肺泡灌洗液中淋巴细胞表型的测定及临床意义

Immunophenotyping of lymphocytes in bronchoalveolar lavage fluid by flow cytometer

目的 建立以流式细胞仪 (FCM)检测肺泡灌洗液 (BALF)中淋巴细胞表型的方法并对其临床意义进行研究。方法 采用FITC CD45 /PE CD14设门并结合PI染色排除非淋巴细胞、细胞碎片等对检测的干扰 ,以FCM分析 43例肺部疾病患者BALF和对照组外周血中淋巴细胞上各抗原的表达 ,并与免疫组化检测方法进行方法学比较。结果 FCM法与免疫组化法相关良好 (r≥ 0 .80 6 ) ,其精密度 (CV≤ 1.0 % )远优于免疫组化法 (CV≥ 9.8% )。 43例BALF中T细胞亚群较外周血有明显变化(P <0 .0 1) ,并且在某些肺部疾病中有独特的改变。结节病 :细胞表面分化抗原 (CD3)和CD4细胞明显增多 ,CD8细胞数目明显下降 ,CD4/CD8比值升高 ;特发性肺间质纤维化 (IPF) :CD8细胞显著增多 ,CD4细胞显著降低 ,CD4/CD8比值明显降低 ;肺部肿瘤 :CD3细胞数目无明显增多 ,但CD4/CD8比值下降 ;肺结核患者T细胞亚群在正常值范围之内 ,BALF和外周血之间差异无显著意义 (P >0 .0 5 )。结论FCM法准确快速、并可进行多参数分析 ,适合成分复杂、常混有痰液和杂质的BALF标本的检测 ;BALF中T细胞亚群的改变对肺部疾病的鉴别诊断有重要意义。

Objective To establish a tri color flow cytometric approach to immunophenotyping of lymphocytes in bronchealveolar lavage fluid (BALF).Methods We used FITC CD45/PE CD14 antibodies to gate lymphocytes and exclude other contamination. Propidium Iodide (PI) was introduced to differ lymphocytes from debris. 43 BALF were tested by flow cytometery as well as by immunoperoxidase method. Their peripheral bloods were also tested as control group by two methods.Results The variation of FCM ( CV ≤1.0%) was much lower that that of PAP method ( CV ≥9.8%). Meanwhile BALF was more clinicaly significant than peripheral blood in T subset analysis ( P <0.01). For BALF, both CD3 and CD4 were significantly increased with decreasing CD8 in sarcoidosis ( n =14). Idiopathic pulmonary fibrosis ( n =16) demonstrated the reverse tendency: CD8 rose but both CD3 and CD4 dropped. As for lung cancer ( n =7), CD3 was normal but CD4/CD8 ratio declined. Tuberculosis of lungs ( n =6) showed a normal CD3, CD4 and CD8.Conclusion The high precision and reliability of flow cytometric approach to immunophenotyping of lymphocyte subsets in BALF suggested that it should be used as a routine test, especially for the BALF which is often contaminated by inorganic particles.