| Xu Xiaopeng,Dai Jun,Gao Yi,et al.Exploring the impacts and mechanisms of GCH1 dephosphorylated mutants on the radiosensitivity of esophageal cancers[J].Chinese Journal of Radiological Medicine and Protection,2024,44(10):819-826 |
| Exploring the impacts and mechanisms of GCH1 dephosphorylated mutants on the radiosensitivity of esophageal cancers |
| Received:March 04, 2024 |
| DOI：10.3760/cma.j.cn112271-20240304-00084 |
| KeyWords:Esophageal cancer Ionizing radiation GTP cyclohydrolase 1 (GCH1) Tetrahydrobiopterin (BH4) |
| FundProject:江苏省卫生健康委面上项目(H2023027);无锡市“太湖人才计划”优秀医学专家团队(2021-9);江阴市“暨阳名医”消化内镜领航专家团队(2023-37);无锡市卫生健康委青年人才项目(Q202206)及中青年后备拔尖人才资助计划(HB2023105);四川省科技计划项目(2022NSFSC0797,2023NSFSC0648) |
| Author Name | Affiliation | E-mail | | Xu Xiaopeng | Department of Gastroenterology, Jiangyin Clinical College of Xuzhou Medical University, Wuxi 214422, China | | | Dai Jun | Department of Gastroenterology, Jiangyin Clinical College of Xuzhou Medical University, Wuxi 214422, China | | | Gao Yi | Department of Gastroenterology, Jiangyin Clinical College of Xuzhou Medical University, Wuxi 214422, China | | | Wang Jian | Department of Gastroenterology, Jiangyin Clinical College of Xuzhou Medical University, Wuxi 214422, China | | | Sun Chuantang | West China School of Basic Medical Science and Forensic Medicine, Sichuan University, Chengdu 610041, China | | | Zhang Shuyu | West China School of Basic Medical Science and Forensic Medicine, Sichuan University, Chengdu 610041, China | | | Liu Pengfei | Department of Gastroenterology, Jiangyin Clinical College of Xuzhou Medical University, Wuxi 214422, China | pengfeimd@163.com |
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| Abstract:: |
| Objective To investigate the impacts and mechanisms of the GCH1-S81A mutants of the rate-limiting enzyme GTP cyclohydrolase 1 (GCH1) at key phosphorylation sites on the radiosensitivity of esophageal cancers during the de novo synthesis of tetrahydrobiopterin (BH4). Methods The KYSE-150 cell lines of esophageal squamous cell carcinoma with stable GCH1 overexpression at different phosphorylation levels were constructed in this study. Based on GCH1's phosphorylation levels and radiation doses, the experimental groups were divided into the blank control group, the empty virus group, the empty virus + irradiation group, the wild-type GCH1 group, the GCH1 phosphorylation group, the GCH1 dephosphorylation group, the GCH1 dephosphorylation + irradiation group, the validation group, and the validation + irradiation group. The Western blot and the CCK-8 assay were employed to detect the infection efficiency and the survival rates of tumor cells in various groups, respectively. The flow cytometry was applied to detect the changes in the apoptosis rate, reactive oxygen species (ROS) level, and lipid peroxide level of tumor cells in various groups. The colony formation assay was used to detect the changes in the radiosensitivity of tumor cells. Besides, the Western blot was performed to detect the changes in the expression of ferroptosis-related proteins. Results The GCH1 dephosphorylation group showed a significantly decreased expression level of phosphorylated GCH1-S81 protein in the cells at 48 h after infection (t=9.35, 16.57,P<0.05). Compared to the empty virus + irradiation group, the GCH1 dephosphorylation + irradiation group exhibited a significantly decreased cell survival rate 24 h after 10 Gy X-ray irradiation (t=26.97,P<0.05). The ROS levels in KYSE-150 cells at 8 h after 10 Gy X-ray irradiation, and the apoptosis rates and lipid peroxide levels at 48 h after irradiation, all showed a significant increase (t=17.89-131.1,P<0.05), which was further aggravated following the addition of GCH1-S81A mutants (t=157.06-97.81,P<0.05). This phenomenon could be inhibited by complementing wild-type GCH1 (t=66.38-23.08,P<0.05). Compared to the empty virus + irradiation group, the GCH1 dephosphorylation + irradiation group manifested decreased colony formation capacity under various X-ray doses (2, 4, 6 and 8 Gy, t=7.31-8.16,P<0.05). The GCH1-S81A mutation reduced the expression level of the ferroptosis-related protein GPX4 (t=4.55,P<0.05), which was further decreased after 10 Gy X-ray irradiation (t=12.98,P<0.05). Conclusions The GCH1-S81A dephosphorylated mutants can inhibit the growth of esophageal carcinoma cells KYSE-150 and enhance their radiosensitivity, which may hold promise as a novel approach to improve the efficacy of radiotherapy for esophageal cancers. |
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