乳腺癌微转移骨髓播散肿瘤细胞检测方法

Detection of bone marrow disseminated tumor cells in micrometastasis of breast cancer

目的探讨乳腺癌患者骨髓中播散肿瘤细胞(DTC)敏感的免疫细胞化学检测方法。方法首先用人源性乳腺癌细胞株MCF-7作为标本,建立成熟的免疫细胞化学染色方法(DAB法、APAAP法);再用人源性乳腺癌细胞株SK-BR3、MDA-MB 231、SUM1315作为标本,向健康人外周血提取的单核细胞中加入细胞株进行检测,从而验证免疫细胞化学染色方法的敏感性;最后收集可手术乳腺癌患者的骨髓标本进行临床检测。选择细胞角蛋白(CK)作为肿瘤细胞标志物。并对乳腺癌骨髓微转移与TNM分期进行相关性分析。结果建立能检测到MCF-7等乳腺癌肿瘤细胞的免疫细胞化学方法DAB法、APAAP法。DAB法、APAAP法检测肿瘤细胞的敏感度为在1×10^(5)个单核细胞中可以找到1个肿瘤细胞;DAB法检测34例乳腺癌患者骨髓标本,6例患者骨髓中存在DTC细胞,5例可疑存在DTC细胞,其余23例未见到DTC细胞。骨髓微转移阳性组患者的T2、T3及N1、N2、N3分期占比较骨髓微转移阴性组患者明显增高,差异有统计学意义(P<0.05)。结论本研究建立较为可靠的免疫细胞化学检测方法,初步表明可用于检测乳腺癌患者骨髓内有无肿瘤细胞播散,有助于判断患者的预后和指导个体化治疗。

Objective To explore the sensitive immunocytochemical assay for disseminated tumor cells(DTC)in bone marrow of breast cancer patients.Methods First,the human breast cancer cell line MCF-7 was used as the specimen,and a mature immunocytochemical staining method(DAB method,APAAP method)was established.Then the human breast cancer cell line SK-BR3,MDA-MB 231 and SUM1315 were used as the specimen,adding cell lines to the mononuclear cells extracted from the peripheral blood of healthy people for detection to verify the sensitivity of the immunocytochemical staining method.Finally,the bone marrow specimens of patients with operable breast cancer were collected for clinical detection.Cytokeratin(CK)was selected as a tumor cell marker.The correlation between bone marrow micrometastasis and TNM staging of breast cancer was analyzed.Results Immunocytochemical methods(DAB method,APAAP method)were established to detect MCF-7 and other human breast cancer tumor cells.The sensitivity of DAB method and APAAP method for detecting tumor cells was that one tumor cell can be found in 1×10^(5) monocytes;DAB method was used to detect bone marrow samples of 34 patients with breast cancer,and there were DTC cells in the bone marrow of 6 patients,DTC was suspected in the bone marrow of 5 patients,and no DTC was found in the remaining 23 patients.The proportion of T2,T3 and N1,N2 and N3 stages in patients with bone marrow micrometastasis positive group was significantly higher than that in patients with bone marrow micrometastasis negative group(P<0.05).Conclusion In this study,a more reliable immunocytochemical detection method has established,which preliminarily indicates that it can be used to detect the presence of tumor cells spread in the bone marrow of breast cancer patients,and it is helpful to judge the prognosis of patients and guide individual treatment.