导致鼻咽癌(NPC)患者临床治疗不良结局的主要因素是辐射抵抗和复发。本研究旨在探讨细胞角蛋白13(CK13)在鼻咽癌放疗中的敏感性及分子基础。为了实现这一点,构建了过表达CK13的人NPC细胞系HNE-3-CK13。用CCK-8法评价了CK13过表达对放疗条件下细胞活力和凋亡的影响,免疫荧光,和蛋白质印迹(WB)。进行下一代测序以鉴定介导放疗反应的CK13的下游基因和信号通路。通过使用克隆形成和WB的挽救实验,研究了候选基因ERRFI1在CK13诱导的放射敏感性增强中的潜在作用。ERRFI1对细胞活力的影响,细胞凋亡,细胞周期,并使用CCK-8,免疫荧光,流式细胞术,定量聚合酶链反应和WB。结果显示,HNE-3中CK13过表达显著抑制放疗下细胞存活,促进细胞凋亡标志物γH2AX表达,导致ERRFI1显著增加。ERRFI1的敲低挽救了由CK13过表达介导的NPC细胞的放疗敏化引起的细胞活力和增殖的降低以及细胞凋亡的增加。在这个过程中,EGFR,AKT,发现GSK-3β参与其中。最后,ERRFI1被证明可以抑制CDK1,CDK2,细胞周期蛋白B1和细胞周期蛋白D1的表达水平,从而导致G2/M细胞比率增加。CK13的过表达增强了NPC细胞的放射敏感性,其特征是细胞活力和增殖降低以及细胞凋亡增加。这种调控可能通过增加ERRFI1的表达和激活EGFR/Akt/GSK-3β信号通路来影响HNE-3细胞的存活,为鼻咽癌的治疗提供了新的潜在治疗靶点。
The main factors contributing to the unfavorable outcome in the clinical treatment of patients with nasopharyngeal carcinoma (NPC) patients are radiation resistance and recurrence. This study aimed to investigate the sensitivity and molecular foundation of cytokeratin 13 (CK13) in the radiotherapy of NPC. To achieve this, a human NPC cell line overexpressing CK13, HNE-3-CK13, was constructed. The effects of CK13 overexpression on cell viability and apoptosis under radiotherapy conditions were evaluated using the CCK-8 assay, immunofluorescence, and western blotting (WB). Next-generation sequencing was performed to identify the downstream genes and signaling pathways of CK13 that mediate radiotherapy response. The potential role of the candidate gene
ERRFI1 in CK13-induced enhancement of radiosensitivity was investigated through rescue experiments using clone formation and WB. The effects of
ERRFI1 on cell viability, cell apoptosis, cell cycle, and the related key genes were further evaluated using CCK-8, immunofluorescence, flow cytometry, quantitative polymerase chain reaction and WB. The results showed that CK13 overexpression in HNE-3 significantly inhibited cell survival under radiotherapy and promoted apoptosis marker γH2AX expression, leading to a significant increase of ERRFI1. Knockdown of
ERRFI1 rescued the decreased cell viability and proliferation and the increased cell apoptosis that were caused by CK13 overexpression-mediated radiotherapy sensitization of NPC cells. In this process, EGFR, AKT, and GSK-3β were found involved. In the end,
ERRFI1 was proven to inhibit expression levels of CDK1, CDK2, cyclin B1, and cyclin D1, resulting an increased G2/M cell ratio. Overexpression of CK13 enhances the radiosensitivity of NPC cells, which is characterized by decreased cell viability and proliferation and increased apoptosis. This regulation may affect the survival of HNE-3 cells by increasing the expression of
ERRFI1 and activating the EGFR/Akt/GSK-3β signaling pathway, providing new potential therapeutic targets for the treatment of NPC.