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《天津医科大学学报》[ISSN:1006-8147/CN:12-1259/R]

卷:

31卷

期数:

2025年03期

页码:

203-209

栏目:

肿瘤疾病专题

出版日期:

2025-05-20

文章信息/Info

Title:

High expression of TERRA enhanced ZBP1 and ferroptosis in glioblastoma

文章编号:

1006-8147(2025)03-0203-07

作者:

黄欣宇; 刘思琪; 郝名英; 耿鑫

（天津医科大学基础医学院生物化学与分子生物学系，天津300070）

Author(s):

HUANG Xinyu; LIU Siqi; HAO Mingying; GENG Xin

（Department of Biochemistry and Molecular Biology，School of Basic Medical Sciences，Tianjin Medical University，Tianjin 300070，China）

关键词:

端粒; 胶质母细胞瘤; 铁死亡; Z-DNA结合蛋白1

Keywords:

telomerase; glioblastoma; ferroptosis; ZBP1

分类号:

R34

DOI:

10.20135/j.issn.1006-8147.2025.03.0203

文献标志码:

A

摘要:

目的：探究非编码RNA（TERRA）的表达增加对Z-DNA结合蛋白1（ZBP1）和胶质母细胞瘤铁死亡的影响。方法：实验分为野生组（WT组）、阴性对照组（Scramble组）、高表达TERRA处理组、抑制TERRA组。慢病毒感染构建高表达TERRA的U251细胞株并通过蛋白印记、RT-qPCR验证效率；qPCR检测ZBP1及其下游信号通路相关基因mRNA水平；通过IF-FISH实验检测TERRA和ZBP1共定位情况；通过Western印迹验证ZBP1、铁死亡相关蛋白含量变化；检测铁死亡下游脂质过氧化指标丙二醛（MDA）、活性氧簇（ROS）以及氧化还原指标谷胱甘肽（GSH）含量。结果：Western印迹显示，慢病毒包装后高表达TERRA处理组比WT组的端粒重复结合因子2（TRF2）蛋白表达量降低（t=3.135，P<0.05）；qPCR验证高表达TERRA处理组比WT组9p、15q和Xp染色体末端TERRA水平显著升高（t=24.77、34.08、13.27，均P<0.001）；高表达TEERA处理组比WT组中ZBP1的蛋白和转录水平升高（t=15.73，P<0.01）；qPCR 检测高表达TEERA处理组的趋化因子 CCL5 （t=17.27，P<0.001）和干扰素调节因子IRF1（t=32.06，P<0.01）转录水平比WT组显著升高；IF-FISH结果显示高表达TERRA处理组比WT组，TERRA和ZBP1的共定位信号更多；使用siRNA后，与WT组相比，抑制TERRA组中的ZBP1、CCL5、IRF1表达水平均无显著差异；Western印迹显示，高表达TERRA处理组比WT组溶质载体家族转运蛋白（SLC7A11）的表达显著降低，谷胱甘肽过氧化物酶4（GPX4）的变化与其一致（t=13.510、8.994，均P<0.05）；与WT组相比，高表达TERRA处理组ROS、MDA水平显著增加（F=128.4、9.824，均P<0.01），GSH含量降低（F=6.815，P<0.05）。使用siRNA后，与WT组相比，抑制TERRA组SLC7A11、GPX4、ROS、MDA、GSH差异均无统计学意义。结论：胶质母细胞瘤中TERRA高表达，增强ZBP1表达并促进肿瘤细胞铁死亡。

Abstract:

Objective： To explore the impact of increased enhancement of long non-coding RNA （TERRA） on Z-DNA binding protein 1 （ZBP1） and ferroptosis in glioblastoma. Methods：The experiment was divided into four distinct treatment groups：the Wild Type group （WT group），the Negative Control group （Scramble group），the high TERRA expression group，and the TERRA inhibition group. Lentiviral infection was used to establish a U251 cell line overexpressing TERRA，with efficiency confirmed by Western blotting and RT-qPCR. The mRNA levels of genes related to ZBP1 and its downstream signaling pathways were detected by qPCR. IF-FISH was used to detect the co-localization signals of TERRA and ZBP1. The changes in ZBP1 and ferroptosis related protein content was verified by Western blotting. The levels of malondialdehyde （MDA），reactive oxygen species （ROS），glutathione （GSH） in the downstream of fer-roptosis were detected. Results：Western blotting revealed that the expression of telomeric repeat binding factor 2 （TRF2） protein in the lentiviral-packaged high TERRA expression group was reduced compared with the WT group （t=3.135，P<0.05）. QPCR validation confirmed a significant increase in TERRA levels at the termini of chromosomes 9p，15q，and Xp in the high TERRA expression group compared with the WT group （t=24.77，34.08，13.27，all P<0.001）. The protein and transcription levels of ZBP1 were elevated in the high TERRA expression group compared with the WT group （t=15.73，P<0.01）. The transcription levels of chemokine CCL5 （t=17.27，P<0.001） and interferon regulatory factor IRF1 were significantly higher （t=32.06，P<0.01） in the high TERRA expression group than those in the WT group，as detected by qPCR. IF-FISH results showed more co-localization signals of TERRA and ZBP1 in the high TERRA expression group compared with the WT group. After siRNA treatment，there were no significant differences in the expression levels of ZBP1，CCL5，and IRF1 in the TERRA inhibition group compared with the WT group. Western blotting indicated a significant decrease in the expression of solute carrier family member （SLC7A11） and a consistent change in glutathione peroxidase 4 （GPX4） in the high TERRA expression group compared with the WT group （t= 13.510，8.994，both P<0.05）. Compared with the WT group，the levels of ROS and MDA were elevated in high TERRA expression group （F=128.4，9.824，both P<0.01），while the level of GSH was decreased （F=6.815，P<0.05）. After siRNA treatment，there were no significant differences in the levels of SLC7A11，GPX4，ROS，MDA，and GSH in the TERRA inhibition group compared with the WT group. Conclusion：High expression of TERRA in glioblastoma enhances ZBP1 expression and promotes ferroptosis in tumor cells.

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相似文献/References:

[1]韩鑫宇,李婷芳,王峰.GADD45A通过端粒替代延长途径调控骨肉瘤细胞增殖[J].天津医科大学学报,2023,29(03):265.
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[2]徐子韫,黄欣宇,耿鑫.TERRA的功能、调控及其与替代性端粒延长机制之间的关系[J].天津医科大学学报,2026,32(02):190.[doi:10.20135/j.issn.1006-8147.2026.02.0190]

备注/Memo

备注/Memo:

基金项目: 国家自然科学基金面上项目（82271265）；天津市教委科研计划（2021ZD037）
作者简介: 黄欣宇（2000-），女，硕士在读，研究方向：医学生物化学与分子生物学；
通信作者：耿鑫，E-mail：gengx@tmu.edu.cn。

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更新日期/Last Update: 2025-06-01