摘要
目的:从基因和蛋白水平探讨冬虫夏草菌粉提取液调节哮喘缓解期儿童体内T辅助淋巴细胞(T helperlymphocyte,Th)失衡及减轻炎症反应的机制,为冬虫夏草应用于哮喘缓解期提供理论依据。方法:采集20例哮喘缓解期儿童外周血3 mL,用Ficoll法分离出外周血单核细胞(peripheral bloodmononuclear cell,PBMC)。每份血样分为空白组,低、高剂量冬虫夏草菌粉提取液(10、20μg/mL)组。细胞培养箱中37℃、5%CO_2培养48 h后,利用实时荧光定量聚合酶链式反应(polymerase chain reaction,PCR)法检测PBMCγ干扰素(interferon-γ,IFN-γ)、白细胞介素4(interleukin-4,IL-4)、IL-10、T细胞表达的T盒(T-box expressed in T cells,T-bet)、GATA连接蛋白3(GATA-binding protein-3,GATA-3)、叉头状转录因子3(forkhead/winged-helix transcription factor-3,Foxp3)mRNA的表达,利用酶联免疫吸附测定法检测细胞培养上清液中IFN-γ、IL-4及IL-10含量。结果:3组PBMC IFN-γmRNA的表达比较,差异无统计学意义;高剂量和低剂量冬虫夏草菌粉提取液组IL-4 mRNA的表达明显低于空白组(P=0.014,P=0.011);高剂量冬虫夏草菌粉提取液组IL-10 mRNA的表达高于空白组(P=0.034);3组IFN-γmRNA/IL-4 mRNA比值比较,差异无统计学意义。高剂量冬虫夏草菌粉提取液组细胞培养上清中IL-4含量低于空白组(P=0.018),IL-10含量及IFN-γ/IL-4比值均高于空白组(P=0.011,P=0.045);3组IFNγ含量比较,差异无统计学意义。高剂量和低剂量冬虫夏草菌粉提取液组PBMC T-bet/GATA-3比值及Foxp3 mRNA的表达均高于空白组(P=0.001,P=0.015);高剂量冬虫夏草菌粉提取液组GATA-3 mRNA的表达明显低于空白组(P=0.028);3组T-bet mRNA的表达比较差异无统计学意义。结论:冬虫夏草菌粉提取液可下调PBMC中GATA-3 mRNA的表达和上调Foxp3 mRNA的表达,增加IL-10的含量,以此减轻炎症反应。
Objective：To explore the effects of Cordyceps extract in regulating the imbalance of Th1/Th2 ratio and inhibiting the inflammatory reaction,and to find the theoretical basis of Cordyceps extract for treating asthma in remission stage. Methods：A total of 20 peripheral venous blood samples（3 mL） were collected from 20 asthmatic children during remission stage,and peripheral blood mononuclear cells（PBMCs） were separated by Ficoll method. PBMCs were separated into three groups（blank group,low-dose group and high-dose group）.The PBMCs were incubated in vitro for 48 hours in the absence（blank group） or presence（low-dose group and high-dose group） of Cordyceps extract at different concentrations（10,20μg/mL）.The expressions of interferon-γ（IFN-γ）,interleukin-4（IL-4）,IL-10,T-box expressed in T cells（T-bet）,GATA-binding protein-3（GATA-3） and forkhead/winged-helix transcription factor-3（Foxp3） mRNAs in PBMCs were measured by real-time fluorescent quantitative polymerase chain reaction,and the contents of IFN-γ,IL-4 and IL-10 in supernatants were measured by enzyme-linked immunosorbent assay. Results：The expressions of IFN-γmRNA showed no significant differences among the three groups.The expressions of the IL-4 mRNA in the high-dose group and the low-dose group were lower than that in the blank group（P=0.014,P=0.011）.The expression of IL-10 mRNA in the high-dose group was higher than that in the blank group（P=0.034）.And the differences of the IFN-γmRNA/IL-4 mRNA ratio presented no statistic significance among the three groups.The level of IL-4 content in the high-dose group was lower than that in the blank group（P=0.018）,but the level of IL-10 content,and ratio of IFN-γ/IL-4 in the high-dose group were higher than those in the blank group（P=0.011,P=0.045）.The differences of the IFN-γpresented no statistic significance among the three groups.The T-bet/GATA-3 ratio and Foxp3 mRNA expression in the high-dose group were higher than those in the blank group（P=0.001,P=0.015）.There was significant difference in expression of GATA-3 mRNA between the high-dose group and the blank group（P=0.028）,and between the low-dose group and the blank group（P=0.019）.The expression differences of T-bet mRNA were insignificant between any two groups. Conclusion：Cordyceps extract can inhibit the proliferation and differentiation of Th2 cells and reduce the expression of related cytokines by down-regulating the expression of GATA-3 mRNA and up-regulating the expression of Foxp3 mRNA in PBMCs.Meanwhile,it can alleviate the chronic allergic inflammation by increasing the content of IL-10.
出处
《中西医结合学报》
CAS
2010年第4期341-346,共6页
Journal of Chinese Integrative Medicine
基金
复旦大学儿科医院重点科研基金资助项目(No.0620-06)
关键词
冬虫夏草
哮喘
单核细胞
缓解期
细胞因子类
转录因子
Cordyceps sinensis
asthma
monocytes
remission stage
cytokines
transcription factors
