目的 分析慢性乙型肝炎患者外周血中IL-10+ CD19+调节性B淋巴细胞(Breg)的表达及其免疫调节机制.方法 病例对照研究.收集201 1年6月-2012年10月常熟市第二人民医院住院患者,急性乙型肝炎(AHB)患者28例,慢性乙型肝炎(CHB)患者31例和健康对照25名.分离外周血单个核细胞(PBMC),经含CpG序列的寡脱氧核苷酸CpG ODN 2006和佛波醇酯(PMA)体外作用后,流式细胞术分析IL-10+ CD19+ Breg、CD4+ CD25high调节性T淋巴细胞(Treg)的表达及其相关性,ELISA分析培养上清IL-10水平.Mann-Whitney U检验分析两组间差异,Spearman相关分析两组连续变量间的相关性.结果 CHB组外周血Breg[1.28％ (1.05％ ～2.20％)]高于AHB组[0.87％(0.55％～1.22％)]和HCs组[0.89％(0.51％～1.37％)](P值分别为0.001和0.006),AHB和HCs组间差异无统计学意义(P=0.669).CpG ODN 2006和PMA作用后CHB组Breg[14.30％(10.70％～16.70％)]较AHB组[10.30％ (7.05％ ～ 13.30％)]和HCs组[10.40％ (6.85％ ～12.60％)]显著升高(P值分别为0.003和0.001)；CHB组Treg[5.80％ (4.20％ ～9.10％)]较AHB组[4.05％(2.53％～5.40％)]和HCs组[4.50％(2.55％～5.50％)]也显著增高(P值分别为＜0.001和0.005).AHB和HCs组间差异均无统计学意义(Breg:P=0.796；Treg:P=0.227).Spearman相关性分析发现CHB组Breg与Treg存在正相关关系(r=0.50,p=0.004),而AHB和HCs组无此相关性(r=-0.15,P=0.462;r=0.09,P=0.669).CHB组IL-10表达高于AHB和HCs组(P＜0.001).AHB和HCs组间差异无统计学意义(P=0.341),Spearman相关性分析表明CHB组IL-10与Breg存在正相关关系(r=0.409,P=0.005).结论 B淋巴细胞Breg和调节性T淋巴细胞在慢性乙型肝炎患者中表达升高,Breg可能通过IL-10的分泌在HBV慢性感染过程中发挥免疫调节作用.%Objective To investigate the population and role of IL-10+ CD19+ regulatory B cell (Breg) in patients with chronic hepatitis B.Methods Patients with acute hepatitis B (AHB) (n =28),chronic hepatitis B (CHB) (n =31) and normal subjects (n =25) were collected from Changshu No.2 People's Hospital between 2011 June and 2012 October.Peripheral blood mononuclear cells (PBMC) were isolated and stimulated with CpG ODN 2006 and PMA.Flow cytometry was used to analyze the population of IL-10-CD19 + Breg,CD4 + CD25high Treg,and ELISA was used to analyze the concentration of IL-10 in culture supernatant.Results The population of Breg in Peripheral blood of the CHB group [1.28％ (1.05％-2.20％)] was higher than that in the AHB group [0.87％(0.55％-1.22％)] and the HCs group [0.89％ (0.51％-1.37％)] (P =0.001,0.006),and the difference between the AHB group and the HCs group was not statistically significant (P=0.669).Breg in the CHB group [14.30％ (10.70％-16.70％)] was higher than that in the AHB group [10.30％ (7.05％-13.30％)] and the HCs group [10.40％(6.85％-12.60％)] (P =0.003,0.001),treg in the CHB group [5.80％ (4.20％-9.10％)] was also higher than that in the AHB group [4.05％ (2.53％-5.40％)] and the HCs group [4.50％ (2.55％-5.50％)] (P ＜0.001,P =0.005),and there was no significantly difference between the AHB group and the HCs group (Breg:P =0.796 ; Treg:P =0.227).Spearman correlation analysis showed that Breg was positively correlated with Treg in the CHB group (r =0.50,P =0.004),however there was no significantly correlation in the AHB group and the HCs group (r =-0.15,P =0.462; r =0.09,P =0.669).The concentration of IL-10 in the CHB group was higher than that in the AHB group and the HCs group (P ＜ 0.001),and the difference between the AHB group and the HCs group was not statistically significant (P=0.341).Spearman correlation analysis showed that IL-10 were positively correlated with the population of Breg in the CHB group (r =0.409,P =0.022).Conclusion The poluations of regulatory B cell and regulatory T cell increased in patients with chronic hepatitis B,and Breg cell might play the immune regulation role through secreting IL-10 in chronic HBV infection.
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