结直肠癌及淋巴结组织CEA mRNA表达的实时荧光定量检测

Study of CEA mRNA expression in cancer and lymph nodes tissues with SYBR Green Ⅰ real-time quantitative polymerase chain reaction

目的:建立检测CEA mRNA表达水平的实时荧光定量逆转录聚合酶链反应(FQ-PCR)方法,并探讨结直肠癌及淋巴结组织中CEA mRNA表达的应用价值。方法:应用SYBR Green Ⅰ作为荧光染料,以GAPDH作为内对照,建立检测CEA mRNA的FQ-PCR方法,并对56例结直肠癌组织、淋巴结组织CEA mRNA的表达水平进行检测。结果:CEA mRNA在结直肠癌组织中均表达,但表达水平在不同病理分化程度(P=0.8661)及不同Duke分期(P=0.2986)中无明显变化;在检测的231个淋巴结中,共有137个淋巴结CEA mRNA表达为阳性,表达水平为18.31±36.85,其阳性表达率为59.3%(137/231),明显高于常规病理检测[29.4%(68/231)],χ2=41.7497,P=0.0001;CEA mRNA在Duke C+D期淋巴结中表达水平(32.85±53.49)显著高于Duke A+B期(6.54±14.57),P=0.0001。结论:FQ-PCR方法简便、特异和可靠;比常规病理检查更加敏感,可有效补充常规病理检查的不足,在预测肿瘤微转移方面具有重要的临床应用价值。

OBJECTIVE：To develop a real-time quantitative RT-PCR method for detection of CEA mRNA expression in colorectal cancer and lymph nodes tissues and explore the functions and clinical significance of CEA mRNA in patients with colorectal cancer. METHODS： Using SYBR Green Ⅰ as dye and GAPDH as inner reference, FQ-PCR method was established and the CEA mRNA expression of tumor and lymph nodes tissues in 56 colorectal cancer patients was detected. RESULTS： CEA mRNA expressed in all colorectal cancer tissues, but the expression levels had no significant difference in different Duke stages （P=0.866 1） and pathology differentitation, P=0.298 6. Among all 231 collected lymph nodes, there were 137 lymph nodes that expressed CEA mRNA, and the expressed levels were 18.31±36.85 and the positive ratio was 59.3%（137/231）which was significantly higher than that of the method by the routine pathology 29.4%（68/231）, χ^2=41.749 7, P=0.000 1. The CEA mRNA expressed levels in lymph nodes of Duke C＋D stages （32.85±53.49）were significantly higher than those of Duke A＋B stages （6.54±14.57）, P=0.000 1. CONCLUSION： The electrophoresis and melting curve analysis of gene amplification product show that the FQ-PCR method is simple, specific and reliable, and this method is more sensitive than the routine pathology method and can complement the lack of the routine pathology, and it has an important clinical value in prognosticating tumor micrometastases.