Department of Otorhinolaryngology Head and Neck Surgery, Shanghai Key Clinical Disciplines of Otorhinolaryngology, Eye & ENT Hospital of Fudan University, Shanghai, China.
Department of Otolaryngology, Shanghai Municipal Hospital of Traditional Chinese Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai, China.
J Clin Lab Anal. 2022 Jan;36(1):e24108. doi: 10.1002/jcla.24108. Epub 2021 Nov 17.
Aberrant expression of exosomal miRNAs has emerged as a research hotspot. However, no studies have been conducted on the dysregulation of exosomal miRNAs derived from cancer-associated fibroblasts (CAFs) in supraglottic laryngeal squamous cell carcinoma (SLSCC).
Cancer-associated fibroblasts and paired normal fibroblasts (NFs) from SLSCC patients were cultured, and exosomes in the culture supernatants were collected and identified. Exosomal miRNA expression was compared in each pair of CAFs and NFs by next-generation sequencing, and expression of selected exosomal miRNAs was validated by reverse transcription-quantitative PCR. Four online bioinformatic algorithms predicted the potential target genes of aberrantly expressed miRNAs, while gene ontology and Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway enrichment and network analysis identified downstream target genes and their interactions.
Three pairs of CAFs and NFs were successfully cultured and purified. CAF-derived exosomal miRNAs were mostly downregulated and included miR-656-3p, miR-337-5p, miR-29a-3p and miR-655-3p; however, some, including miR-184-3p, miR-92a-1-5p, miR-212-3p and miR-3135b, were upregulated. Bioinformatics analysis revealed involvement of these miRNAs in biological processes, cellular components and molecular functions. KEGG analysis revealed the top 30 pathways involvement in cancer initiation and progression and in cell cycle regulation. An interaction network showed miR-16-5p, miR-29a-3p, miR-34c-5p, miR-32-5p and miR-490-5p as the top five miRNAs and CCND1, CDKN1B, CDK6, PTEN and FOS as the top five target genes.
SLSCC patients showed aberrant expression of CAF-derived exosomal miRNAs. The top five miRNAs and their target genes may jointly constitute a carcinogenic tumour microenvironment and act as biomarkers for SLSCC intervention.
外泌体 miRNA 的异常表达已成为研究热点。然而，目前尚无研究报道关于源自声门上喉鳞状细胞癌（SLSCC）癌相关成纤维细胞（CAFs）的外泌体 miRNA 的失调。
培养 SLSCC 患者的 CAFs 和配对的正常成纤维细胞（NFs），收集和鉴定培养上清液中的外泌体。通过下一代测序比较每对 CAFs 和 NFs 中的外泌体 miRNA 表达，并通过逆转录定量 PCR 验证选定的外泌体 miRNA 的表达。四个在线生物信息学算法预测了异常表达 miRNA 的潜在靶基因，而基因本体和京都基因与基因组百科全书（KEGG）通路富集和网络分析确定了下游靶基因及其相互作用。
成功培养和纯化了三对 CAFs 和 NFs。CAF 来源的外泌体 miRNA 大多下调，包括 miR-656-3p、miR-337-5p、miR-29a-3p 和 miR-655-3p；然而，一些包括 miR-184-3p、miR-92a-1-5p、miR-212-3p 和 miR-3135b 在内的 miRNA 则上调。生物信息学分析显示这些 miRNA 参与了生物学过程、细胞成分和分子功能。KEGG 分析显示了参与癌症发生和进展以及细胞周期调节的前 30 个通路。相互作用网络显示 miR-16-5p、miR-29a-3p、miR-34c-5p、miR-32-5p 和 miR-490-5p 作为前五个 miRNA，CCND1、CDKN1B、CDK6、PTEN 和 FOS 作为前五个靶基因。
SLSCC 患者的 CAF 来源的外泌体 miRNA 表现出异常表达。前五个 miRNA 及其靶基因可能共同构成致癌性肿瘤微环境，并作为 SLSCC 干预的生物标志物。
