摘要
目的 探讨糖酵解核心酶烯醇化酶在甲状腺癌组织的表达及与肿瘤免疫逃逸和增殖的关系.方法 选取来商丘市第一人民医院接受手术切除的79例甲状腺癌标本和对应癌旁组织作为研究对象,采用蛋白质组学分析甲状腺癌组织和癌旁组织差异表达的蛋白质,选取差异最为显著的蛋白烯醇化酶作为研究对象,采用免疫组织化学分析甲状腺癌和癌旁组织中烯醇化酶表达水平;采用免疫荧光分析CD8阳性淋巴细胞浸润比例;采用蛋白质免疫印迹分析癌旁组织和甲状腺癌组织粒酶B(Granzme B)和穿孔素(perforin)表达水平;并皮尔森相关系数分析烯醇化酶表达水平与CD8阳性淋巴细胞浸润、粒酶B(Granzme B)和穿孔素(perforin)表达之间的关系.组间计量数据比较采用t检验.结果 癌旁组织和甲状腺癌组织总共鉴定差异表达蛋白1 329个,其中上调表达738例,下调表达591个.738个上调基因主要参与糖代谢、氨基酸代谢、细胞增殖相关信号通路、细胞凋亡通路和细胞黏附分子和细胞外基质-受体间相互作用.其中糖代谢中烯醇化酶表达差异最为显著,作为本研究的重点.甲状腺癌组织中烯醇化酶表达水平(143.65±21.79)明显高于癌旁组织表达水平(40.54±9.01),差异有统计学意义(t=38.870,P＜0.05).甲状腺癌组织中CD8淋巴细胞浸润比例[(2.49±5.30)％]明显低于癌旁组织表达水平[(5.30±1.24)％],差异有统计学意义(t=18.380,P＜0.05).甲状腺癌组织粒酶和穿孔素表达水平(1.34±0.16、1.18±0.27)明显低于癌旁组织表达水平(0.74±0.12、0.60±0.15),差异有统计学意义(t=26.600、16.810,P＜0.05).烯醇化酶的表达水平与CD8淋巴细胞的浸润呈负相关(r=-0.411,P＜0.05),与粒酶和穿孔素水平也呈负相关(r=-0.325、-0.514,P＜0.05).结论 烯醇化酶在甲状腺癌组织中表达水平显著增加,与CD8淋巴细胞的浸润和杀伤能力成负相关.
Abstract
Objective To investigate the expression of enolase,a core enzyme of glycolysis,in thyroid cancer tissues and its relationship with tumor immune escape and proliferation.Methods A total of 79 cases of thyroid cancer specimens and corresponding adjacent tissues were selected as the research ob-jects.The differentially expressed proteins in thyroid cancer tissues and adjacent tissues were analyzed by proteomics.The most significant difference protein enolase was selected as the research object.The expres-sion level of enolase in thyroid cancer and adjacent tissues was analyzed by immunohistochemistry.The proportion of CD8 positive lymphocyte infiltration was analyzed by immunofluorescence.The expression lev-els of Granzyme B and perforin in adjacent tissues and thyroid cancer tissues were analyzed by Western blotting.The relationship between the expression level of enolase and the infiltration of CD8 positive lym-phocytes,Granzyme B and perforin was analyzed by Pearson correlation coefficient.The t test was used for comparison of measurement data between groups.Results A total of 1 329 differentially expressed pro-teins were identified in adjacent tissues and thyroid cancer tissues,among which 738 were up-regulated and 591 were down-regulated.The 738 up-regulated genes were mainly involved in glucose metabolism,amino acid metabolism,cell proliferation-related signaling pathways,apoptosis pathways,cell adhesion molecules and extracellular matrix-receptor interactions.The difference of enolase expression in glucose metabolism was the most significant,which was the focus of this study.The expression level of enolase in thyroid canc-er tissues(143.65±21.79)was significantly higher than that in adjacent tissues(40.54±9.01,t=38.870,P＜0.05).The proportion of CD8 lymphocyte infiltration in thyroid cancer tissues[(2.49± 5.30)％]was significantly lower than that in adjacent tissues[(5.30±1.24)％,t=18.380,P＜0.05].The expression levels of granzyme and perforin in thyroid cancer tissues(1.34±0.16,1.18±0.27)were significantly lower than those in adjacent tissues(0.74±0.12,0.60±0.15,t=26.600,16.810,P＜0.05).The expression level of enolase was negatively correlated with CD8 lymphocyte infiltration(r=-0.411,P＜0.05),and also negatively correlated with granzyme and perforin levels(r=-0.325,-0.514,P＜0.05).Conclusion The expression level of enolase in thyroid cancer tissues is significant-ly increased,and is negatively correlated with the infiltration and killing ability of CD8 lymphocytes.
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