严重烧伤后调节性T细胞免疫活性变化及其与患者预后的关系

Immunological activity change of regulatory T cells and its relation with outcome of patients with severe burn

目的 探讨严重烧伤后患者外周血调节性T细胞（Treg）免疫活性的变化规律,并探讨其在MODS和患者不良预后中的临床意义. 方法对106例烧伤总面积（TBSA）≥30%的患者进行动态观察.试验分组：（1）按烧伤面积将患者分为三组：Ⅰ组41例（TBSA 30%～49%）,Ⅱ组34例（TBSA 50%～69%）,Ⅲ组31例（TBSA 70%～99%）;（2）根据MODS诊断标准,将患者进一步分为MODS组（21例）和非MODS组（85例）;（3）根据MODS患者的预后情况,将其分为死亡组（16例）和生存组（5例）;同时设正常对照组.分别于伤后1,3,7,14,21 d采集患者外周静脉血,采用免疫磁性分离系统对外周血CD4＋CD25＋Treg进行分离纯化,采用流式细胞术检测Treg表面分子T淋巴细胞毒性相关抗原4（CTLA-4）的表达情况,ELISA法检测Treg孵育液上清中IL-10的分泌水平. 结果 （1）与正常对照组比较,严重烧伤患者各烧伤面积组CD4＋CD25＋Treg表面分子CTLA-4表达水平及IL-10分泌水平均明显增强（P＜0.01）,烧伤患者各组间差异有统计学意义（P＜0.05或＜0.01）;（2）严重烧伤患者伤后不同时相点CTLA-4阳性表达水平及IL-10分泌水平均明显升高,MODS组CTLA-4阳性表达率及IL-10分泌水平在伤后3～21 d显著高于非MODS组（P＜0.01）;（3）MODS患者死亡组CTLA-4阳性表达率及IL-10分泌水平在伤后3～21 d均显著高于生存组（P＜0.05或＜0.01）. 结论严重烧伤后CD4＋CD25＋Treg表面分子表达及细胞因子分泌在不同烧伤面积、是否并发MODS及是否生存等不同组间存在显著差异,CD4＋CD25＋Treg可通过分泌抑制性细胞因子参与严重烧伤后机体免疫失衡、诱发多脏器功能障碍发生甚至最终造成患者死亡的病理过程.

Objective To investigate the immunological activity change of regulatory T cells （Treg） and discuss its significance in the outcomes of patients with multiple organ dysfunction syndrome （MODS） and severe burn. Methods A total of 106 patients with total burn surface area （TBSA） larger than 30% were included in the study and randomly divided into three groups according to the burn area： Group Ⅰ （TBSA of 30%-49%, n = 41）, Group Ⅱ （TBSA of 50% -69%, n = 34） and Group Ⅲ （TBSA of 70%-99%, n = 31）. According to the development of MODS, patients were divided into MODS group （n =21） and non-MODS group （n =85）. The patients with MODS were further divided into non-survival group （n = 16） and survival group （n = 5） based on their outcomes. Healthy volunteers were served as normal control （n = 25）. Peripheral blood samples were collected at days 1,3,7, 14 and 21 postburn. The immunomagnetic separation technique was applied to separate and purify CD4＋ CD25＋Tregs in peripheral blood, and phenotypes （CTLA-4） were analyzed by flow cytometry and the contents of interleukin-10 released in the supernatants were determined by ELISA. Results Expression of CTLA-4 and level of IL-10 were significantly increased in burn patients compared with normal control group, with statistical differences. The expression of CTLA-4 and level of IL-10 were significantly increased in patients with severe burns at all time points. The expression of CTLA-4 and level of IL-10 in MODS group were much higher than those in non-MODS group at days 3-21 postburn （P 〈 0.01）. Among the MODS patients, the expression of CTLA-4 and level of IL-10 in the survival group were obviously lower than those in the non-survival group at days 3-21 postburn （P 〈 0.05 or P 〈 0.01）. Conclusions After severe burn injury, expressions of the markers on CD4 ＋ CD25 ＋ Treg surface and secretion of cytokines produced by CD4 ＋ CD25 ＋ Tregs show significant difference in patients with different born areas, MODS development and survival state. CD4 ＋ CD25 ＋ Treg may play an important role in the pathogenesis of immunoregulation, MODS and mortality of burn patients through secretion of inhibitory cytokines.