Pan Yan, Abdureyim Marhaba, Yao Qing, Li Xuejun
Department of Pharmacology, Health Science Center, School of Basic Medical Sciences, Peking University, Beijing, China.
Department of Biochemistry and Molecular Biology, Ningxia Medical University, Yinchuan, China.
Front Cell Dev Biol. 2021 Feb 19;9:604038. doi: 10.3389/fcell.2021.604038. eCollection 2021.
Tumor cell adhesion to the endothelium is one pattern of tumor-endothelium interaction and a key step during tumor metastasis. Endothelium integrity is an important barrier to prevent tumor invasion and metastasis. Changes in endothelial cells (ECs) due to tumor cell adhesion provide important signaling mechanisms for the angiogenesis and metastasis of tumor cells. However, the changes happened in endothelial cells when tumor-endothelium interactions are still unclear. In this study, we used Affymetrix Gene Chip Human Transcriptome Array 2.0. and quantitative real-time PCR (qPCR) to clarify the detailed gene alteration in endothelial cells adhered by prostate tumor cells PC-3M. A total of 504 differentially expressed mRNAs and 444 lncRNAs were obtained through chip data analysis. Gene Ontology (GO) function analysis showed that differentially expressed genes (DEGs) mainly mediated gland development and DNA replication at the biological level; at the cell component level, they were mainly involved in the mitochondrial inner membrane; and at the molecular function level, DEGs were mainly enriched in ATPase activity and catalytic activity. Kyoto Encyclopedia of Genes and Genomes (KEGG) signal pathway analysis showed that the DEGs mainly regulated pathways in cancer, cell cycle, pyrimidine metabolism, and the mTOR signaling pathway. Then, we constructed a protein-protein interaction functional network and mRNA-lncRNA interaction network using Cytoscape v3.7.2. to identify core genes, mRNAs, and lncRNAs. The miRNAs targeted by the core mRNA PRKAA2 were predicted using databases (miRDB, RNA22, and Targetscan). The qPCR results showed that miR-124-3p, the predicted target miRNA of PRKAA2, was significantly downregulated in endothelial cells adhered by PC-3M. With a dual luciferase reporter assay, the binding of miR-124-3p with PRKAA2 3'UTR was confirmed. Additionally, by using the knockdown lentiviral vectors of miR-124-3p to downregulate the miR-124-3p expression level in endothelial cells, we found that the expression level of PRKAA2 increased accordingly. Taken together, the adhesion of tumor cells had a significant effect on mRNAs and lncRNAs in the endothelial cells, in which PRKAA2 is a notable changed molecule and miR-124-3p could regulate its expression and function in endothelial cells.
肿瘤细胞与内皮细胞的黏附是肿瘤-内皮细胞相互作用的一种模式，也是肿瘤转移过程中的关键步骤。内皮细胞的完整性是阻止肿瘤侵袭和转移的重要屏障。肿瘤细胞黏附导致内皮细胞（ECs）发生变化，为肿瘤细胞的血管生成和转移提供了重要的信号传导机制。然而，肿瘤-内皮细胞相互作用时内皮细胞发生的变化仍不清楚。在本研究中，我们使用Affymetrix基因芯片人类转录组阵列2.0和定量实时PCR（qPCR）来阐明前列腺肿瘤细胞PC-3M黏附的内皮细胞中详细的基因改变。通过芯片数据分析共获得504个差异表达的mRNA和444个lncRNA。基因本体论（GO）功能分析表明，差异表达基因（DEGs）在生物学水平主要介导腺体发育和DNA复制；在细胞成分水平，它们主要参与线粒体内膜；在分子功能水平，DEGs主要富集于ATP酶活性和催化活性。京都基因与基因组百科全书（KEGG）信号通路分析表明，DEGs主要调控癌症、细胞周期、嘧啶代谢和mTOR信号通路。然后，我们使用Cytoscape v3.7.2构建了蛋白质-蛋白质相互作用功能网络和mRNA-lncRNA相互作用网络，以鉴定核心基因、mRNA和lncRNA。使用数据库（miRDB、RNA22和Targetscan）预测核心mRNA PRKAA2靶向的miRNA。qPCR结果显示，PRKAA2预测的靶标miRNA miR-124-3p在PC-3M黏附的内皮细胞中显著下调。通过双荧光素酶报告基因检测，证实了miR-124-3p与PRKAA2 3'UTR的结合。此外，通过使用miR-124-3p的敲低慢病毒载体下调内皮细胞中miR-124-3p的表达水平，我们发现PRKAA2的表达水平相应增加。综上所述，肿瘤细胞的黏附对内皮细胞中的mRNA和lncRNA有显著影响，其中PRKAA2是一个显著变化的分子，miR-124-3p可以调节其在内皮细胞中的表达和功能。
