丁酸对TNFα所致肠上皮屏障损伤的保护作用

Butyric Acid Protects Intestinal Epithelial Barrier from Injury Induced by TNFα

目的探讨丁酸对TNFα所致肠上皮屏障损伤的保护作用。方法用CCK-8细胞活力检测探索TNFα对Caco2发挥损伤作用的最佳浓度和时间,并以此为基础探索在TNFα发挥损伤作用最佳作用时间附近丁酸对Caco2细胞的保护情况,随后探索TNFα和丁酸共同作用于Caco2细胞时的最佳时间和浓度,并检测Caco2细胞单层上皮屏障的FITC-dextran渗透率,紧密连接ZO-1和Occludin的mRNA表达情况及免疫荧光观察TNFα和丁酸共同作用后细胞的生长情况及ZO-1和Occludin在Caco2中的表达和分布。结果100 ng/mL的TNFα刺激48 h能明显降低Caco2的细胞存活率(P<0.0001);丁酸作用于Caco2细胞48 h,0.2 mM/L的丁酸能明显提高Caco2的细胞存活率(P<0.0001)。当TNFα和丁酸共同作用时,与TNFα干预组相比,丁酸可以明显降低TNFα所致的肠上皮单层屏障的FITC-dextran渗透率(P<0.0001),提高ZO-1(P<0.01)和Occludin(P<0.01)的表达及稳定其在Caco2细胞中的分布。结论丁酸可以减轻TNFα所致的肠上皮屏障损伤,为进一步明确丁酸在溃疡性结肠炎治疗中作用机制提供实验基础。

Objective To explore the protective effect of butyric acid on intestinal epithelial barrier damage induced by TNFα.Methods CCK-8 cell viability assay was used to determine the optimal concentration and time of TNFαin exerting damaged effects on Caco2 damage,and based on this,the protective effect of butyric acid on Caco2 cells was explored around the optimal time when TNFαexerted damage.Subsequently,the optimal time and concentration of TNFαand butyric acid acting on Caco2 cells were explored,and the FITC-dextran permeability of the Caco2 cell monolayer epithelial barrier,the mRNA expression of ZO-1 and Occludin,and the growth of cells after TNFαand butyric acid co-treatment were detected,as well as the expression and distribution of ZO-1 and Occludin in Caco2 cells were observed by immunofluorescence.Results TNFα,at a concentration of 100 ng/mL significantly decreased the cell viability of Caco2 after 48 hours of stimulation(P<0.0001);butyrate at a concentration of 0.2mM/L significantly increased the cell viability of Caco2 after 48 hours of treatment(P<0.0001).When TNFαand butyrate were co-administered,compared with the TNFαintervention group,butyrate significantly decreased the FITC-dextran permeability of the intestinal epithelial monolayer barrier caused by TNFα(P<0.0001),and increased the expression of ZO-1(P<0.01)and Occludin(P<0.01),and stabilized their distribution in Caco2 cells.Conclusion Butyrate can alleviate TNFα-induced damage to intestinal epithelial barrier,providing experimental basis for further elucidating the mechanism of butyrate in the treatment of ulcerative colitis.