姜酮缓解葡聚糖硫酸钠盐诱导小鼠结肠炎的作用机制研究

Mechanism of zingerone alleviating colitis induced by dextran sodium sulfate in mice

目的探究姜酮(zingerone,Zin)对葡聚糖硫酸钠盐(dextran sodium sulfate,DSS)诱导小鼠结肠炎的治疗作用及其潜在免疫调节机制。方法将SPF级6~8周龄C57BL/6雄性小鼠分为对照组(Water组)、模型组(DSS组)、姜酮干预组(DSS+Zin组,20 mg/kg),每组5只。利用DSS溶液建立急性溃疡性结肠炎模型,比较各组小鼠体质量变化、疾病活动指数(disease activity index,DAI)、结肠长度变化及组织病理学评分。RT-qPCR和多因子试剂盒检测小鼠结肠组织中炎症因子mRNA和蛋白质的表达。流式细胞术检测姜酮对体内外T细胞亚群及转录因子RORγt表达的影响。双荧光素酶报告基因系统分析姜酮对RORγt的转录活性及Rorc启动子活性的影响。结果与对照组小鼠比较,DSS组小鼠结肠长度缩短,DAI评分及组织病理学评分升高(P<0.05),IL-17A、IL-21、IL-22、IL-6、IL-1β、IFN-γmRNA表达水平上调(P<0.05),TGF-βmRNA表达水平降低(P<0.05),IL-17A、IL-17F、IL-22、IFN-γ蛋白表达增加(P<0.05);姜酮处理后可显著减轻上述临床症状,并逆转上述炎症因子的表达(P<0.05)。流式细胞术结果显示:姜酮在体内外均可抑制Th17细胞分化及RORγt表达、促进Treg细胞分化(P<0.05),但Th1亚群的分化不受影响。双荧光素酶基因结果显示姜酮处理后的RORγt的转录活性及Rorc启动子活性均被显著抑制(P<0.05)。结论姜酮可通过调Th17/Treg细胞平衡,抑制RORγt表达,从而发挥治疗小鼠溃疡性结肠炎作用。

ObjectiveTo investigate the efficacy of zingerone(Zin)on dextran sodium sulfate(DSS)-induced colitis in mice and its potential immunomodulatory mechanism.MethodsSPF C57BL/6 male mice aged 6 to 8 weeks were divided into control group(Water),model group(DSS),and Zin intervention group(DSS+Zin),with 5 mice in each group.Acute ulcerative colitis model was constructed using DSS solution,and the changes in body weight,disease activity index(DAI),colon length,and histopathological scores were compared among the groups.RT-qPCR and corresponding reagents and kits were employed to detect the mRNA and protein expression of inflammatory cytokines in the colonic tissue,respectively.The effects of Zin on T cell subsets and on the expression of transcription factor RORγt were determined by flow cytometry in vitro and in vivo.Moreover,the transcriptional activity of RORγt and activity of Rorc promoter were examined using dual-luciferase reporter system.ResultsAs compared with the control group,the mice in the DSS group showed shorter colon length,higher scores of DAI and histopathological index(P<0.05),with increased mRNA levels of IL-17A,IL-21,IL-22,IL-6,IL-1βand IFN-γ(P<0.05),while decreased mRNA level of TGF-β(P<0.05).The protein levels of IL-17A,IL-17F,IL-22,and IFN-γwere also upregulated in the DSS group(P<0.05).However,Zin treatment significantly relieved the above clinical symptoms and reversed the expression of above inflammatory factors(P<0.05).Flow cytometry indicated that Zin inhibited the differentiation of Th17 cells and the expression of RORγt,and promoted the differentiation of Treg cells both in vitro and in vivo(P<0.05),but the differentiation of Th1 subset was not affected.The dual-luciferase reporter assay demonstrated that the transcriptional activity of RORγt and Rorc promoter activity were remarkably suppressed after Zin treatment(P<0.05).ConclusionZin alleviates DSS-induced colitis in mice by regulating the balance of Th17/Treg cells and inhibiting RORγt expression.