慢性乙型肝炎患者外周血调节性B细胞与调节性T细胞的表达频率及其相关性

Expression and roles of regulatory B cells and T cells in patients with chronic hepatitis B

目的:分析慢性乙型肝炎患者外周血IL-10+CD19+调节性B细胞(regulatory B cells,Breg)与CD4+CD25high调节性T细胞(regulatory T cells,Treg)的比例及两者的相互作用。方法:收集急性乙肝(acute hepatitis B,AHB)患者28例,慢性乙型肝炎(chronic hepatitis B,CHB)患者31例和健康体检者(健康组)25例。分离外周血单个核细胞(peripheral blood mononuclear cells,PBMC),经CpG ODN 2006和佛波酯体外作用后,流式细胞术分析Breg、Treg的表达及其相关性,ELISA检测培养上清IL-10水平。通过免疫磁珠剔除CD4+CD25+T细胞后分析Breg与Treg表达频率间的相关性。结果:CpG ODN 2006和佛波酯作用后CHB组Breg和Treg百分率较AHB组和健康组均明显升高(Breg:P值分别为0.003和0.001;Treg:P值分别为<0.001,0.005),AHB组和健康组间两类细胞表达频率差异无统计学意义(Breg:P=0.796;Treg:P=0.227);Spearman相关性分析显示CHB组Breg与Treg表达百分比存在正相关关系(r=0.50,P=0.004),而AHB组和健康组无此相关性(r=-0.15,P=0.462;r=0.09,P=0.669)。CHB组IL-10表达高于AHB组和健康组(Z值分别为-6.596、-6.373,P均<0.001),AHB组和健康组间差异无统计学意义(Z=-0.952,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=3.303,P=0.005)。CHB组患者Breg和IL-10的表达在剔除CD4+CD25+T细胞前后间的差异无统计学意义。结论:Breg和Treg在CHB患者中表达频率升高,Breg可能通过IL-10的分泌参与慢性乙肝的免疫调节,且该作用不依赖于Treg。

Objective： To investigate the expression and roles of IL-10＋ CD19＋ regulatory B cells （Breg） and CD4 ＋ CD25^high regulatory T cell（Treg） in patients with chronic hepatitis B. Methods： A total of 28 patients with acute hepatitis B （AHB）, 31 patients with chronic hepatitis B （CHB） and 25 normal subjects（HCs group） were enrolled in this study. Peripheral blood mononuclear cells （PBMC） were isolated and stimulated with CpG ODN 2006 and PMA. The expression of IL-10 ＋ CD19 ＋ Breg and CD4 ＋ CD25high Treg was analyzed with flow cytometry. The level of IL-10 in culture supernatant was analyzed with ELISA. CD4＋ CD25 ＋ T cells were removed with Immunomagnetic beads to analyze the interactions between Breg and Treg. Results： After stimulation with CpG ODN 2006 and PMA, compared with the AHB group and the HCs group, percentages of Breg and Treg in the CHB group increased significantly （ Breg ：P = 0. 003,0.001 ;Treg：P 〈0.001, 〈0.005） ,and no difference was found between the AHB group and the HCs group （Breg ：P =0.796; Treg： P = 0.227 ）. Spearman correlation analysis found that in the CHB group, Breg and Treg was positively correlated （r =0.50,P =0. 004） ,but in the AHB group and the HCs group,there was no such correlation （r = -0.15,P =0.462;r =0.09,P =0. 669）. IL-10 expression in the CHB group was higher than that in the AHB group and the HCs group（Z = -6.596, -6.373,P 〈0. 001 ）, and the difference between the AHB group and the HCs group was not statistically significant （Z = -0.952, P = O. 341 ）. Spearman correlation analysis showed that in the CHB group, IL-10 and Breg were positively correla- ted （r=3.303 ,P =0. 005）. Before and after removal of CD4＋ CD25 ＋T cells ,in the CHB group, the difference between Breg and IL-10 expression was not statistically significant. Conclusion： The expression of Breg and Treg increased in patients with chronic hepatitis B, Breg might play a regulatory role by IL-10 secretion.