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中文摘要：
目的观察高迁移率族蛋白B1（HMGB1）对健康人T淋巴细胞增殖的影响，并对其机制进行初步探讨。方法分离健康人外周血单个核细胞（PBMCs），调整细胞浓度后接种于细胞培养板并加入重组人高迁移率族蛋白B1（rhHMGB1）进行刺激。以四甲基偶氮唑盐微量Ⅱ酶反应比色法（MTT）检测细胞数量和细胞活性，观察HMGB1对T淋巴细胞增殖活性的影响。采用四色流式细胞术（FCM）分析CD3^＋淋巴细胞CD4表达。细胞中自细胞介素-2（IL-2）、IL-2α受体（IL-2Rα）基因表达水平采用逆转录一聚合酶链反应（RT—PCR）分析。结果①500～1000μg／L rhHMGB1作用48h后T淋巴细胞增殖反应显著抑制，低于这一剂量对其增殖活性影响不显著。②不同rhHMGB刺激时间和作用剂量对CD4^＋T淋巴细胞未造成明显改变，但rhHMGB1能时间-剂量依赖性增加Th2亚群比例，并因此降低Th1／Th2比值，刺激后T淋巴细胞免疫功能出现Th1优势向Th2优势偏移。③经植物血凝素激活后12hT淋巴细胞IL-2和IL-2Ra基因表达达到峰值；rhHMGB1与T淋巴细胞共同培养12h后，10～100μg／L剂量可明显上调IL-2和IL-2Ra基因表达；而较高剂量rhHMGB1（100～1000μg／L）刺激持续48h上述效应衰竭，并表现出相反的变化趋势。结论HMGB1对T淋巴细胞包括增殖、分化和细胞因子分泌等免疫功能具有直接调节效应。剂量蓄积和持续刺激可诱导T淋巴细胞功能亚群从促炎优势向抗炎优势转化。
英文摘要：
Objective To investigate the effect of high mobility group box-1 protein （HMGB1） on immune function of human T lymphocytes in vitro and explore its potential role in cell-mediated immune dysfunction. Methods Fresh blood was obtained from healthy adult volunteers and peripheral blood mononuclear cells （PBMCs） were isolated, then rhHMGB1 was added to PBMCs. Cell viability was assessed by thiazolyl blue （MTT） assay. Four-color flow cytometric （FCM） analysis was used for the measurement of CD3, CD8 expression. Reverse transcription-polymerase chain reaction amplification was performed to detect respective gene expression of interleukin-2 （IL-2）, IL-2 receptor （IL-2R） alpha. Results ①Proliferation of T lymphocytes was not affected by rhHMGB1 in low concentrations, while continued exposure of T cells to 500 - 1 000 μg/L rhHMGB1 for 48 hours resulted in a decrease in MTT assay. ② Different stimulating time and dosages of rhHMGB1 did not alter CD4 expression of CD3^＋ T lymphocytes, rhHMGB1 stimulation provoked a dose-dependent and time-dependent increase in Th2 subset and lowering in ratio of Thl to Th2. ③ Compared with the untreated cells, when the cells were co-incubated with rhHMGB1 （10μg/L） for 12 hours, mRNA expressions of IL-2 and IL-2R were significantly up-regulated. At 48 hours, in contrast, gene expression was relatively lower in T cells after exposure to 100 - 1 000μg/L rhHMGB1. Conclusion These data demonstrated that HMGB1 had a dual influence on immune functions of T lymphocytes.
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