补肾固表方对反复呼吸道感染小鼠Toll样受体4和CD4＋CD25＋foxp3＋调节性T细胞的影晌

目的：探讨补肾固表方（Bushen Gubiao Recipe,BGR）对氢化可的松诱导的反复呼吸道感染（recurrent respiratory tract infection,RRTI）小鼠天然免疫Toll样受体4（toll-like receptor 4,TLR4）和CD4＋CD25＋foxp3＋调节性T细胞（CD4＋CD25＋foxp3＋regulatory T cell,CD4＋CD25＋foxp3＋Treg）的影响及量效关系。方法：应用氢化可的松连续腹腔注射BALB/c小鼠14 d,作为RRTI肾阳虚模型,随机分为BGR大、小剂量组,核酪组,模型组,分别灌胃BGR大、小剂量煎剂,核酪口服液和蒸馏水,正常组灌胃蒸馏水,干预4周后,评价小鼠外观行为学,免疫组织化学SABC法检测肺组织TLR4和核因子-κB（nuclear factor-kappaB,NF-κB）p65的表达,荧光定量聚合酶链反应检测肺组织TLR4 mRNA的表达,流式细胞术检测血CD4＋CD25＋foxp3＋Treg细胞水平。结果：BGR能显著改善RRTI小鼠外观和行为异常,提高肺气道TLR4、NF-κB p65免疫阳性表达,与模型组比较差异有统计学意义（P〈0.05）,未见中药大、小剂量组间差异（P〉0.05）,核酪组未见明显变化（P〉0.05）。大剂量BGR和核酪能够降低CD4＋CD25＋foxp3＋Treg水平,与模型组比较差异有统计学意义（P〈0.05）,中药大、小剂量组间未见显著差异（P〉0.05）。结论：补肾固表方能够改善小鼠肾阳虚状态;通过上调TLR4/NF-κB信号通路,提高气道天然免疫功能;下调CD4＋CD25＋foxp3＋Treg活性,增强辅助性T细胞（T-helper,Th）1型免疫反应,调节Th1/Th2平衡。

Effects of Chinese herbal medicine Bushen Gubiao Recipe on toll-like receptor 4 and CD4(+)CD25(+)foxp3(+) regulatory T cells in mice with recurrent respiratory tract infections

Objective： To evaluate the effects of Bushen Gubiao Recipe（BGR）,a compound traditional Chinese herbal medicine,on toll-like receptor 4（TLR4） signaling pathway and CD4＋CD25＋foxp3＋regulatory T cells（CD4＋CD25＋foxp3＋Tregs） in mice with recurrent respiratory tract infections（RRTIs）.Methods： A mouse model of kidney-yang deficiency which simulated physical characteristics of RRTIs was established by intraperitoneal injection of hydrocortisone for 14 d.The model mice were divided into 4 groups,model group,high-dose BGR group,low-dose BGR group,and nucleic acid and casein oral solution group.They were administered respectively with distilled water,high-dose BGR（50 g/kg body weight）,low-dose BGR（25 g/kg body weight） and nucleic acid and casein oral solution.Besides,a normal control group was set up and gastrogavaged with distilled water.The effect of intervention was evaluated 4 weeks later by estimating the changes in behaviors of mice.Expressions of TLR4 and nuclear factor-kappa B（NF-？？） p65 in lung tissue were detected by immunohistochemical SABC method,the expression of TLR4 mRNA in lung tissue was detected by fluorescence quantitative-polymerase chain reaction（FQ-PCR）,and the level of blood CD4＋CD25＋foxp3＋ Tregs was determined by flow cytometry.Results： A kidney-yang deficiency mouse model with RRTIs was successfully established by intraperitoneal injection of hydrocortisone.BGR could improve the abnormal behavioral condition of the mice and enhance the expressions of TLR4 and NF-？？ p65 in the lung tissue.Expression of TLR4 mRNA in high-dose BGR group was higher than that in model group（P0.05）,while the difference was not statistically significant between high-dose BGR group and low-dose BGR group（P0.05）.Levels of CD4＋CD25＋foxp3＋Tregs in high-dose BGR group and nucleic acid and casein oral solution group were lower than that in model group（P0.05）,while the difference was not statistically significant between high-dose BGR group and low-dose BGR group（P0.05）.Conclusion： BGR can improve the behavior of the kidney-yang deficiency mice,and improve the innate immune function by up-regulating TLR/NF-？？ signaling pathway.BGR can adjust the immune imbalance of T-helper cell（Th） 1/Th2 through reducing the activity of CD4＋ CD25＋foxp3＋Tregs and enhancing the immune response of Th1 type.