KRT6A调控胰腺导管腺癌细胞生物学行为的作用及作为诊断与预后判断靶标的研究

The role of KRT6A in regulating the biological behavior of pancreatic ductal adenocarcinoma PANC1 cells and its role as a target for diagnosis and prognosis

目的:通过生物信息学分析以及细胞生物学实验研究角蛋白6A(KRT6A)对胰腺导管腺癌(PDAC)诊断、预后判断、免疫微环境以及PDAC细胞PANC1增殖、凋亡等生物学行为的影响。方法:通过GEPIA平台整合TCGA(The Cancer Genome Atlas)数据库与GTEx(Genotype-Tissue)数据库中的数据,分析KTRT6A在PDAC组织中的表达情况,并通过CIBERSORT工具分析KRT6A表达与PDAC组织中免疫细胞浸润的关系,然后通过GSEA方法研究与KRT6A基因表达相关的肿瘤信号通路。选取长海医院病理科保存的60例PDAC组织与癌旁组织标本进行免疫组化分析,验证KRT6A在肿瘤组织中表达情况;通过干扰RNA敲减PANC1细胞中KRT6A的表达,采用CCK-8实验以及流式细胞术检测敲减KRT6A对细胞的增殖、凋亡的影响。结果:利用TCGA与GTEx数据库数据分析发现,KRT6A在人PDAC组织中呈高表达,且与患者较差的生存期存在关联(P=0.015)。利用CIBERSORT软件以及GSEA分析发现,KRT6A高表达的PDAC组织中M2型巨噬细胞浸润性升高(P=0.034),且与Wnt通路(NES:1.7359272,P<0.05)、磷酸戊糖途径(PPP)(NES:1.5613053,P<0.05)等信号通路上调有关联(P<0.05或P<0.01);免疫组化结果进一步验证了KRT6A在PDAC组织中呈高表达(P<0.001)。增殖和凋亡实验发现,干扰KRT6A能够显著抑制PANC1细胞的增殖(P<0.05)以及凋亡(P<0.001)。结论:KRT6A在人PDAC组织中呈高表达,敲减其表达能够抑制PANC1细胞的增殖和凋亡,具有作为PDAC诊断与预后判断新靶标的潜力。

Objective:To study,through bioinformatics analysis and cellular biology experiments,the effects of keratin 6A(KRT6A)on the diagnosis,prognosis and immune microenvironment of pancreatic ductal adenocarcinoma(PDAC)and biological behaviors of PDAC PANC1 cells,such as proliferation and apoptosis.Methods:The GEPIA platform was used to integrate the data from the TCGA(The Cancer Genome Atlas)and the GTEx(Genotype-Tissue)to analyze the KTRT6A expression in PDCA tissues.CIBERSORT software was then used to analyze the correlation between KRT6A expression and immune cell infiltration in PDCA tissues,and GSEA analysis was used to study the signaling pathway related to KRT6A gene expression.Immunohistochemical analysis was performed on cancer and adjacent tissue samples from 60 PDAC patients preserved in the Department of Pathology of Changhai Hospital to verify the expression of KRT6A in tumor tissues.The expression of KRT6A gene in PANC1 cells was inhibited by interfering with siRNA.The effects of KRT6A on the proliferation and apoptosis of PDAC cells were detected by CCK-8 assay and flow cytometry.Results:Data analysis of the TCGA and the GTEx found that KRT6A was highly expressed in human PDAC tissues and was significantly correlated with poor survival period of patients(P=0.015).CIBERSORT software and GSEA analysis showed that the infiltration of M2-type macrophages in PDCA tissues with high KRT6A expression was increased(P=0.034),and there was a significant correlation between high KRT6A expression and the up-regulation of Wnt pathway(NES:1.7359272,P<0.05),pentose phosphate pathway(PPP)(NES:1.5613053,P<0.05)and other signaling pathways(P<0.05 or P<0.01).Immunohistochemical results further verified the high expression of KRT6A in PDCA tissues(P<0.001).Proliferation and apoptosis experiments showed that interfering with KRT6A gene could significantly inhibit the proliferation(P<0.05)and apoptosis(P<0.001)of PANC1 cells.Conclusion:KRT6A is highly expressed in human PDAC tissues,and knocking down its expression can inhibit the proliferation and apoptosis of PANC1 cells,and has the potential to be a new target for PDAC diagnosis and prognosis.