黄芩苷对人淋巴细胞Toll样受体的调节

目的:研究黄芩苷对人淋巴细胞Toll样受体(TLR)的调节,探讨其在免疫调节方面的作用。方法:1.尼龙毛分离法分离人外周血T和B细胞后使用UF-50流式分析寻找最佳黄芩苷作用浓度;2.采用实时定量荧光RT-PCR,分析在最佳黄芩苷作用浓度处理前后TLRs的表达情况;3.通过加入鼠抗人TLR4单抗进行受体抑制实验并应用实时定量荧光RT-PCR探讨黄芩苷对TLRs的调节位点。结果:1.1 mg/ml的黄芩苷在体外可以显著促进人T、B淋巴细胞增殖(P<0.05);2.使用1mg/ml黄芩苷相对于未用黄芩苷处理的T、B细胞,其TLR3、TLR7、TLR8及TLR9 mRNA的表达均有显著增加(P<0.05),T细胞增加倍数分别为21、15、57和66倍,B细胞增加的倍数分别为24、20、61和63倍,而TLR1、TLR2、TLR4、TLR5、TLR6及TLR10 mRNAs表达变化不显著(P>0.05);3.在1 mg/ml黄芩苷作用下人T、B细胞TLR3、7、8和9 mRNA在12小时时表达已开始增加,除T细胞的TLR7 mRNA表达在48小时时略有下降外,其余表达基本一直持续增加,至48小时到达峰值;4.人T、B细胞在被黄芩苷作用前使用鼠抗人TLR4单克隆抗体封闭,会显著降低黄芩苷对TLR3、7、8和9 mRNA表达的诱导作用(P<0.05),T细胞下降比率分别为47.6%、66.7%、50.9%和45.5%;B细胞下降比率分别为50.0%、45.0%、50.8%和50.8%。结论:1.黄芩苷在体外能显著促进T、B淋巴细胞增殖;2.人外周血T、B细胞组成性表达含量差异较大的TLR1-10 mRNA,黄芩苷在体外可以显著上调人T、B淋巴细胞TLR3、TLR7、TLR8及TLR9 mRNAs的表达,但对TLR1、TLR2、TLR4、TLR5、TLR6及TLR10 mRNAs未见明显影响;3.黄芩苷可能通过TLR4发挥对人天然免疫能力的调节。

Regulation of baicalin on TLRs of the lymphocytes in human

Objective:To study the effect of baicalin on TLRs of the lymphocyte and function of immune regulation.Methods:1.After T and B lymphoytes were separated by nylon wool cotton column,we used UF-50 to test the optimum concentrations of baicalin to the proliferation of T,B lymphocytes.2.The different expression of TLRs mRNA before procession and after procession was analyzed by the real-time fluorescence quantitative polymerase chain reaction.3.The regulation point of baicalin for TLRs was studied by the receptor inhibition test using monoclonal antibodies rat against human TLR4 and the real-time fluorescence quantitative polymerase chain reaction.Results:1.Baicalin could significantly promote the proliferation of T and B lymphocytes of human in vitro at the concentration of 1 mg/ml(P<0.05);2.The mRNA of TLR3,TLR7,TLR8 and TLR9 on the lymphocyte was up-regulate by 1 mg/ml baicalin in vitro(P<0.05),the increased fold was 21,15,57 and 66 respectively in T lymphocytes,the increased fold was 24,20,61 and 63 respectively in T lymphocytes;3.The expression of mRNA of TLR3,TLR7,TLR8 and TLR9 on the T,B lymphocytes began to increase at 12 h,and kept increasing,nearly reached peak levels at 48 h,except TLR7 mRNA of T lymphocytes decreased slightly at 48 h;4.When T and B lymphocytes was inhibited by using monoclonal antibodies rat against human TLR4 before treated by baicalin,the up-regulation of mRNA of TLR3,TLR7,TLR8 and TLR9 was significantly decreased(P<0.05),which made the decrease rate was 47.6%,66.7%,50.9%,45.5% respectively in T lymphocytes and the decrease rate was 50.0%,45.0%,50.8%,50.8% respectively in B lymphocytes.Conclusion:1.To confirm that baicalin could significantly promote the proliferation of T and B lymphocytes of human in vitro;2.To confirm that T and B lymphocytes constitutively expressed different quantity of mRNAs for TLR1-10,the expression of mRNA of TLR3,TLR7,TLR8 and TLR9 on the lymphocyte was significantly up-regulate by baicalin in vitro,but baicalin had no influence for other TLRs;3.To confirm that Baicalin may regulate innate immunity of human through TLR4.