An Xiaohua, Huang Xiaoxue, Yu Qiujie, Tang Yiyue, Wang Yan, Chen Huasu, Zhang Yafei, Huang Qianhao, Rao Yudi, Hu Guomei, Zha He
Department of Laboratory Medicine, The Third Affiliated Hospital of Zunyi Medical University (The First People's Hospital of Zunyi), Zunyi, Guizhou, China.
Scientific Research Center, The Third Affiliated Hospital of Zunyi Medical University (The First People's Hospital of Zunyi), Zunyi, Guizhou, China.
Front Bioinform. 2025 Aug 1;5:1605681. doi: 10.3389/fbinf.2025.1605681. eCollection 2025.
To verify the expression profile of long non-coding RNAs (lncRNAs) and mRNAs in cervical cancer, identify their clinical significance in HPV16-associated cervical cancer, and annotate the biological function of mRNAs. Three pairs of cancerous and paracancer tissues were selected in cervical squamous cell carcinoma (IB2 stage), high-throughput sequencing was utilized to determine the expression levels of lncRNAs and mRNAs. The detection results were validated by GEPIA database analysis and RT-qPCR. Functional annotations of differential mRNAs were conducted through Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, and protein-protein interaction (PPI) network states. Furthermore, the association between antisense lncRNA and mRNA in cervical cancer was analyzed to predict the biological functions of lncRNA. Finally, recombinant lentivirus CV224-HPV16 E6/E7 was transfected into HcerEpic to establish a stable cell line with overexpressed HPV16 E6/E7, then differential lncRNAs were detected by RT-qPCR. Compared to paracancerous tissues, there were 3,608 lncRNAs significantly upregulated and 4,383 lncRNAs significantly downregulated in cervical cancer tissues (Fold change >2 and < 0.05). Additionally, 3,666 mRNAs were significantly upregulated, while 2,220 mRNAs were significantly downregulated (Fold change >2 and < 0.05). GO/KEGG enrichment analysis showed that differentially expressed mRNA played a significant role in cell cycle and cell senescence, and was related to signal pathways such as cAMP and MAPK, forming a complex network among the proteins encoded by these mRNAs. Further analysis indicated that the 20 antisense lncRNAs with the most remarkable differences might exert biological functions by influencing their corresponding mRNAs. The results of RT-qPCR revealed that CDKN2B-AS1, HAGLROS and GATA6-AS1 were potentially regulated by HPV16 E6/E7, which were in accordance with those obtained from chip detection. In this study, differentially expressed lncRNAs associated with HPV16 infection were screened and explored their transcriptional molecular functions and biological pathways, providing a molecular basis for predicting diagnostic markers of cervical cancer.
为验证长链非编码RNA（lncRNAs）和mRNA在宫颈癌中的表达谱，确定它们在HPV16相关宫颈癌中的临床意义，并注释mRNA的生物学功能。选取3对宫颈鳞状细胞癌（IB2期）癌组织和癌旁组织，利用高通量测序确定lncRNAs和mRNAs的表达水平。检测结果通过GEPIA数据库分析和RT-qPCR进行验证。通过基因本体论（GO）、京都基因与基因组百科全书（KEGG）通路富集分析以及蛋白质-蛋白质相互作用（PPI）网络状态对差异mRNA进行功能注释。此外，分析宫颈癌中反义lncRNA与mRNA之间的关联，以预测lncRNA的生物学功能。最后，将重组慢病毒CV224-HPV16 E6/E7转染至HcerEpic中，建立HPV16 E6/E7过表达的稳定细胞系，然后通过RT-qPCR检测差异lncRNAs。与癌旁组织相比，宫颈癌组织中有3608个lncRNAs显著上调，4383个lncRNAs显著下调（倍数变化>2且<0.05）。此外，3666个mRNAs显著上调，而2220个mRNAs显著下调（倍数变化>2且<0.05）。GO/KEGG富集分析表明，差异表达的mRNA在细胞周期和细胞衰老中起重要作用，并且与cAMP和MAPK等信号通路相关，在这些mRNA编码的蛋白质之间形成复杂网络。进一步分析表明，差异最显著的20个反义lncRNAs可能通过影响其相应的mRNA发挥生物学功能。RT-qPCR结果显示，CDKN2B-AS1、HAGLROS和GATA6-AS1可能受HPV16 E6/E7调控，这与芯片检测结果一致。本研究筛选了与HPV16感染相关的差异表达lncRNAs，探索了它们的转录分子功能和生物学途径，为预测宫颈癌诊断标志物提供了分子基础。
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