中文摘要：

目的观察高迁移率族蛋白B1（HMGBl）对调节性T细胞（Treg）Toll样受体4（TLR4）表达的影响，初步探讨其与Treg免疫活性的关系。方法用免疫磁珠法分离正常C3H／HeN（TLR4受体野生型）小鼠脾脏CD4^＋CD25^＋Treg。采用可溶性抗CD3抗体（1mg／L）和抗CD28抗体（1mg／L）辅助活化，观察不同浓度HMGBl（0、10、100、1000μg／L）刺激不同时间（24、48、72h）后Treg TLR4表达的时间-效应关系和剂量-效应关系。结果以100μg／L浓度的HMGBl刺激，Treg TLR4在24、48和72h表达水平均明显受抑（P均〈0．01），但各时间点组间比较差异无统计学意义（P均〉0．05）；不同剂量HMGBl刺激48h可诱导Treg TLR4表达水平下调，其中以100μg／L、1000μg／L HMGBl作用时Treg TLR4表达降低尤为明显（P〈O．05和P〈0．01）。结论HMGBI能显著诱导Treg表达TLR4下调，进而参与调节Treg的免疫活性。

英文摘要：

Objective To investigate the effect of high mobility group box-1 protein （HMGB1） on Toll-like receptor 4 （TLR4） expression of regulatory T cells （Tregs） and its relationship to the immunologic activity of Tregs in mice. Methods CD4^＋CD25^＋Tregs were isolated from C3H/HeN （TLR4 wild type） murine splenocytes with a CD4＋ CD25＋Tregs isolation kit. Tregs were stimulated on Tregs by HMGB1 in 96-well （1×10^5 cells/well） cell plates cultured with anti-mouse CD3 （1 rag/L） and anti-mouse CD28 （1 mg/L）. After being stimulated, Tregs were harvested to determine the expression of TLR4 and analyze the time-and dose-dependent response between HMGB1 （0, 10, 100, and 1 000/μg/L） and TLR4 expression by flow cytometry at different time points （24, 48 and 72 hours）. Results Compared with the normal control group, the expression of TLR4 on Tregs stimulated by HMGB1 （100/μg/L） was significantly down-regulated at 24, 48, and 72 hours, respectively （all P〈0.01）, and no differences in TLR4 expression on Tregs were observed among various intervals （all P 〉 0.05）. Meanwhile, HMGB1 at various doses （10, 100, and 1 000/μg/L） also could induce the significant down-regulation of TLR4 expression on Tregs, and it maintained low value on Tregs when stimulated by HMGBI at doses of 100/μg/L or 1 000/μg/L （P〈0. 05 and P〈0. 01）, nevertheless, no differences in TLR4 expressions were observed between 100 /μg/L and 1 000/μg/L groups （P〉0.05）. Conclusion HMGB1 can markedly down-regulate the expression of TLR4 on Tregs, thereby possibly modulating the immunologic activity of Tregs.