Objective To investigate the significance of changes in expression of co-stimulatory molecules on T lymphocytes in patients with chronic hepatitis B (CHB) infection.Methods In a casecontrol study,a total of 82 CHB cases including 50 male cases and 32 female cases (the mean age was 42.32 ±3.74) were enrolled in the First Affiliated Hospital to Soochow University from October 2012 to November 2013,together with 30 cases health control (15 male cases and 15 female cases,and the mean age was 42.32 ± 3.74).Patients were divided into three groups:40 cases of non-treated,30 cases of effective-treated and 12 cases of ineffective-treated with anti-viral drugs and immune-related therapy.The expression levels of CD28,CTLA-4,PD-1,Tim-3 on T cells subset from peripheral blood were determined by Flow Cytometry.Serum load of HBV DNA was detected by Real-time PCR and the serology markers such as HBeAg and ALT were detected by conventional methods The Kruskal-Wallis test was used to analysis groups comparison.Independent samples t-test and Mann-Whitney U test were used to two sample comparison.Spearman's rank correlation test was used to analyze the correlation.Results The expression levels of CD28,CTLA-4 and PD-1 on CD4 in health control group was the highest compared to ineffectivetreated group [404.65 (331.65-536.09) vs 277.15 (249.90-344.25) (H=29.81,P＜0.001);32.89 (29.69-39.69) vs 19.26 (11.90-20.56) (H =43.13,P ＜0.001),respectively].The expression level of PD-1 on CD8 in pre-treated group was the highest,while expression level in health control group was the lowest [15.47 (12.50-17.78) vs 3.05 (1.41-3.97) (H=56.60,P＜0.001)].Similarly,the expression level of Tim-3 on CD4 in ineffective-treated group was the highest,while Tim expression on CD4 in health control group was the lowest [199.62 (55.61-239.45) vs 70.62 (53.88-112.32) (H =41.03,P ＜ 0.001)].The expression level of Tim-3 on CD8 in pre-treated group was the highest,while it was the lowest in health control group [82.50 (78.69-84.58) vs 3.07 (1.56-6.87) (H=74.84,P ＜0.001)].Compared to the group with low viral load,the expression level of PD-1 on CD8 both in the pre-treated group with high viral load and the post-treated group was significantly increased [17.87 (13.38-20.94) (U=25.00,P＜0.001); 16.95 (5.39-18.27) (U=63.50,P＜0.001),respectively].Importantly,in the post-treated group,the PD-1 expression on CD4 T (r =0.689,P ＜0.001) and on CD8 T (r =0.751,P ＜ 0.001) was also positively correlated with ALT.Conclusions The abnormal expression of these co-stimulatory molecules may be present in the antiviral treatment process of CHB.It may provide new clues for the reasonable treatment of CHB.%目的 探讨慢性乙型肝炎患者外周血T淋巴细胞表面共刺激分子的表达量变化及其意义.方法 临床病例对照研究.收集2012年10月至2013年1 1月在苏州大学附属第一医院感染科住院的慢性乙型肝炎患者82例,其中男50例,女32例,年龄(42.25±14.22)岁,健康对照组30名来自本院预防保健科,其中男15例,女15例,年龄(42.32±3.74)岁；采用流式细胞术和定量PCR等技术测定慢性乙型肝炎患者外周血T淋巴细胞表面CD28、CTLA-4、PD-1、Tim-3的表达、T淋巴细胞亚群(CD4+、CD8+、CD4 +/CD8+),其中未经抗病毒及相关免疫治疗的有40例,经过抗病毒治疗有效的有30例,无效的12例,并结合HBV病毒载量、乙型肝炎e抗原和丙氨酸转氨酶等指标作综合分析.组间比较采用单因素方差分析和Kruskal-Wallis H检验,两样本均数的比较采用t检验和Mann-Whitney U检验,相关分析用Spearman'S检验.结果 CD4+ CD28+组间表达量健康对照组最高[404.65(331.65～536.09)],治疗无效组最低[277.15(249.90～344.25)],差异有统计学意义(H=29.81,P＜0.001)；CD4+ PD-1+组间表达量未治疗组最高[32.89 (29.69 ～ 39.69)],治疗无效组最低[19.26(11.90 ～20.56)],差异有统计学意义(H =43.13,P＜0.001)；CD8+ PD-1+组间表达量未治疗组最高[15.47(12.50 ～17.78)],健康对照组最低[3.05(1.41 ～ 3.97)],差异有统计学意义(H=56.60,P＜0.001)；CD4+ Tim-3+组间表达量治疗无效组最高[199.62(55.61 ～239.45)]、健康对照组最低[70.62(53.88 ～ 112.32)],差异有统计学意义(H=41.03,P＜0.001)；CD8+ Tim-3+组间表达量未治疗组最高[82.50(78.69 ～84.58)],健康对照组最低[3.07(1.56 ～6.87)],差异有统计学意义(H=74.84,P＜0.001)；与低病毒载量组相比,治疗前CD8+ PD-1+在高病毒载量组中的表达量17.87(13.38 ～20.94)]升高,差异有统计学意义(U=25.00,P＜0.001),治疗后CD8+ PD-1+在高病毒载量组中的表达量[16.95(5.39 ～ 18.27)]升高,差异有统计学意义(U=63.50,P＜0.001)；治疗后PD-1表达量与ALT也有明显的正相关性,其中CD4+ PD-1+(r=0.689,P＜0.001)、CD8+ PD-1+(r =0.751,P＜0.001).结论 在慢性乙型肝炎患者抗病毒治疗过程中,可能伴随有CD28、PD-1、CTLA-4和Tim-3的异常表达.共刺激分子的检测可为临床合理治疗慢性乙肝提供新线索.
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