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    牛磺酸影响人外周血T淋巴细胞增殖及分化功能的体外研究

    Effect of taurine on the proliferation and differentiation of human peripheral blood T lym⁃phocytes in vitro

      摘要
    • 摘要: 目的：研究牛磺酸（Tau）对人外周血T淋巴细胞体外活化增殖及分化的影响及其作用机制。方法：无菌分离健康志愿者外周血淋巴细胞，建立植物凝集素（PHA）刺激T 细胞体外活化增殖模型。实验设置对照组（control 组）、PHA 刺激组（PHA 5 μg/mL 组）及Tau 处理组（PHA+Tau 80 mmol/L 组、PHA+Tau 160 mmol/L 组）。细胞计数试剂盒（CCK-8）法检测T 淋巴细胞增殖率；瑞—吉染色法观察细胞形态及计算转化率；实时荧光定量PCR 检测T 细胞增殖标志物Ki67，Th1 转录因子T-bet 及细胞因子干扰素-γ（IFN-γ）、肿瘤坏死因子-α（TNF-α），Th2 转录因子GATA-3 及白介素-4/6（IL-4/6），凋亡相关因子Fas、FasL 基因表达水平；活性氧（ROS）试剂盒检测细胞内ROS水平。结果：与control组相比，PHA 5 μg/mL组T淋巴细胞转化率、增殖率、增殖标志物Ki67、Th1 相关因子T-bet、IFN-γ、TNF-α、凋亡相关因子Fas 基因表达及胞内ROS 水平升高（P< 0.05），凋亡相关因子FasL基因，Th2相关因子GATA-3、IL-6基因表达水平降低（P< 0.05）。与PHA 5 μg/mL组相比，PHA+Tau 80 mmol/L组与PHA+Tau 160 mmol/L 组T 淋巴细胞转化率、增殖率、Ki67 基因表达及胞内ROS 水平均下降（P< 0.05），Th2 相关因子IL-4、IL-6 基因表达水平均升高（P< 0.05），Th1 相关因子IFN-γ、FasL 基因表达水平差异均无统计学意义（P> 0.05），PHA+Tau 80 mmol/L 组Th1相关因子T-bet、TNF-α、Th2相关因子GATA-3及Fas 基因表达水平差异无统计学意义（P> 0.05），PHA+Tau 160 mmol/L 组Th1相关因子T-bet、TNF-α、Th2相关因子GATA-3 及Fas基因表达水平升高（P< 0.05）。结论：Tau可能通过下调Ki67表达和增强Fas-AICD途径抑制T细胞增殖转化，并可能通过降低胞内ROS含量，提高Th1/Th2活化水平，调节Th1/Th2平衡偏向Th1分化，发挥调节T淋巴细胞活化增殖及分化的作用。

       

      Abstract: Objective:To study the effect and mechanism of taurine(Tau)on the activation, proliferation, and dif-ferentiation of human peripheral blood T lymphocytes in vitro.Methods:Peripheral blood lymphocytes were aseptically isolated from healthy volunteers to establish the model of activation and proliferation of T cells stimu-lated by phytohaemagglutinin (PHA) in vitro.The control group, PHA stimulation group (PHA 5 μg/mL group), and Tau treatment group (PHA+Tau 80 mmol/L group, PHA+Tau 160 mmol/L group) were set up.Cell counting kit-8(CCK-8)assay detected the T lymphocyte proliferation rate.The cell morphology was observed with Wright-Giemsa staining and the conversion rate was calculated.T lymphocyte proliferation marker Ki67, Th1 transcrip-tion factor T-bet and cytokines interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), Th2 transcription factors GATA-3 and interleukin-4/6(IL-4/6)and the expres-sions of apoptosis-related factors Fas and FasL gene were detected by real-time fluorescence quantitative polymerase chain reaction (RT-qPCR).Reactive ox-ygen species (ROS) kits were used to detect the level of intracellular ROS.Results:Compared with the control group, T lymphocyte conversion rate, proliferation rate, proliferation marker Ki67, Th1-related factors T-bet, IFNγ, TNF-α, apoptosis-related factor Fas gene expression and intracellular ROS level of PHA 5 μg/mL group in-creased (P< 0.05), while apoptosis-related factor FasL gene, Th2-related factors GATA-3 and IL-6 gene expres-sion levels decreased(P< 0.05).Compared with the PHA 5 μg/mL group, T lymphocyte conversion rate, prolifer-ation rate, Ki67 gene expression and intracellular ROS level in PHA+Tau 80 mmol/L group and PHA+Tau 160 mmol/L group decreased (P< 0.05).The gene expression levels of Th2-related factors IL-4 and IL-6 increased(P< 0.05).There were no significant differences in the gene expression levels of Th1-related factors IFN-γ and FasL (P> 0.05).There were no significant differences in the gene expression levels of Th1-related factors T-bet and TNF-α, and Th2-related factors GATA-3 and Fas in PHA+Tau 80 mmol/L group(P> 0.05).The gene expres-sion levels of Th1-related factors T-bet and TNF-α, and Th2-related factors GATA-3 and Fas increased in PHA+Tau 160 mmol/L group (P< 0.05).Conclusion:Tau may inhibit the proliferation and conversion of T lympho-cytes by down-regulating the expression of Ki67 and enhancing the Fas-AICD pathway.Moreover, Tau may play a role in regulating the activation, proliferation and differentiation of T lymphocytes by reducing intracellular ROS level, increasing Th1/Th2 activation levels and regulating the balance of Th1/Th2 towards Th1 differentia-tion.

       

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