Li Nuoxuan, Wang Liang, Yao Jie, Yang Hong
Department of Head and Neck Surgery, Guangzhou Medical University Affiliated Cancer Hospital, No.78 Hengzhigang Road, Yuexiu District, Guangzhou, 510030 Guangdong China.
Cytotechnology. 2025 Jun;77(3):96. doi: 10.1007/s10616-025-00755-1. Epub 2025 May 6.
Thyroid cancer (TC) is a leading malignancy of the endocrine system. We investigated mechanism of the N6-methyladenosine (m6A) reader heterogeneous nuclear ribonucleoprotein C (HNRNPC) facilitating immune escape in TC by stabilizing programmed death ligand 1 (PD-L1). HNRNPC expression in TC tissues was analyzed using databases. Human TC cells (BHT-101, B-CPAP, SW579) and human thyroid follicular epithelial cells (Nthy-ori3-1) were cultured in vitro. SW579 cells were treated with pcDNA3.1-HNRNPC (oe-HNRNPC) and small interfering (si)-PD-L1, and B-CPAP cells were transfected with si-HNRNPC. HNRNPC and PD-L1 expression levels were assessed by RT-qPCR and Western blot. Cell proliferation, migration and invasion were evaluated by CCK-8, colony formation, and Transwell assays. Carboxyfluorescein diacetate succinimidyl ester-labelled CD8 T cell proliferation and effector cytokine (interferon-γ, tumor necrosis factor-α) levels were measured by flow cytometry and ELISA. The correlation between HNRNPC and PD-L1 expression in TC tissues, m6A modification sites on PD-L1 messenger RNA (mRNA), and HNRNPC-PD-L1 interaction were analyzed by databases and RIP assay. PD-L1 m6A modification was determined by Me-RIP assay. PD-L1 mRNA stability was detected by treating cells with actinomycin D. HNRNPC was notably highly expressed in TC cells. HNRNPC promoted TC cell proliferation, migration and invasion, facilitating immune escape. Mechanistically, HNRNPC mediated m6A modification to strengthen PD-L1 mRNA stability and up-regulate PD-L1 expression. Moreover, knockdown of PD-L1 partially reversed the promotional effect of HNRNPC on immune escape in TC cells. HNRNPC bolstered PD-L1 stability and up-regulated PD-L1 expression through m6A modification, thus promoting immune escape in TC.
甲状腺癌（TC）是内分泌系统的一种主要恶性肿瘤。我们研究了N6-甲基腺苷（m6A）阅读器异质核糖核蛋白C（HNRNPC）通过稳定程序性死亡配体1（PD-L1）促进TC免疫逃逸的机制。利用数据库分析TC组织中HNRNPC的表达情况。体外培养人TC细胞（BHT-101、B-CPAP、SW579）和人甲状腺滤泡上皮细胞（Nthy-ori3-1）。用pcDNA3.1-HNRNPC（过表达HNRNPC，oe-HNRNPC）和小干扰（si）-PD-L1处理SW579细胞，用si-HNRNPC转染B-CPAP细胞。通过RT-qPCR和蛋白质印迹法评估HNRNPC和PD-L1的表达水平。通过CCK-8、集落形成和Transwell实验评估细胞增殖、迁移和侵袭能力。通过流式细胞术和酶联免疫吸附测定法测量羧基荧光素二乙酸琥珀酰亚胺酯标记的CD8 T细胞增殖和效应细胞因子（干扰素-γ、肿瘤坏死因子-α）水平。通过数据库和RNA免疫沉淀（RIP）实验分析TC组织中HNRNPC与PD-L1表达的相关性、PD-L1信使核糖核酸（mRNA）上的m6A修饰位点以及HNRNPC-PD-L1相互作用。通过甲基化RNA免疫沉淀（Me-RIP）实验确定PD-L1的m6A修饰。用放线菌素D处理细胞检测PD-L1 mRNA稳定性。HNRNPC在TC细胞中显著高表达。HNRNPC促进TC细胞增殖、迁移和侵袭，促进免疫逃逸。机制上，HNRNPC介导m6A修饰以增强PD-L1 mRNA稳定性并上调PD-L1表达。此外，敲低PD-L1可部分逆转HNRNPC对TC细胞免疫逃逸的促进作用。HNRNPC通过m6A修饰增强PD-L1稳定性并上调PD-L1表达，从而促进TC中的免疫逃逸。
