恶性胸腔积液患者T淋巴细胞亚群的变化意义(一)

详细内容

作者：王英禹，张瑞，张志培，周勇安，孙晓雁

【关键词】 胸腔积液；T淋巴细胞；CD4；CD8

　　Value of measurement of Tlymphocyte subsets in patients with malignant pleural effusions

　　【Abstract】 AIM: To measure Tlymphocyte subsets (TLS) in peripheral blood (PB) and pleural effusion (PE) in patients with malignant pleural effusions (MPE) and to study the changes of TLS in PB and PE in patients with MPE before and after chemotherapy. METHODS: The levels of TLS in PB and PE in patients with MPE or benign pleural effusion (BPE) and in PB of normal person were examined and pared; The levels of TLS in PB and PE were also pared before and after chemotherapy. RESULTS: The distribution of TLS was similar in PB and PE in patients with MPE; the level of CD8+ T cell in PB of patients with MPE was higher than that in normal person (30.6±2.4 vs 20.9±1.8, P0.05), but the ratio of CD4+/CD8+ was lower than that in normal person (1.78±0.21 vs 2.71±0.35, P0.05), and the level of CD4+ T cell was similar with the normal; pared with BPE patients, MPE ones had lower level of CD4+ T cell and ratio of CD4+/CD8+ (P0.05), but higher level of CD8+ T cell in their PB and PE (P0.05); chemotherapy could depress CD8+ T cell (30.6±2.4 vs 24.7±2.3, 31.6±2.4 vs 21.9±4.1, P0.05) and elevate the ratio of CD4+/CD8+ (1.78±0.21 vs 2.19±0.19, 1.69±0.30 vs 2.62±0.40, P0.05) in PB and PE in patients with MPE. CONCLUSION: The level of TLS in PE can be known by measuring it in PB; it is helpful to measure the level of TLS in PB and PE for distinguishing MPE from BPE; cellmediated immunity depression in MPE can be partly ameliorated by chemotherapy.

　　【Keywords】 pleural effusion； Tlymphocyte； CD4； CD8

　　【摘要】 目的：探讨检测恶性胸腔积液(MPE)患者外周血(PB)和胸水(PE)中T淋巴细胞亚群(TLS)的意义及化疗前后TLS的变化. 方法：对比良性胸腔积液(BPE)、MPE患者外周血和胸水及健康正常人外周血中TLS的水平；对比MPE患者外周血和胸水中TLS的水平；对比MPE患者化疗前、后外周血和胸水中TLS的水平. 结果：TLS在胸腔积液患者外周血和胸水中分布相似；MPE患者外周血中CD8+ T细胞水平高于正常人（30.6±2.4 vs 20.9±1.8，P0.05），CD4+T细胞相仿，CD4+/CD8+低于正常（1.78±0.21 vs 2.71±0.35，P0.05）；与BPE患者相比，MPE患者外周血和胸水中CD4+T细胞、CD4+/CD8+较低（P0.05），而CD8+T细胞较高（P0.05）；化疗后MPE患者外周血和胸水中CD8+T细胞水平明显降低（30.6±2.4 vs 24.7±2.3，31.6±2.4 vs 21.9±4.1， P0.05），CD4+/CD8+明显升高（1.78±0.21 vs 2.19±0.19，1.69±0.30 vs 2.62±0.40，P0.05）. 结论：检测外周血可以了解胸水中TLS水平；检测外周血和胸水中TLS水平有助于BPE和MPE的鉴别；化疗可部分改善MPE患者的细胞免疫抑制状态.

　　【关键词】 胸腔积液；T淋巴细胞；CD4；CD8

　　0引言

　　肿瘤的发生、发展与细胞免疫功能降低密切相关, T淋巴细胞在其中起中心调控作用〔1〕. 为比较良性胸腔积液（benign pleural effusion, BPE）和恶性胸腔积液(malignant pleural effusion, MPE)患者的T淋巴细胞亚群(TLS)分布水平及对原发病的诊断价值，和化疗对其影响，我们选择BPE，MPE患者的胸腔积液和外周血及正常健康人外周血进行TLS检测.

　　1对象和方法

　　1.1对象诊断明确MPE 32(男21，女11)例，平均年龄57.9岁，鳞腺癌4例，腺癌28例；BPE (结核性) 29(男17，女12)例，平均年龄45.7岁；正常对照组选自体检合格、肝、肾、肺功能完全正常健康人27(男14，女13)例，平均年龄34.2岁. 患者于治疗前行胸腔穿刺术，收集胸腔积液，同时抽取静脉血10 mL. MP患者经NP方案化疗2个周期后（盖诺25 mg/m2 iv d1，d8；DDP 40～50 mg d1～3，21 d为一周期），收集胸腔积液，同时抽取静脉血10 mL. 采集体检正常健康人静脉血10 mL.

　　1.2方法胸腔积液用200 μm尼龙网过滤组织于离心管；加入5 mL RPMI；18℃，200 g离心10 min，去除上清液，混悬细胞沉淀于20 mL RPMI，重复离心1次，重混悬细胞沉淀于5 mL RPMI, 加入PBS冲洗细胞，室温下，200 g离心10 min，去除上清液. 重复此步骤. 混悬细胞沉淀于适量RPMI备用. 用台盼蓝计数活细胞百分率. 将肝素化的新鲜去红细胞混悬液室温下加入等量PBS混匀，每10 mL去红细胞混悬液/PBS混悬液中加入3 mL FicollHypaque液. 18℃，900 g，离心30 min. 移除上层的血浆和血小板，缓慢的转移中层的单个核细胞层到锥形管，用3倍体积的HBSS洗涤细胞，18℃，490 g，离心10 min. 去除上清液. 用HBSS重新混匀细胞、洗涤并离心1次，去除上清液. 混悬细胞于RPMI备用. 用台盼蓝计数活细胞百分率. 计数外周血和胸腔积液淋巴细胞，用RPMI调整细胞度至2.5×109/L，将l06淋巴细胞悬于100 μL Hanks缓冲液(含1 g/L牛血清蛋白、0.1 g/L叠氮钠)中，分别与20 μL抗CD3APC(UCHT1)、抗CD4FITC(RPAT4)及抗CD8Cychrome(RPAT8) mAb或相应IgG对照蛋白(均购自美国BD Pharmingen公司)于4℃孵化30 min，Hanks缓冲液洗涤2次，5 g/L多聚甲醛固定. 24 h内用FACStar Plus流式细胞仪检测CD3，CD4，CD8细胞，计算CD4/CD8比值.
　　
　　统计学处理：用SPSS 10.0统计软件，数据以x±sx表示. 组间比较用MannWhitney U检验. 组内前后比较用Wilcoxon signedrank检验.