胃癌组织中NLRC5表达的意义及其影响胃癌侵袭转移的可能机制

目的 观察核苷酸结合结构域受体家族含CARD结构域-5(NLRC5)、β-catenin在胃癌组织中的表达情况.分析NLRC5与胃癌临床病理特征及预后的关系.并初步探讨NLRC5与Wnt/β-catenin信号通路间的关系.方法 收集根治性胃癌切除术患者石蜡标本93例,免疫组化检测胃癌组织中NLRC5和β-catenin的表达情况;分析NLRC5的表达与胃癌临床病理特征的关系.采用胃癌细胞株MKN-45和MGC-803分别转染pEGFP-C2-NLRC5重组质粒,qRT-PCR法和Western blot法验证NLRC5转染效率.采用MTT细胞增殖和细胞划痕实验对NLRC5进行功能分析,观察体外实验中NLRC5表达变化对胃癌细胞的生物学行为的影响,采用Western blot法检测Wnt/β-catenin信号通路相关蛋白的变化.结果 NLRC5在93例胃癌标本中阳性表达67例(72.04％),阴性表达26例(27.96％).NLCR5的表达情况与胃癌的TNM分期、淋巴结转移和复发情况密切相关.Kaplan-merier生存分析结果显示NLCR5阴性表达的患者预后好于阳性表达患者(P＜0.05).93例胃癌标本中β-catenin阳性表达63例(67.74％),阴性表达30例(32.26％).Cox多因素回归分析显示,NLRC5和β-catenin的阳性表达、低TNM分期、淋巴结转移是影响胃癌患者预后的独立危险因素.胃癌细胞在过表达NLRC5后,MTT细胞增殖和划痕实验结果显示,细胞的增殖和迁移能力显著增强(P ＜0.05,P＜0.01),Western blot实验结果显示,Wnt/β-catenin信号通路中关键蛋白β-catenin和下游关键靶蛋白C-myc、金属基质蛋白酶-7的表达水平明显提高.结论 NLRC5广泛表达于胃癌组织中,其表达与胃癌的TNM分期和淋巴结转移密切相关.Kaplan-merier生存分析显示:NLRC5高表达是影响胃癌预后的危险因素.Cox多因素回归分析显示:NLRC5高表达是影响胃癌预后的独立危险因素.NLRC5与β-catenin的表达呈正相关性,NLRC5的表达变化影响胃癌细胞的生物学行为和Wnt/β-catenin信号通路的关键蛋白的表达.提示NLRC5可能通过调控Wnt/β-catenin信号通路影响胃癌的浸润和转移.

The significance of NLRC5 expression in gastric carcinoma and its possible mechanism of invasion and metastasis of gastric carcinoma

Objective To observe the expression of nucleotides-binding domain receptor family containing CARD domain-5 (NLRC5) and β-catenin in gastric cancer,analyzc the relationship bctween NLRC5 and clinicopathological features and prognosis of gastric cancer,and preliminary explore the relationship between NLRC5 and Wnt/β-catenin signaling pathway.Methods A total of 93 patients with gastric cancer who underwent radical gastrectomy were enrolled.All patients were diagnosed by immunohistochemical staining to investigate the expression of NLRC5 and β-catenin in gastric carcinoma,and to analyze the relationship between the expression of NLRC5 and the clinicopathological features of gastric cancer.Gastric cancer cell lines MKN-45 and MGC-803 were transfected by pFPFP-C2-NLRC5 recombinant plasmid,qRT-PCR and Western blot were used to verify the transfection efficiency of NLRC5.The effect of NLRC5 expression on the biological behavior was observed by MTT cell proliferation and cell scratches test (P ＜0.05,P ＜0.01),and the changes of related protein in Wnt/β-catenin signaling pathway were detected by Western blot in gastric cancer cells.Results The expression of NLRC5 in 93 cases of gastric cancer was positive in 67 cases (72.04％) and negative in 26 cases (27.96％).The expression of NLCR5 was closely related to the TNM staging,lymph node metastasis and recurrence of gastric cancer.Kaplan-merier survival analysis was performed in gastric cancer patients with different expression of NLCR5.The results showed that patients with NLCR5-negative expression were better than those with positive expression (P ＜ 0.05).The expression of β-catenin was positive in 63 cases (67.74％) and negative in 30 cases (32.26％).Cox multivariate regression analysis showed that the positive expression of NLRC5 and β-catenin,low TNM staging and lymph node metastasis were independent prognostic factors of gastric cancer.MTT and scratches showed that overexpression of NLRC5 could significantly promote the proliferation and migration of gastric cancer cells.Western blot showed that overexpression of NLRC5 could significantly increase the key protein β-catenin and the key downstream of Wnt/β-catenin signaling pathway target protein C-myc,matrix metalloproteinase-7 expression levels.Conclusion NLRC5 is widely expressed in gastric cancer tissues,and its expression is closely related to TNM staging and lymph node metastasis of gastric cancer.Kaplan-merier survival analysis shows that NLRC5 overexpression is a risk factor for prognosis of gastric cancer.Cox multivariate regression analysis shows that NLRC5 overexpression is an independent risk factor for the prognosis of gastric cancer patients.NLRC5 and β-catenin are positively correlated,and the expression of NLRC5 affects the biological behavior of gastric cancer cells and the expression of key proteins in Wnt/β-catenin signaling pathway,suggesting that NLRC5 may affect the invasion andmetastasis of gastric cancer by regulating Wnt/β-catenin signaling pathway.