Department of Radiotherapy, The Third Affiliated Hospital of Qiqihar Medical University, Qiqihar City, China.
Department of Otolaryngology, The Third Affiliated Hospital of Qiqihar Medical University, Tiefeng District, 27 Taishun Street, Qiqihar City 161000, China.
Mol Biotechnol. 2023 Nov;65(11):1857-1868. doi: 10.1007/s12033-023-00691-5. Epub 2023 Feb 23.
To screen microRNAs (miRNAs) and analyze their role in the nasopharyngeal carcinoma (NPC) development through differential analysis and cytological validation of the nasopharyngeal carcinoma dataset. The Gene Expression Omnibus (GEO) database of NPC-related data were utilized to screen for differential miRNAs, downstream target genes and relevant pathways, and the relationships among them were verified by luciferase reporter assay and cell co-culture. To analyze the function of miRNAs and downstream target genes, a series of mimics, inhibitors or Small interfering RNAs (siRNAs) targeting the downstream target genes were transfected into NPC cells or normal epithelial cells by cell transfection techniques. Cell Counting Kit-8 (CCK8), Transwell, Enzyme-linked immunosorbent assay (ELISA) apoptosis, and western blotting were adopted to determine the changes in cell activity, invasiveness, and apoptosis after differential miRNA and target gene overexpression or downregulation. Differential analysis of miRNA dataset showed that the expression of miR-26b was significantly downregulated in NPC, in agreement with the validation results of nasopharyngeal carcinoma cell lines. And downregulation of miR-26b expression in normal nasopharyngeal epithelial cells transformed the cells to tumors. CEP135 was identified as the miR-26b downstream target gene by mRNA dataset analysis, and a luciferase reporter test revealed a direct targeting link between the two. Upregulation of CEP135 levels in nasopharyngeal cancer cell lines increased cell activity, accelerated cell migration, and inhibited apoptosis. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed that CEP135 exerted the above effects on cells via the NF-κB pathway, and co-culture with NF-κB pathway blockers reversed cell biological behavior to the level of the control group. MiR-26b downregulation leads to CEP135 overexpression and NF-κB pathway activation in NPC, which enhances proliferation, migration, and prevents apoptosis of nasopharyngeal carcinoma cells. Therefore, the study further clarifies the biological behavior mechanism of NPC and suggests new therapeutic options for NPC.
通过对鼻咽癌（NPC）数据集的差异分析和细胞学验证，筛选 microRNAs（miRNAs）并分析其在 NPC 发展中的作用。利用 NPC 相关数据的基因表达综合数据库（GEO）筛选差异表达的 miRNAs、下游靶基因和相关通路，并通过荧光素酶报告基因检测和细胞共培养验证它们之间的关系。为了分析 miRNAs 和下游靶基因的功能，通过细胞转染技术将针对下游靶基因的一系列 miRNA 模拟物、抑制剂或小干扰 RNA（siRNA）转染到 NPC 细胞或正常上皮细胞中。采用细胞计数试剂盒-8（CCK8）、Transwell、酶联免疫吸附测定（ELISA）凋亡和 Western blot 检测差异 miRNA 和靶基因过表达或下调后细胞活性、侵袭和凋亡的变化。miRNA 数据集的差异分析表明，miR-26b 在 NPC 中的表达明显下调，与鼻咽癌细胞系的验证结果一致。下调正常鼻咽上皮细胞中 miR-26b 的表达可将细胞转化为肿瘤。通过 mRNA 数据集分析鉴定出 CEP135 是 miR-26b 的下游靶基因，荧光素酶报告基因检测显示两者之间存在直接靶向关系。在鼻咽癌细胞系中上调 CEP135 水平可增加细胞活性、加速细胞迁移并抑制凋亡。京都基因与基因组百科全书（KEGG）富集分析显示，CEP135 通过 NF-κB 通路对细胞发挥上述作用，与 NF-κB 通路阻滞剂共培养可将细胞生物学行为逆转至对照组水平。miR-26b 的下调导致 NPC 中 CEP135 的过表达和 NF-κB 通路的激活，从而增强鼻咽癌细胞的增殖、迁移和阻止凋亡。因此，该研究进一步阐明了 NPC 的生物学行为机制，并为 NPC 提供了新的治疗选择。
