补肾解毒法对多发性硬化小鼠CD4^+T细胞的作用

Effect of Kidney-tonifying and Detoxicating Prescription on CD4^+T Cells in Mice with Multiple Sclerosis

目的研究补肾解毒法对多发性硬化小鼠中枢神经炎症及外周血CD4^+T细胞亚群(Th1、Th2、Th17、Treg细胞)的影响。方法以实验性自身免疫性脑脊髓炎(experimental autoimmune encephalomyelitis,EAE)模型作为多发性硬化动物模型,以C57BL/6小鼠皮下多点注射髓鞘少突胶质细胞糖蛋白35-55(myelin oligodendrocyte glycoprotein 35-55,MOG35-55)的方法复制EAE模型。将小鼠随机分为正常组,模型组,泼尼松组,中药高、中、低剂量组,每组8只小鼠。模型复制后第30天取小鼠脊髓,光镜下观察各组小鼠脊髓炎性细胞浸润和脱髓鞘等病理改变,流式细胞仪检测小鼠脾脏Th1、Th2、Th17、Treg细胞亚群,ELISA法检测脾脏细胞培养上清液中干扰素-γ(interferon gamma,IFN-γ)、白细胞介素-4(interleukin-4,IL-4)、白细胞介素-10(interleukin-10,IL-10)、白细胞介素-17A(interleukin-17A,IL-17A)水平。结果中药高剂量组、泼尼松组炎症评分和脱髓鞘评分明显低于模型组(P<0.05)。与模型组比较,中药高剂量组、泼尼松组脾细胞培养后上清液中致炎因子IFN-γ、IL-17A降低明显,而抑炎因子IL-4、IL-10升高明显,Treg比例明显升高,Th1和Th17比例明显降低,差异均具有统计学意义(P<0.05)。结论补肾解毒法能够调节EAE小鼠失衡的CD4^+T细胞亚群,显著抑制EAE模型的炎性反应。

Objective To investigate the effect of kidney-tonifying and detoxicating prescription on inflammation in the central nervous system and peripheral CD4^+T lymphocyte subsets(Th1,Th2,Th17,and Treg cells)in mice with multiple sclerosis.Methods Experimental autoimmune encephalomyelitis(EAE)was used for the animal model of multiple sclerosis.C57 BL/6 mice were given subcutaneous multi-point injection of myelin oligodendrocyte glycoprotein 35-55 to establish a mouse model of EAE.The mice were randomly divided into normal group,model group,prednisone group,and high-,medium-,and low-dose kidney-tonifying and detoxicating prescription groups,with 8 mice in each group.The spinal cord was harvested on day 30 after model establishment,and pathological changes of the spinal cords such as inflammatory infiltration and demyelination were observed under a light microscope.Flow cytometry was used to measure Th1,Th2,Th17,and Treg cells in the spleen,and ELISA was used to measure the levels of interferon-gamma(IFN-γ),interleukin-4(IL-4),interleukin-10(IL-10),and interleukin-17 A(IL-17 A)in the supernatant of splenic cell culture.Results The high-dose kidney-tonifying and detoxicating prescription group and the prednisone group had significantly lower scores of inflammation and demyelination than the model group(P<0.05).Compared with the model group,the high-dose kidney-tonifying and detoxicating prescription group and the prednisone group had significant reductions in the levels of IFN-γand IL-17 Aand significant increases in the levels of IL-4 and IL-10 in the supernatant of splenic cell culture,as well as a significant increase in the percentage of Treg cells and significant reductions in the percentage of Th1 and Th17 cells(P<0.05).Conclusion Kidney-tonifying and detoxicating prescription can regulate the imbalance of CD4^+T lymphocyte subsets and significantly inhibit inflammatory response in EAE mice.