慢性乙型肝炎患者外周血CD8＋T淋巴细胞亚群T淋巴细胞受体β链互补决定区3谱型研究

Characteristics of CDR3 of TCR β on CD8＋ T cells in chronic hepatitis B patients

目的研究慢性乙型肝炎（CHB）患者外周血CD8＋T淋巴细胞亚群T淋巴细胞受体（TCR）p链V区（BV）中互补决定区3（CDR3）谱型特点。方法采集8例CHB患者（ALT〉2倍正常值上限）的抗凝血，分离外周血单个核细胞，磁珠分选CD8＋T淋巴细胞亚群，提取RNA并应用逆转录聚合酶链反应扩增TCRBV中CDR3基因的24个家族，采用免疫指纹技术进行TCRBV各家族基因扫描和谱型分析。不同细胞亚群间数据比较采用配对t检验。结果8例患者外周血淋巴细胞TCRBV家族CDR3谱型发生明显偏移，表现为单克隆性、寡克隆性及偏峰性克隆增生。8例患者CD8＋T淋巴细胞亚群发生TCRBVCDR3谱型偏移家族总个数高于去除CD8。T淋巴细胞群的外周血单个核细胞[（10．6±4．7）个比（4．1±3．1）个，t=6．619，P〈0．01）]；比较单、寡克隆增生2种形式偏移的家族，CD8＋T淋巴细胞亚群也高于代表CD4＋T淋巴细胞的去除CD8＋T淋巴细胞群的外周血单个核细胞[（8．8±4．5）个比（3．9±2．8）个，t=5．706，P〈0．01]。对其中3例患者磁珠分选前后TCRBV谱型的比较发现，分选后CD8＋T淋巴细胞亚群发生TCRBV谱型偏移家族数均高于分选前。结论通过淋巴细胞亚群分选方式分析TCRBV家族CDR3谱型变化可以减少不同淋巴细胞亚群之间的峰型重叠干扰；应用这一方法分析外周血CD8＋T淋巴细胞亚群克降增牛程度有助于了解CHB患者炎疖的发牛机制．

Objective To analyze the characteristics of CDR3 of TCRβ on CD8＋ T cells in chronic hepatitis B patients. Methods Eight patients with chronic hepatitis B （ALT 〉 2 ULN） were enrolled in this study. CD8＋ T cells were isolated from peripheral blood. RT-PCR was proformed to amplify the CDR3 of TCRβ, and the PCR products were sequenced and analyzed. Results The chronic hepatitis B patients showed obvious clonal expansion of T cell, and three perturbation patterns of T cell expansion were showed in the CDR3 of TCR β, including monoclonicity, oligoclonicity and skewed peak patterns. The number of perturbation families of CD8＋ subpopulation was significantly higher than that of CD8- subpopulation （10.6 ± 4.7 vs. 4.1± 3.1, t = 6.619, P 〈 0.01）. In 3 out of 8 patients, the number of perturbation families of CD8＋ subpopulation was also higher than that of PBMCs without depleting CD8＋ subpopulation. Conclusion The characteristics of CDR3 of TCR＋ may help to understand the inflammatory response in CHB patients.