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目的:探讨α1-抗胰蛋白酶(α1-antitypsin,α1-AT)基因第5外显子和组织因子途径抑制物(tissue factor pathway inhibitor,TFPI)基因T287C多态性的交互作用及其与大肠癌侵袭和转移的关系。方法:选择河南省新乡医学院第一附属医院2011年7月至2015年7月期间收治的经病理学确诊的大肠癌TNMⅠ、Ⅱ、Ⅲ和Ⅳ期患者各180例,以180例TNM 0期患者作为对照组,用PCR-RFLP技术检测各组患者外周血白细胞两基因的多态性,以Hardy-Weinberg平衡检验分析样本的群体代表性,以Khoury和Wagener模型分析两基因多态性的交互作用。ELISA法检测患者血清α1-AT和TFPI蛋白表达,细胞划痕试验和Transwell侵袭试验分别检测α1-AT和TFPI对人结肠癌SW480细胞迁移和侵袭能力的影响,Western blotting检测SW480细胞胰蛋白酶、组织因子(tissue factor,TF)和蛋白酶激活受体-2(protease-actieated receptor 2,PAR-2)表达水平。结果:α1-AT(MZ)、α1-AT(ZZ)、TFPI(TC)和TFPI(CC)基因型者大肠癌侵袭与转移的风险均显著增加,且在α1-AT(MZ)和TFPI(TC)之间、α1-AT(MZ)和TFPI(CC)之间、α1-AT(ZZ)和TFPI(TC)及α1-AT(ZZ)和TFPI(CC)之间均存在正向交互作用(均γ>1)。Ⅰ、Ⅱ、Ⅲ和Ⅳ期患者血清α1-AT(或TFPI)表达明显低于0期组,且Ⅰ、Ⅱ、Ⅲ和Ⅳ期患者之间血清α1-AT(或TFPI)表达也有明显差异(P<0.01)。同一组携带突变基因型个体的血清α1-AT(或TFPI)表达明显低于携带野生型个体(P<0.01)。体外细胞培养实验显示,α1-AT可明显抑制SW480细胞胰蛋白酶的表达,而TFPI则明显抑制TF表达,而两者均可明显降低细胞PAR-2的表达及细胞迁移和侵袭能力。结论:α1-AT(MZ)、α1-AT(ZZ)、TFPI(TC)和TFPI(CC)基因型均是大肠癌侵袭与转移的高危因素,基因多态性的交互作用增加了大肠癌侵袭与转移的风险。
Objective: To investigate the interaction between exon 5 of α1-antitypsin (α1-antitypsin, α1-AT) gene and T287C polymorphism of tissue factor pathway inhibitor (TFPI) gene and its relationship with colorectal cancer Invasion and metastasis of the relationship. Methods: 180 cases of TNMⅠ, Ⅱ, Ⅲ and Ⅳ pathologically diagnosed colorectal cancer admitted to the First Affiliated Hospital of Xinxiang Medical College from July 2011 to July 2015 were enrolled. 180 cases of TNM stage 0 Patients as control group, using PCR-RFLP technology to detect peripheral blood leukocyte two gene polymorphism in each group, Hardy-Weinberg equilibrium test to analyze the sample population representative, Khoury and Wagener model analysis of two gene polymorphism interaction effect. The expression of α1-AT and TFPI in serum of the SW480 cells was detected by ELISA. The effects of α1-AT and TFPI on the migration and invasion of human colon cancer SW480 cells were detected by cell scratch assay and Transwell invasion assay respectively. Western blotting was used to detect the expression of trypsin, The expression of tissue factor (TF) and protease-actieated receptor 2 (PAR-2) Results: The risk of invasion and metastasis of colorectal cancer were significantly increased in α1-AT (MZ), α1-AT (ZZ), TFPI (TC) and TFPI (CC) TC), there was a positive interaction between α1-AT (MZ) and TFPI (CC), between α1-AT (ZZ) and TFPI (TC) (All γ> 1). The serum levels of α1-AT (or TFPI) in patients with stage Ⅰ, Ⅱ, Ⅲ and Ⅳ were significantly lower than those in stage 0, and there was also significant difference in the expression of serum α1-AT (or TFPI) between patients with stage Ⅰ, Ⅱ, P <0.01). The expression of α1-AT (or TFPI) in the same group of individuals with mutant genotypes was significantly lower than that in wild-type individuals (P <0.01). In vitro cell culture experiments showed that α1-AT can significantly inhibit the expression of trypsin in SW480 cells, while TFPI significantly inhibited the expression of TF, both of which can significantly reduce the cell PAR-2 expression and cell migration and invasion ability. CONCLUSION: The genotypes of α1 -AT, Z1, TFPI and TFPI are both risk factors of invasion and metastasis of colorectal cancer. The interaction of gene polymorphisms increases the invasion of colorectal cancer With the transfer of risk.