四神丸及其拆方对结肠炎小鼠Treg细胞及PD-1/PD-L1表达的影响

Effect of Sisheng Pill and its disassembly on Expression of Treg cells and PD-1/PD-L1 in mice with colitis

目的探讨四神丸、二神丸及五味子散对结肠炎小鼠Treg细胞及PD-1/PD-L1表达的影响。方法将60只雄性BALB/c小鼠随机平均分为正常组、模型组、四神丸组、二神丸组、五味子散组、美沙拉嗪组,除正常组以外,其他组均采用葡聚糖硫酸钠复制慢性复发性溃疡性小鼠结肠炎模型。造模成功后,模型组和正常对照组小鼠均以饮用水灌胃,四神丸组小鼠给予2.5 g/kg四神丸溶液灌胃,二神丸组小鼠给予10 g/kg二神丸溶液灌胃,五味子散组小鼠给予6 g/kg五味子散溶液灌胃,美沙拉嗪组小鼠给予300 mg/kg美沙拉嗪混悬液灌胃,连续给药10天。观察实验小鼠体质量、结肠长度、结肠重量、结肠重量指数、病理损伤评分评价其疗效,并采用流式细胞术(FCM)检测小鼠外周血中Treg及其亚群CD4^(+)CD25^(+)Foxp3^(+)(Treg)、CD4^(+)CD25^(+)PD-1^(+)(PD-1^(+)Treg)和CD4^(+)CD25^(+)PD-L1^(+)(PD-L1^(+)Treg)细胞水平;结果与模型组相比,四神丸、二神丸及五味子散治疗组小鼠结肠长度增加,结肠体质量、结肠重量指数和结肠镜下病理损伤评分均显著降低(P<0.05或P<0.01),同时各治疗组PD-1^(+)Treg和PD-L1^(+)Treg细胞水平明显下调,Treg细胞水平明显上升(P<0.05或P<0.01)。结论四神丸、二神丸、五味子散可通过升高Treg细胞水平,抑制PD-1/PD-L1表达而治疗实验性结肠炎。

Objective To investigate the effects of Sishen Pill,Ershen Pill and Wuweizi Powder on Treg cells and PD-1/PD-L1 expression in mice with colitis.Methods Sixty male BALB/C mice were randomly divided into normal group,model group,Sishen pill group,Ershen pill group,Wuweizi powder group and mesalazine group.Except the normal group,the other groups were used to replicate the chronic recurrent ulcerative colitis model.After successful modeling,mice in model group and normal control group were given drinking water by instillation,mice in Sishen Pill group were given 2.5 g/kg Sishen pill solution,mice in Ershen pill group were given 10 g/kg Ershen pill solution,and mice in Wuweizi powder group were given 6 g/kg Schisandzi powder solution by instillation.Mice in mesalazine group were given 300 mg/kg mesalazine suspension by gavage for 10 consecutive days.Body weight,colonic length,colonic weight,colonic mass index and pathological injury score of experimental mice were observed to evaluate its efficacy.Flow cytometry(FCM)was used to detect the levels of Treg and its subsets CD4^(+)CD25^(+)Foxp3^(+)(Treg),CD4^(+)CD25^(+)PD-1^(+)(PD-1^(+)Treg)and CD4^(+)CD25^(+)PD-L1^(+)(PD-L1^(+)Treg)in peripheral blood of mice.Results Compared with the model group,the colon length of mice in the treatment groups of Sishen Pill,Ershen Pill and Wuweizi powder increased,and the colon body weight,colon mass index and pathological injury score under colonoscopy were significantly decreased.Meanwhile,the levels of PD-1^(+)Treg and PD-L1^(+)Treg cells in the treatment groups were significantly decreased,while the levels of Treg cells were significantly increased(P<0.05 or P<0.01).Conclusion Sishen pill,Ershen Pill and Wuweizi powder can treat experimental colitis by increasing Treg cell level and inhibiting PD-1/PD-L1 expression.