密连接相关蛋白及炎症细胞因子基因表达和细胞上清液中炎症因子浓度。[结果]丙酸钠处理24 h后,处理组细胞活力显著下降(P<0.05),细胞的TEER值则显著上升(P<0.05)。RT-qPCR结果显示,紧密连接相关蛋白Claudin中,不同浓度丙酸钠处理12和24 h后,显著下调Claudin-1 mRNA的表达(P<0.05),但显著上调Claudin-2、Claudin-3和Claudin-4 mRNA的表达(P<0.05)。紧密连接相关蛋白ZO中,处理12和24 h时,1和10 mmol·L^(-1)丙酸钠分别显著上调ZO-2 mRNA和ZO-1 mRNA的表达(P<0.05),但10 mmol·L^(-1)丙酸钠则显著下调ZO-2 mRNA的表达(P<0.05)。不同浓度丙酸钠处理12和24 h均显著上调炎症细胞因子IL-8、IL-18和TNF-αmRNA的表达(P<0.05),显著下调IL-6 mRNA的表达(P<0.05)。ELISA测定结果表明,丙酸钠显著刺激IL-8和TNF-α的分泌(P<0.05),对IL-18和IL-6的分泌则无显著影响(P>0.05)。[结论]丙酸钠选择性上调紧密连接蛋白家族相关基因的表达,一定程度上维护小肠上皮细胞屏障功能的完整性,同时调节细胞炎症反应,揭示丙酸在调节小肠上皮细胞屏障功能以及炎症反应过程中发挥重要作用。

[Objectives] This study aims to investigate the effects of sodium propionate on the expression of tight junction and inflammatory cytokines in IPEC-J2 cells.[Methods] IPEC-J2 cells were treated by different doses of sodium propionate(1 and 10mmol·L^-1 , 0mmol·L^-1 as control group),and the tight junction permeability and expression of tight junction complex and inflammatory cytokines were measured by trans-epithelial electrical resistance(TEER),RT-qPCR and enzyme linked immunosorbent assay(ELISA),respectively.[Results] Compared with the control group,sodium propionate dose-dependently increased the TEER of IPEC-J2 cells after being incubated for 12 h and 24 h( P <0.05). The expression of tight junction complex Claudin- 2,Claudin-3 and Claudin- 4 were significantly up-regulated by different levels of sodium propionate( P <0.05),whereas the expression of Claudin- 1 was down-regulated( P < 0.05 ). In addition,1mmol·L^-1 and 10mmol·L^-1 sodium propionate up-regulated the expression of ZO- 2 and ZO- 1 ,respectively( P <0.05 ). However, ZO- 2 expression decreased by 10mmol·L^-1 sodium propionate( P <0.05). The mRNA expression results of inflammatory cytokines showed that the sodium propionate administration significantly up-regulated the expression of pro-inflammatory cytokines except IL- 6 (P <0.05). Moreover,ELISA results showed that sodium propionate significantly stimulated the release of IL-8 and TNF-α in IPEC-J2 cells( P <0.05),but had no effects on the concentration of IL-6 and IL^-18( P >0.05).[Conclusions] Taken together,this study indicates that sodium propionate selectively up-regulates the mRNA expression of tight junction related protein,partly maintains the intestinal epithelial barrier function,and regulates inflammation process,reveals the role of propionate in the regulation of small intestinal epithelial barrier function and inflammatory reaction.