一氧化氮抑制T细胞产生细胞因子但促进单核细胞细胞因子的分泌

Nitric oxide inhibits the cytokines production of T cells but promotes the cytokines production of monocytes

目的研究一氧化氮(nitric oxide,NO)对T细胞和单核细胞分泌细胞因子的作用及机制。方法用抗人CD3抗体或抗人CD3抗体联合抗人CD28抗体或LPS刺激培养外周血单个核细胞(PBMCs)、纯化T细胞或纯化单核细胞,加或不加一氧化氮培养,ELISA检测培养细胞上清液中细胞因子的含量;流式细胞术检测T细胞表面活化分子CD69和CD25的表达、T-box转录因子(T-bet)的表达,STAT1和STAT4的磷酸化程度以及T细胞的分裂情况。结果一氧化氮呈时间和剂量依赖方式抑制T细胞产生IFN-γ、TNF-α和IL-2,但促进单核细胞IL-1β、IL-6和TNF-α的分泌,且不诱导细胞的凋亡。进一步研究发现其抑制细胞晚期活化分子的表达和细胞的分裂,以及抑制T细胞相关转录因子T-bet的表达,但不抑制STAT1和STAT4的磷酸化程度。结论一氧化氮通过非细胞毒性作用抑制活化的T细胞产生细胞因子,但促进活化的单核细胞的细胞因子的分泌,其机制是通过抑制转录因子T-bet信号通路而抑制T细胞细胞因子的分泌,该研究将为临床应用提供理论依据。

To investigate the effect and mechanism of NO on the production of cytokines by T lymphocytes and monocytes, PBMCs and purified CD4+, CD8+T cells or CD14+monocytes were stimulated with anti-CD3 plus antiCD28 or LPS in the presence or absence of NO. The levels of IFN-γ, TNF-α, IL-1β, IL-2 and IL-6, in the culture supernatants were detected by ELISA;the production of cytokines and transcription factors including T-bet, pSTAT-1 and p-STAT-4 were analyzed by FACs. Data showed that NO suppressed the activation and dividing from T lymphocytes through a non-cytotoxic mechanism. In addition, NO directly inhibited the production of IFN-γ,TNF-α and IL-2 by purified CD4+and CD8+T cells in a dose-time-dependent manner, but promoted the production of TNF-α, IL-1β and IL-2 by CD14+monocytes. Further study revealed that NO down-regulated the expression of T-bet but not the phosphorylation of STAT1 or STAT4. All these results indicated that the main mechanism of NO inhibiting the cytokine production is down-regulating the expression of T-bet. The further study on the mechanism of NO immunosuppression is of great significance for the clinical study of immunosuppressive drugs.