基于生物信息学探讨miR-15b-5p在头颈部鳞状细胞癌中的诊断价值和潜在作用机制

Diagnostic value and potential mechanism of miR-15b-5p in head andneck squamous cell carcinoma: an exploration based on bioinformatics

目的采用生物信息学探讨miR-15b-5p在头颈部鳞状细胞癌(HNSCC)中的诊断价值和潜在作用机制。方法(1)从GEO数据库和TCGA数据库中收集HNSCC的miR-15b-5p表达数据和/或HNSCC患者的临床资料。通过Meta分析miR-15b-5p在HNSCC中的表达情况及其诊断HNSCC的价值,使用SPSS 26.0软件分析miR-15b-5p表达水平与HNSCC患者临床特征的关系。利用实时荧光定量PCR检测HNSCC细胞和正常细胞中的miR-15b-5p表达水平。(2)通过miRWalk数据库和GEPIA2数据库分别筛选miR-15b-5p的下游靶基因和HNSCC的差异表达基因,对两者取交集后获得miR-15b-5p在HNSCC中的关键靶基因,对关键靶基因进行富集分析。(3)利用UALCAN数据库分析部分靶基因在HNSCC中的表达水平和甲基化水平。利用GEPIA2数据库分析部分关键靶基因表达情况与HNSCC患者无病生存期和总生存期的关系。结果(1)Meta分析结果显示,miR-15b-5p在HNSCC中呈高表达(P<0.05)。与正常细胞相比,miR-15b-5p在HNSCC细胞中的表达水平升高(P<0.05)。miR-15b-5p表达水平诊断HNSCC的敏感度、特异度、阳性似然比、阴性似然比和诊断优势比分别为0.70、0.72、2.15、0.48和7.33,汇总受试者工作特征曲线下面积为0.81。男性、年龄<60岁、N_(1)~N_(3)期患者的miR-15b-5p表达水平分别高于女性、年龄≥60岁、N_(0)期患者(P<0.05)。(2)共获得54个miR-15b-5p在HNSCC中的关键靶基因。功能富集分析结果提示,关键靶基因与细胞外基质密切相关;通路富集分析结果提示关键靶基因与多条信号通路密切相关,其中包含视黄醇代谢通路。(3)与正常组织相比,关键靶基因RDH12和UGT1A7在HNSCC组织中表达下调,关键靶基因UGT1A10和UGT1A7在HNSCC组织中的甲基化水平降低(P<0.05)。与UGT1A7表达量低的HNSCC患者相比,UGT1A7表达量高的HNSCC患者的无病生存期更长(P<0.05),但两者的总生存期差异无统计学意义(P>0.05)。结论miR-15b-5p与HNSCC的发生、发展密切相关,对HNSCC有潜在的诊断价值。miR-15b-5p可能通过靶向作用UGT1A7和RDH12基因来参与调控视黄醇代谢信号通路,从而影响HNSCC的发生、发展。

Objective To investigate the diagnostic value and potential mechanism of miR-15b-5p in head and neck squamous cell carcinoma(HNSCC)by using bioinformatics.Methods(1)Expression data of miR-15b-5p in HNSCC and/or clinical data of HNSCC patients were collected from the GEO and TCGA databases.Expression of miR-15b-5p in HNSCC and its value for diagnosing HNSCC were analyzed by Meta-analysis.The SPSS 26.0 software was used to analyze the relation between miR-15b-5p expression and clinical characteristics of HNSCC patients.The real-time fluorescent quantitative PCR was employed to detect miR-15b-5p expression in HNSCC cells and normal cells.(2)The downstream target genes of miR-15b-5p and differentially expressed genes of HNSCC were screened from the databases of miRWalk and GEPIA2,respectively,after performing intersection of the two,the key target genes of miR-15b-5p in HNSCC were obtained.The enrichment analysis was performed on the key target genes.(3)Expressions and methylation levels of partial target genes in HNSCC were analyzed by using the UALCAN database.The relation of expressions of partial key target genes with disease-free survival and overall survival of HNSCC patients was analyzed by employing the GEPIA2 database.Results(1)The results of Meta-analysis revealed that miR-15b-5p exhibited a high expression in HNSCC(P<0.05).Compared with normal cells,expression of miR-15b-5p was elevated in HNSCC cells(P<0.05).The sensitivity,specificity,positive likelihood ratio,negative likelihood ratio,and diagnostic odds ratio of miR-15b-5p expression in the diagnosis of HNSCC were 0.70,0.72,2.15,0.48,and 7.33,respectively,and area under the summary receiver operating characteristic curve was 0.81.Male patients,and patients with age<60 years old,in N_(1)-N_(3) stage obtained a higher miR-15b-5p expression as compared with female patients,and patients with age≥60 years old,in N_(0) stage,respectively(P<0.05).(2)A total of 54 key target genes of miR-15b-5p in HNSCC were obtained.The results of functional enrichment analysis revealed that the key target genes were closely related to extracellular matrix,and the results of pathway enrichment analysis indicated that the key target genes were closely related to multiple signaling pathways,containing retinol metabolic pathway.(3)Compared with normal tissues,expressions of the key target genes RDH12 and UGT1A7 were down-regulated in HNSCC tissues,and methylation levels of the key target genes UGT1A10 and UGT1A7 were decreased in HNSCC tissues(P<0.05).HNSCC patients with high UGT1A7 expression obtained a longer disease-free survival than those with low UGT1A7 expression(P<0.05),but there was no statistically significant difference in overall survival between the two(P>0.05).Conclusion There is a close correlation between miR-15b-5p and the occurrence and development of HNSCC,and miR-15b-5p has a potential diagnostic value on HNSCC.MiR-15b-5p may participate in the regulation of retinol metabolism signaling pathway by targeting UGT1A7 and RDH12 genes,thereby affecting the occurrence and development of HNSCC.