TLR配体对URSA患者蜕膜调节性T细胞功能影响的研究

目的：检测原因不明复发性自然流产（URSA）患者蜕膜CD4^＋ CD25^high Treg在TLR4、TLR9配体刺激下功能的变化，探讨Toll样受体（TLRS）参与URSA发生的免疫机制。方法：选择2013年1～12月就诊于广州市妇女儿童医疗中心妇产科的URSA患者，其中B超确认胚胎已停育35例为研究对象，同期选择35例要求人工流产的计划外妊娠的正常早孕妇女为对照组。收集URSA患者、对照组清宫手术后的新鲜蜕膜组织，磁珠分离CD4^＋ CD25^high Treg，在脂多糖（LPS）、GpG—ODN2006刺激性下，分析CD4^＋CD25^high Treg增殖及对CD4^＋CD25^-T细胞增殖抑制的差异。结果：磁珠分选CD4^＋CD25^high Treg或CD4^＋CD25^-T细胞纯度可达到96％以上。与正常对照组相比，URSA患者蜕膜CD4^＋CD25^high Treg增殖能力相对较弱，在LPS、GpG—ODN2006的刺激性，增殖能力提高有限，二者差异有统计学意义（P〈0．05）。与正常对照组比较，uRsA患者蜕膜CD4^＋CD25^highTreg对CD4^＋CD25^-T抑制率明显下降，二者间存在显著差异（P〈0．05）；在LPS、GpG—ODN2006刺激下，URSA患者蜕膜CD4^＋ CD25^highTreg对CD4^＋CD25^-T抑制率变化不大，差异无统计学意义（P〉0．05）。对照组在LPS刺激性蜕膜CD4^＋CD25^high Treg对CD4^＋CD25^-T抑制率增强；在GpG—ODN2006刺激下，正常对照组CD4^＋ CD25^high Treg对CD4^＋ CD25^- T的增殖抑制作用明显减弱，差异有统计学意义（P〈0．05）。结论：URSA患者蜕膜中的CD4^＋CD25^high Treg功能受到抑制，TLRs配体刺激反应性下降可能参与URSA的发生。

Effects of TLRs agonists on decidual regulatory T cells in URSA patients

Objective： To detect the changes of decidual CD4^＋ CD25^high regulatory T with TLR4 and TLR9 agonists in URSA patients and explore the immune mechanism of TLRs involved in the patho- genesis of URSA. Methods： A total of 35 URSA patients with early embryo growth arrest were selected from January 2013 and December 2013. Meanwhile, 35 healthy pregnant women undergoing elective ter-minations during the first trimester were selected as URSA patients with spontaneous abortion and control group. Decidual samples were obtained from patients with induced abortion. Decidual CD4^＋ CD25^highTregs and CD4^＋ CD25^- T cells were separated by MACS. The proliferation of CD4^＋ CD25^high Tregs and the difference of suppressive function of CD4^＋ CD25^high Tregs with irritation of TLR4, TLR9 agonists were analyzed. Results： The MACS purification of CD4^＋ CD25^high Tregs and CD4^＋ CD25^- T cells was higher than 96 %. Decidual CD4^＋ CD25^high Tregs proliferated weakly, although with the irritation of LPS, GpG-ODN2006, limited proliferative capacity was increased in URSA patients and the differences were statistically significant compared with normal control group （P〈0.05）. Compared with normal con- trol group, inhibition of decidual CD4^＋ CD25^high Tregs on CD4^＋CD25^- T cells was decreased significantly in URSA patients and there was significant difference （P〈0.05） between the two groups. With LPS, GpG-ODN2006 irritating, inhibition of decidual CD4^＋ CD25^highTregs on CD4^＋ CD25^- T cells was changed and the differences were not statistically significant in URSA patients （P〉0.05）. Inhibition of decidual CD4^＋ CD25^high Tregs on CD4^＋ CD25^- T cells was enhanced significantly with LPS irritating, but decreased significantly under GpG-ODN2006 stimulation in control group（P〈0.05）. Conclusion. The function of decidual CD4^＋ CD25^high Tregs is suppressed in URSA patients and low reactivity of stimulation of TLRs ligands may involve in the pathogenesis of URSA.