氢气对脓毒症大鼠免疫抑制状态的影响

Effect of hydrogen on immunosuppressive status of septic rats

目的评价氢气对脓毒症大鼠免疫抑制状态的影响。方法SPF级健康成年雄性SD大鼠,7~8周龄,体重220~260 g。实验Ⅰ采用随机数字表法分为2组:脓毒症组(Sep组,n=36)和假手术组(Sham组,n=12)。采用盲肠结扎穿孔法(CLP)制备大鼠脓毒症模型。于CLP术后即刻、1、2、3和4 d时采用流式细胞术检测外周血组织相容性DR抗原(HLA-DR)/CD14^(+)单核细胞水平。外周血HLA-DR/CD14^(+)单核细胞水平<30%为脓毒症免疫抑制模型制备成功。实验Ⅱ随机选取脓毒症免疫抑制大鼠12只,采用随机数字表法分为2组(n=6):脓毒症免疫抑制组(Sep-IS组)、脓毒症免疫抑制^(+)氢气处理组(Sep-IS+H组);另取正常大鼠12只,采用随机数字表法分为2组(n=6):假手术组(Sham组)和假手术^(+)氢气组(Sham^(+)H组)。Sep-IS^(+)H组于脓毒症免疫抑制成功制备即刻、制备后6 h时,Sham^(+)H组于相应时点吸入67%氢气1 h。于上述时点氢气吸入结束即刻(T0、T1)和模型制备后12 h(T_(2))时采用流式细胞术检测外周血辅助T淋巴细胞17(Th17细胞)、调节性T淋巴细胞(Treg细胞)和HLA-DR/CD14^(+)单核细胞水平。结果实验Ⅰ与Sham组相比,Sep组CLP术后1~4 d时外周血HLA-DR/CD14^(+)水平降低(P<0.05)。实验Ⅱ与Sham组相比,Sep-IS组、Sep-IS^(+)H组各时点外周血HLA-DR/CD14^(+)单核细胞水平降低,Treg细胞、Th17细胞水平升高(P<0.05),Sham^(+)H组上述指标差异无统计学意义(P>0.05)。与Sep-IS组相比,Sep-IS^(+)H组T_(1,2)时外周血HLA-DR/CD14^(+)单核细胞水平升高,T_(2)时外周血Th17细胞水平升高,T_(1,2)时外周血Treg细胞水平降低(P<0.05)。结论氢气可改善脓毒症大鼠免疫抑制状态。

Objective To evaluate the effect of hydrogen on the immunosuppressive status of septic rats.Methods SPF healthy adult male Sprague-Dawley rats,aged 7-8 weeks,weighing 220-260 g,were studied.This study was performed in two parts.PartⅠThe rats were divided into 2 groups:sepsis group(Sep group,n=36)and sham operation group(Sham group,n=12).The model of sepsis was established by cecal ligation puncture in anesthetized rats.The histocompatibility DR antigen(HLA-DR)/CD14^(+)monocyte level in peripheral blood was detected by flow cytometry immediately after CLP and at 1,2,3 and 4 days after CLP.The establishment of sepsis-induced immunosuppression model was considered successful when the levels of HLA-DR/CD14^(+)monocyte in peripheral blood were<30%.PartⅡTwelve rats with sepsis-induced immunosuppression were randomly selected and divided into 2 groups(n=6 each)by a random number table method:sepsis immunosuppression group(Sep-IS group),sepsis immunosuppression plus hydrogen treatment group(Sep-IS+H group).Another 12 rats were selected and divided into 2 groups(n=6 each)by a random number table method:sham operation group(Sham group)and sham operation plus hydrogen group(Sham^(+)H group).In Sep-IS+H group,67%hydrogen was inhaled for 1 h starting from the time point immediately after successful establishment of sepsis-induced immunosuppression and from 6 h after establishment,and 67%hydrogen was inhaled for 1 h at the corresponding time points in Sham^(+)H group.The levels of helper T lymphocytes 17(Th17 cells),regulatory T lymphocytes(Treg cells)and HLA-DR/CD14^(+)monocyte in peripheral blood were determined by flow cytometry immediately after the end of hydrogen inhalation mentioned above(T0,T1)and at 12 h after establishing the model(T_(2)).Results PartⅠCompared with Sham group,the levels of HLA-DR/CD14^(+)monocyte in peripheral blood were significantly decreased at 1-4 days after CLP in Sep group(P<0.05).PartⅡCompared with Sham group,the level of HLA-DR/CD14^(+)monocytes in peripheral blood was significantly decreased,and the levels of Treg and Th17 cells were increased at each time point in Sep-IS and Sep-IS+H groups(P<0.05),and no significant change was found in the parameters mentioned above in Sham^(+)H group(P>0.05).Compared with Sep-IS group,the level of HLA-DR/CD14^(+)monocytes in peripheral blood was significantly increased at T_(1,2),the levels of Th17 cells were increased at T_(2),and the levels of Treg cells were decreased at T_(1,2)in Sep-IS+H group(P<0.05).Conclusion Hydrogen can improve the immunosuppressive state of septic rats.