磁激活细胞分选结合荧光激活细胞分类检测胃癌骨髓微转移的临床价值

Detecting Bone Marrow Micrometastasis of Gastric Cancer by Magnetic Activated Cell Sorting Combined with Fluorescent Activated Cell Sorting

背景与目的:检测胃癌患者骨髓微转移有多种方法,但不同的方法差异很大。在乳腺癌中,以磁激活细胞分选(m agnetic activated cellsorting,M ACS)结合荧光激活细胞分类(fluorescentactivated cellsorting,FACS)检测血细胞中的肿瘤细胞,有较高的敏感性和特异性。本研究探讨应用这一方法检测胃癌骨髓微转移的临床意义。方法:选择2002年12月～2003年6月行手术治疗的胃癌患者35例,提取其骨髓有核细胞,以结合抗细胞角蛋白(cytokeratin,CK)7/8抗体的M ACS微型免疫磁珠、标记异硫氰酸荧光素(fluorescein isothiocyanate,FITC)的抗CK抗体,以及标记叶绿素蛋白的抗CD45抗体标记后,以M S+/RS+型阳性分选柱进行两次肿瘤细胞富集。取富集前后的细胞样本进行FACS检测,将检测结果与各临床病理学参数进行比较。结果:M ACS富集前,仅3例(8.6%)检测到骨髓中存在的微转移细胞;而富集后,有25例(71.4%)患者检测到了骨髓微转移细胞。组织学分级为中分化、低分化和未分化的患者中,肿瘤细胞频数分别为1.4×10-8～2.4×10-5、2.2×10-7～3.7×10-5和4.0×10-6～8.6×10-5,3组比较,有显著性差异(P=0.026)。骨髓微转移状况与肿瘤TNM分期密切相关(r=439,P=0.008),而与肿瘤大小、浸润深度等参数无关。结论:M

BACKGROUND & OBJECTIVE: Several methods are used to detect bone marrow micrometastasis of gastric cancer with different accuracies. In breast cancer, tumor cells in blood can be detected sensitively and specifically by magnetic activated cell sorting (MACS) and fluorescent activated cell sorting (FACS). This study was to investigate the clinical value of this method in detecting bone marrow micrometastasis of gastric cancer. METHODS: Thirty-five patients, who received operation for gastric cancer from Dec. 2002 to Jun. 2003, were selected. Mononuclear cells were separated from their bone marrows. After marked by MACS minibeads conjugated with cytokeratin (CK) 7/8 antibodies, anti-CK-fluorescein isothiocyanate (FITC), and anti-CD45-perdinin chlorophyll protein (PerCP), tumor cells were enriched twice by MS+/RS+ positive separation column. FACS analysis was conducted on these samples before and after MACS enrichment. The results were compared with clinicopathologic parameters. RESULTS: Disseminated tumor cells were detected in bone marrow of 3 samples(8.6%) before MACS enrichment, and 25 samples (71.4%) after enrichment. The frequencies of tumor cells were 1.4×10-8-2.4×10-5, 2.2×10-7 -3.7×10-5, and 4.0×10-6-8.6×10-5 in patients with moderately differentiated, poorly differentiated, and undifferentiated carcinoma, respectively, with significant differences (P = 0.026). Bone marrow micrometastasis positively correlated with tumor TNM stage (P = 0.008), while had no correlation with tumor size, depth of wall invasion, and other clinicopathologic parameters. CONCLUSIONS: MACS combined with FACS may improve detection rate of bone marrow micrometastasis of gastric cancer. The patients with poor differentiation and in advanced TNM stage have more disseminated tumor cells in bone marrow.