柠檬酸转运体SLC25A1对食管鳞癌细胞体外放射敏感性的影响及其分子机制

doi:10.3969/j.issn.1004-616x.2020.02.009

癌变·畸变·突变 ›› 2020, Vol. 32 ›› Issue (2): 132-138.doi: 10.3969/j.issn.1004-616x.2020.02.009

柠檬酸转运体SLC25A1对食管鳞癌细胞体外放射敏感性的影响及其分子机制

师盼^1,2, 林重华^2,3, 舒易方⁴, 瞿家权^2,3, 宋淑亮¹

1. 山东大学海洋学院, 山东威海 264209;
2. 陆军军医大学第一附属医院消化内科胆汁淤积肝病中心, 重庆 400038;
3. 吉首大学医学院医学影像技术系, 湖南吉首 416000;
4. 陆军军医大学第一附属医院肿瘤科放疗中心, 重庆 400038

收稿日期:2019-11-08 修回日期:2020-03-10 出版日期:2020-03-30 发布日期:2020-04-10
通讯作者: 宋淑亮,E-mail:songshuliang@sdu.edu.cn E-mail:songshuliang@sdu.edu.cn
作者简介:师盼,E-mail:pan.shi@cldcsw.org。
基金资助:
国家自然科学基金（81703043，81770583）；山东省自然科学基金（ZR2017MH040）；湖南省卫生厅科研课题（B2017070）

Role of mitochondrial citrate transporter SLC25A1 in radiosensitivity of human esophageal squamous cell carcinoma in vitro

SHI Pan^1,2, LIN Chonghua^2,3, SHU Yifang⁴, QU Jiaquan^2,3, SONG Shuliang¹

1. Marine College, Shandong University, Weihai 264209, Shandong;
2. Cholestatic Liver Diseases Center and Department of Gastroenterology, Southwest Hospital, Third Military Medical University, Chongqing 400038;
3. Department of Imaging Technology, Jishou University, Jishou 416000, Hunan;
4. Department of Oncology, Southwest Hospital, Third Military Medical University, Chongqing 400038, China

Received:2019-11-08 Revised:2020-03-10 Online:2020-03-30 Published:2020-04-10

摘要/Abstract

摘要： 目的： 探讨线粒体柠檬酸转运体SLC25A1对食管癌细胞体外放射敏感性的影响及其分子机制。方法： 采用癌症基因组图谱（TCGA）数据库及UALCAN交互式网络工具比较分析184例食管癌组织与11例正常组织的SLC25A1 mRNA表达水平的差异；采用亚致死剂量X射线多次暴露法建立放射抵抗及对照组食管癌细胞系，放射克隆存活法线性二次靶模型分析两株食管癌细胞株放疗反应性的差异；多次X射线照射（总剂量6.0 Gy）上述两株食管癌细胞后，Hoechst33258染色荧光显微镜观察细胞凋亡形态学变化，流式细胞术检测细胞活性氧水平；免疫印迹法分析慢病毒小发夹RNA稳定转染敲低SLC25A1表达对DNA损伤修复关键信号分子H2AX和DNA-PKcs磷酸化水平的影响。结果： TCGA肿瘤组织数据库比较分析证实，与正常食管组织比较，食管癌组织SLC25A1 mRNA表达水平升高1.65倍（P=1.64×10^-4）；多轮亚致死剂量暴露法构建的放射抵抗食管鳞癌细胞株SLC25A1蛋白表达水平较对照组升高2.36倍（P < 0.05）；与对照组比较，稳定敲低SLC25A1表达的放射抵抗食管癌细胞株凋亡率增加1.58倍（P < 0.05）；活性氧水平下调（65.3±14.3）%（P=0.007）；并下调磷酸化H2AX（Ser139）和磷酸化DNA-PKcs（Ser2056）水平（P均 < 0.05）。结论： SLC25A1可能是一种参与食管鳞癌发病和放射抵抗的癌基因，通过下调活性氧水平，抑制DNA损伤修复，诱导细胞凋亡并可作为食管鳞癌放射增敏的潜在分子靶标。

关键词: 线粒体柠檬酸转运体, 食管鳞癌, 放射增敏, 活性氧产物, DNA损伤修复

Abstract: OBJECTIVE: To investigate the effect and molecular mechanism of mitochondrial citrate transporter SLC25A1 on sensitivity of esophageal squamous cell carcinoma to radiotherapy in vitro. METHODS: The Cancer Genome Atlas (TCGA) tumor tissue database and UALCAN interactive network tool were used to compare differences in SLC25A1 expression between 184 cases of esophageal cancer and 11 cases of normal tissues. Radioresistant esophageal carcinoma cell line and the control were established by multiple round of X-ray sublethal dose exposures. Radioresistance and radiosensitivities were characterized by the radiation clonal survival assay and the linear quadratic target model. Morphologic changes of cell apoptosis were observed by fluorescence microscopy using the Hoechst 33258 staining after multiple X-ray exposures at a total dose of 6.0 Gy. Flow cytometry was used to detect levels of reactive oxygen species after the same X-ray exposures. Western blotting was used to analyze the expression alteration of levels of phosphor-H2AX and phosphor-DNA-PKcs,and the key signal kinases in DNA repair by SLC25A1 knockdown with lentivirus vector carrying small hairpin interfering RNA. RESULTS: A comparative analysis of TCGA tumor tissue database confirmed that the expression level of SLC25A1 mRNA in esophageal cancer tissues was 1.65-folds higher than that in normal esophageal tissues (P=1.64×10^-4). The expression level of SLC25A1 protein was increased by 2.36-folds when compared with the control (P < 0.05). The apoptosis rate of esophageal cancer cell lines with stable SLC25A1 expression compared to the control was increased by 1.58-folds (P < 0.05). ROS level in the SLC25A1 knockdown radioresistant esophageal carcinoma cell line was decreased by (65.3±14.3)% (P=0.007). Levels of phosphorylation of H2AX (Ser139) and DNA-PKCs (Ser2056) in SLC25A1 knockdown radioresistant esophageal carcinoma cell lines were significantly reduced when compared with the control (all P < 0.05). CONCLUSION: SLC25A1 may be an oncogene involved in tumor pathogenesis and radiation resistance. The data suggest that it can inhibit DNA damage repair and induce cell apoptosis by down-regulating reactive oxygen species,and can serve as a potential molecular target for radiation sensitization of esophageal squamous cell carcinoma.

Key words: SLC25A1, esophageal squamous cell carcinoma, radiosensitization, reactive oxygen species, DNA repair

中图分类号:

R735.1

师盼, 林重华, 舒易方, 瞿家权, 宋淑亮. 柠檬酸转运体SLC25A1对食管鳞癌细胞体外放射敏感性的影响及其分子机制[J]. 癌变·畸变·突变, 2020, 32(2): 132-138.

SHI Pan, LIN Chonghua, SHU Yifang, QU Jiaquan, SONG Shuliang. Role of mitochondrial citrate transporter SLC25A1 in radiosensitivity of human esophageal squamous cell carcinoma in vitro[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2020, 32(2): 132-138.

参考文献

[1] MALHOTRA G K, YANALA U, RAVIPATI A, et al. Global trends in esophageal cancer[J]. J Surg Oncol, 2017, 115(5):564-579.
[2] ZENG H M, ZHENG R S, ZHANG S W, et al. Esophageal cancer statistics in China, 2011:Estimates based on 177 cancer registries[J]. Thorac Cancer, 2016, 7(2):232-237.
[3] ROEDER F, NICOLAY N H, NGUYEN T, et al. Intensity modulated radiotherapy (IMRT) with concurrent chemotherapy as definitive treatment of locally advanced esophageal cancer[J]. Radiat Oncol, 2014, 9:191.
[4] 陈海军, 刘玉琴, 李雨泽, 等. 维生素E琥珀酸酯对人食管癌TE-1和Eca-109细胞增殖及凋亡的影响[J]. 癌变·畸变·突变, 2015, 27(5):357-360.
[5] KLEINBERG L, GIBSON M K, FORASTIERE A A. Chemoradiotherapy for localized esophageal cancer:regimen selection and molecular mechanisms of radiosensitization[J]. Nat Clin Pract Oncol, 2007, 4(5):282-294.
[6] HE E H, PAN F, LI G C, et al. Fractionated ionizing radiation promotes epithelial-mesenchymal transition in human esophageal cancer cells through PTEN deficiency-mediated akt activation[J]. PLoS One, 2015, 10(5):e0126149.
[7] FERNANDEZ H R, GADRE S M, TAN M J, et al. The mitochondrial citrate carrier, SLC25A1, drives stemness and therapy resistance in non-small cell lung cancer[J]. Cell Death Differ, 2018, 25(7):1239-1258.
[8] KOLUKULA V K, SAHU G, WELLSTEIN A, et al. SLC25A1, or CIC, is a novel transcriptional target of mutant p53 and a negative tumor prognostic marker[J]. Oncotarget, 2014, 5(5):1212-1225.
[9] QU J Q, YI H M, YE X, et al. MiRNA-203 reduces nasopharyngeal carcinoma radioresistance by targeting IL8/AKT signaling[J]. Mol Cancer Ther, 2015, 14(11):2653-2664.
[10] CHANDRASHEKAR D S, BASHEL B, BALASUBRAMANYA S A H, et al. UALCAN:a portal for facilitating tumor subgroup gene expression and survival analyses[J]. Neoplasia, 2017, 19(8):649-658.
[11] HLOUSCHEK J, HANSEL C, JENDROSSEK V, et al. The mitochondrial citrate carrier (SLC25A1) sustains redox homeostasis and mitochondrial metabolism supporting radioresistance of cancer cells with tolerance to cycling severe hypoxia[J]. Front Oncol, 2018, 8:170.
[12] CATALINA-RODRIGUEZ O, KOLUKULA V K, TOMITA Y, et al. The mitochondrial citrate transporter, CIC, is essential for mitochondrial homeostasis[J]. Oncotarget, 2012, 3(10):1220-1235.
[13] SCHÜRMANN R, VOGEL S, EBEL K, et al. The physico-chemical basis of DNA radiosensitization:implications for cancer radiation therapy[J]. Chemistry, 2018, 24(41):10271-10279.
[14] SKVORTSOVA I, DEBBAGE P, KUMAR V, et al. Radiation resistance:Cancer stem cells (CSCs) and their enigmatic pro-survival signaling[J]. Semin Cancer Biol, 2015, 35:39-44.
[15] KIM W, YOUN H, KANG C, et al. Inflammation-induced radioresistance is mediated by ROS-dependent inactivation of protein phosphatase 1 in non-small cell lung cancer cells[J]. Apoptosis, 2015, 20(9):1242-1252.
[16] TRACHOOTHAM D, ALEXANDRE J, HUANG P. Targeting cancer cells by ROS-mediated mechanisms:a radical therapeutic approach?[J]. Nat Rev Drug Discov, 2009, 8(7):579-591.
[17] MORCIANO P, CARRISI C, CAPOBIANCO L, et al. A conserved role for the mitochondrial citrate transporter Sea/SLC25A1 in the maintenance of chromosome integrity[J]. Hum Mol Genet, 2009, 18(21):4180-4188.

柠檬酸转运体SLC25A1对食管鳞癌细胞体外放射敏感性的影响及其分子机制

Role of mitochondrial citrate transporter SLC25A1 in radiosensitivity of human esophageal squamous cell carcinoma in vitro

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