非小细胞肺癌患者EphB3高表达促进血管生成拟态结构的形成

High expression of EphB3 promotes the formation of vasculogenic mimicry in non-small cell lung cancer

目的研究非小细胞肺癌(NSCLC)患者组织样本中促红细胞生成素产生肝细胞受体B3(EphB3)的表达以及与血管生成拟态(VM)形成之间的相关性。方法收集TCGA数据库中有关EphB3的基因表达数据,对比分析515例肺腺癌组织和59例正常组织、503例肺鳞癌组织和52例正常组织的基因表达差异。利用GEPIA2网站筛选出前100个与EphB3相关相似的基因。在仙桃数据库中进行基因本体(GO)功能注释和京都基因与基因组百科全书(KEGG)通路富集分析。收集56例NSCLC患者组织样本,将样本固定在10%多聚甲醛溶液中并用石蜡包埋,按照SP两步法操作,检测其癌组织及癌旁组织中EphB3表达情况。常规CD31免疫组化染色至DAB显色步骤,向组织上滴加PAS染色溶液,镜下观察血管生成拟态结构。结合患者临床特征与EphB3、VM表达情况进行相关性分析。结果EphB3在肺鳞癌和肺腺癌中的表达量均高于对应的癌旁组织,差异均有统计学意义(P均<0.05)。GO/KEGG功能富集分析结果显示KEGG通路富集到Notch信号通路,GO分析结果显示与细胞间连接、细胞黏附分子结合、表皮细胞分化等相关(P<0.05)。免疫组化结果显示,56份样本中癌组织的面积分数为(13.04±1.77)%,癌旁组织为(3.92±0.61)%(P<0.05),其中有41例呈阳性表达,15例呈阴性表达,阳性表达率为73.21%,阴性表达率为26.79%。EphB3在癌组织中的高表达与年龄、吸烟史、T分期有关(P<0.05)。CD31/PAS双染结果显示,在29例样本中,EphB3阳性表达并存在血管生成拟态11例,EphB3阴性表达并出现血管生成拟态1例,EphB3的表达与血管生成拟态形成有关(P<0.05)。结论非小细胞肺癌中EphB3的高表达会促进血管生成拟态的形成,有望成为非小细胞肺癌抗血管生成治疗的潜在治疗靶点。

Objective To investigate the expression level of erythropoietin producing hepatocellular receptor B3(EphB3)in tissue samples of patients with non-small cell lung cancer(NSCLC)and its correlation with vasculogenic mimicry(VM).Methods The gene expression data of EphB3 in TCGA database were collected,and the differences of expression between 515 cases of lung adenocarcinoma and 59 cases of paracancerous tissues,503 cases of lung squamous cell carcinoma and 52 cases of paracancerous tissues were compared and analyzed.The first 100 genes related to EphB3 were screened by GEPIA2 website.Functional annotation of gene ontology(GO)and enrichment analysis of kyoto encyclopedia of gene and genome(KEGG)pathway were carried out in Xiantao database.Tissue samples from 56 patients with NSCLC were collected and fixed in 10%paraformaldehyde solution and embedded in paraffin.The expression of EphB3 in cancer tissues and adjacent no-cancer tissues was detected according to streptavidin-perosidase(SP)method.From CD31 immunohistochemical staining to DAB staining step,PAS staining solution was added to the tissue,and the VM could be observed under microscope.The correlation between clinical characteristics and the expression of EphB3 and VM in patients was analyzed.Results The expression level of EphB3 in lung squamous cell carcinoma and lung adenocarcinoma was higher than that in the adjacent nocancer tissues(P<0.05).The results of GO/KEGG functional enrichment analysis showed that KEGG pathway was enriched in Notch signal pathway,and GO analysis showed that it was related to cell-cell junction,cell adhesion molecular binding,epidermal differentiation(P<0.05).The results of immunohistochemistry showed that the area fraction of cancer tissue and paracancerous tissue were(13.04±1.77)%and(3.92±0.61)%(P<0.05),respectively.Among the 56 samples,41 cases were positive,15 cases were negative,the positive expression rate was 73.21%,and the negative expression rate was 26.79%.The high expression level of EphB3 in cancer tissues was related to age,smoking and T stage(P<0.05).The results of CD31/PAS staining showed that among the 29 samples,11 cases showed positive expression of EphB3 with VM,while only 1 case showed negative expression of EphB3 with VM.The expression of EphB3 was related to the formation of VM(P<0.05).Conclusion The high expression of EphB3 in non-small cell lung cancer could promote the formation of vasculogenic mimicry,which was expected to be a potential therapeutic target for anti-angiogenic therapy in non-small cell lung cancer.