CRIM1在肝细胞癌中的表达及其与上皮-间质转化关系的初步研究

Relationship of CRIM1 and epithelial-mesenchymal transition in hepatocellular carcinoma

目的 探讨半胱氨酸丰富跨膜成骨蛋白调控因子1(CRIM1)在肝细胞癌(简称肝癌)中的表达及临床意义,初步探讨CRIM1表达水平与上皮细胞-间质转化(EMT)之间的关联性。方法 回顾性分析2013年1月—2017年12月扬州大学附属苏北人民医院收治的114 例肝癌手术患者的临床病理资料,收集患者的术后肝癌组织和对应癌旁组织病理学标本,石蜡切片包埋,并行免疫印迹试验(Western blotting)及免疫组织化学染色检测,观察指标:(1)肝癌组织和癌旁组织中CRIM1蛋白及EMT相关蛋白(E-钙黏蛋白、波形蛋白)表达情况;(2)肝癌组织中CRIM1蛋白表达与患者临床病理因素的关系。运用Western blotting检测肝癌患者癌组织、癌旁组织中CRIM1及EMT相关蛋白(E-钙黏蛋白、波形蛋白)表达水平情况,采用t检验方法进行灰度值分析;采用免疫组化检测肝癌组织和癌旁组织中CRIM1蛋白表达情况,根据组化评分将其分为高表达组、低表达组,并用χ2检验和Spearman相关性分析方法分析CRIM1蛋白的不同表达与患者临床病理因素的关系。最后通过Kaplan Meier Plotter数据库分析CRIM1与肝癌患者生存率之间的关联性。 结果 Western blotting显示肝癌组织和癌旁组织中CRIM1蛋白表达灰度值分别为0.15±0.03、0.80±0.04,E-钙黏蛋白表达水平分别为0.20±0.05、0.56±0.06,两者在癌旁组织中表达水平显著高于癌组织(t=14.21、4.69,P<0.05),而波形蛋白在肝癌组织和癌旁组织中表达水平分别为0.74±0.08、0.45±0.06,在癌组织中表达水平显著高于癌旁组织(t=2.87,P<0.05)。免疫组化进一步验证肝癌组织及癌旁组织中CRIM1表达水平,114 例患者中肝癌组织中,46 例CRIM1蛋白高表达,68 例CRIM1蛋白低表达,CRIM1的表达与患者甲胎蛋白水平、肿瘤大小、有无症状具有关联(r=-0.43、-0.34、-0.24, χ2=9.38、5.25、8.12,P<0.05)。Kaplan Meier Plotter数据库分析表明,CRIM1高表达与低表达两者生存情况差异有统计学意义(P=0.04)。结论 在肝癌中CRIM1的表达与肿瘤EMT过程呈负关联,CRIM1有望成为判断肝癌预后的潜在分子标志物。

Objective To study the expression and the clinical significance of cysteine rich transmembrane BMP regulator 1(CRIM1) in hepatocellular carcinoma(HCC) and discuss the association between CRIM1 and epithelial-mesenchymal transition(EMT). Methods The cases were came from the Subei People′s Hospital Affilated Hospital of Yangzhou University from January 2013 to December 2017. CRIM1 and EMT related proteins (E-cadherin, Vimentin) in parts of HCC tissues and their paired peritumoural tissues were tested by Western blotting. The gray value was test by t test. The observation indicators: (1) expression of CRIM1 protein and EMT-related protein (E-cadherin, Vimentin) in liver cancer tissues and paracancerous tissues. (2) The relationship between CRIM1 protein expression and clinicopathological factors in patients with liver cancer. The expression of CRIM1 in HCC tissues and adjacent tissues was detected by immunohistochemistry(IHC), which was divided into high expression group and low expression group according to the histochemical score, and the relation between the expression of CRIM1 and the clinicopathological factors of the patients was analyzed by chi-square test and Spearman correlation analysis. Finally, the relation between CRIM1 and overall survival of HCC patients was analyzed by Kaplan Meier Plotter database.Results The expression of CRIM1 in tumor and matched paratumor specimens were 0.15 ± 0.03, 0.8 ± 0.04, and E-cadherinin tumor and matched paratumor specimenswere 0.20±0.05, 0.56±0.06, their expression in paracancerous tissues was higher than HCC tissues(t=14.21, 4.69, P<0.05),while the expression of Vimentin in tumor and matched paratumor specimens were 0.74 ± 0.08, 0.45 ± 0.06, the expression in tumor tissues were significantly higher than adjacent tissues(t=2.87, P<0.05). The expression of CRIM1 in HCC tissues was further verified by immunohistochemistry, which shows that CRIM1 was overexpressed in paracancerous tissues. In 114 patients, 46 cases of CRIM1 protein were highly expressed in liver cancer tissues, and 68 cases of CRIM1 protein were low expressed. The expression of CRIM1 obviously related with the level of α-fetoprotein (AFP), tumor size and symptom(r=-0.43, -0.34, -0.24, χ2=9.381, 5.248, 8.117, P<0.05). However, other clinicopathological features were not correlated with CRIM1 expression, including age, tumour differentiation, tumor number. Finally, the overall survival was different between CRIM1 high expression and low expression according to the Kaplan Meier Plotter database. Conclusions The expression of CRIM1 is negatively correlated with the EMT process in HCC. CRIM1 might be a potential molecular marker for prognosis.