慢性乙肝患者抗病毒治疗前后特异性T细胞免疫观察

Study on antigen-specific T cells in chronic hepatitis B patients accepting antiviral therapy

目的了解抗病毒治疗前后慢性乙型肝炎患者特异性T淋巴细胞对HBV抗原蛋白免疫应答的变化及其特征。方法收集17例慢性乙型肝炎患者抗病毒治疗前及治疗后1个月、3个月的外周血单个核细胞，以HBV特异性抗原蛋白HBsAg、HBcAg和HBeAg为刺激物，酶联免疫斑点法检测其分泌IFN-γ产生斑点的情况。同时对血清HBVDNA和HBsAg、HBeAg等病毒学指标及谷丙转氨酶（ALT）等生化学指标进行检测并分析其相关性。结果治疗前，所有患者ALT、总胆红素（TBiL）均高于正常上限，17例患者HBVDNA均大于10。拷贝／ml；治疗1个月后，ALT复常率为35．3％，9例患者HBVDNA降为检测下限以下；治疗3个月后，ALT复常率为58．8％，有11例患者HBVDNA降为检测下限以下。抗病毒治疗前、治疗1个月、治疗3个月患者针对HBV特异性蛋白总的T细胞反应阳性率分别为64．7％、76．5％和82．4％，其差别无统计学意义。不论治疗前后，患者对HBcAg的特异性T细胞反应频率和平均反应强度最高；治疗后，对3种蛋白的特异性T细胞反应频率和平均反应强度各有不同程度的增加，其中以对HBcAg蛋白的平均反应强度的增强最明显，治疗前和治疗3个月，治疗1个月和治疗3个月之间的差别都有统计学意义。患者对HBcAg蛋白的特异性T细胞反应平均反应强度与病毒载量有明显负相关，与血清AIJT无明显相关性。结论本研究结果提示抗病毒治疗后，患者对HBV的特异性T细胞免疫应答有所增强，这种改变可能与HBV DNA的下降有关。检测HBV特异性T细胞反应对了解患者的免疫状态有重要的意义。

Objective To explore the responses of antigen-specific T cells stimulated by hepatitis B virus（HBV）-specific proteins in chronic hepatitis B patients accepting antiviral therapy. Methods Seventeen patients with chronic hepatitis B （CHB） accepting antiviral therapy were included in this study. The peripheral blood monocular cell （PBMC） were separated from the whole blood collected at the three different time of before and one and three months after accepting antiviral therapy. ELISPOT assay was used to detect the frequency and strength of secreting IFN-γ cells of PBMC stimulated by HBsAg, HBcAg and HBeAg. HBV virus loading, HBsAg, HBeAg, ALT and AST in serum were detected at the same time. Results After three months therapy, ALT, TBiL were improved in all patients, and HBV DNA level were dropped and undetectable in 11 cases. The rates of T cell response in patients to HBV specific proteins were 64.7%, 76.5% and 82.4% at the time of before and one and three months after accepting antiviral therapy, respectively. The frequency of responses of antigen-specific T cells stimulated by HBcAg was higher than that stimulated by HBsAg or HBeAg, and the frequency was enhanced after antiviral therapy. The average response magnitude was expressed as spot forming cells （SFC） per million input cells. SFC of T cell responses to HBcAg was also higher than to HBsAg or HBeAg. There was no significant difference in SFC of T cell responses to HBsAg or HBeAg at the time of before and after antiviral therapy, but there were sianificant difference in SFC of T cell responses to HBcAg at the time of before and after antiviral therapy. SFC of T cell responses to HBcAg was negatively associated with HBV DNA, and no associated with level of ALT in serum. Conclusion The responses of antigen-specific T cells were improved in CHB patients accepting antiviral therapy which associated with the decrease of HBV DNA. It suggested to investigate HBV specific T cell responses was important.