Mechanisms of copper transporter 1 gene in regulating radiation induced intestinal injury
10.3760/cma.j.cn112271-20221226-00496
- VernacularTitle:铜吸收转运蛋白介导放射性肠损伤的作用及机制研究
- Author:
Yixian WANG
1
;
Li LIU
;
Wei MO
;
Wei ZHU
;
Yahui FENG
;
Yang JIAO
;
Jianping CAO
Author Information
1. 苏州大学放射医学与防护学院 放射医学与辐射防护国家重点实验室，苏州 215123
- Keywords:
Ionizing radiation;
Radiation induced intestinal injury;
Copper transporter 1;
Cuproptosis
- From:
Chinese Journal of Radiological Medicine and Protection
2023;43(6):401-408
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effects and mechanisms of copper transporter 1 (CTR1) in radiation induced intestinal injury in vitro. Methods:Human small intestinal epithelial cells (HIEC) were irradiated with 2, 4, 6, 8 Gy of X-rays and rat intestinal epithelial cells (IEC-6) were irradiated with 5, 10, 15, 20 Gy of X-rays. At 2, 4, 8, 24, and 48 h after irradiation, the expression of CTR1 was detected by Western blot assay. In some experiments, HIEC and IEC-6 cells were transfected with CTR1 shRNA and then exposed to X-rays. Copper levels were detected by Inductively Coupled Plasma Mass Spectrometry (ICP-MS). The radiosensitivity of cells was verified by colonogenic assay, the cellular reactive oxygen species (ROS) level and DNA damage were detected to further explore the related mechanism. In addition, Western blot was applied to detect the expressions of antioxidants and cuproptosis associated proteins in enterocytes after silencing CTR1 or irradiation.Results:The expression of CTR1 was increased by X-ray irradiation in a dose-dependent manner ( t=3.53, 3.45, 6.37, 11.11, 11.13, P<0.05). CTR1 expression was successfully diminished by CTR1 shRNA adenovirus vectors. According to the survival curves, the enhancement ratios of the radiosensitivity of HIEC and IEC-6 cells with CTR1 knocking-down were 1.146 and 1.201, respectively. Radiation-induced copper accumulation was alleviated after CTR1 silencing in IEC-6 cells ( t=3.10, P<0.05). At 0.5 h after irradiation, the ROS production in the CTR1 knockdown group was significantly lower than that in the control group ( t=5.23, 2.96, P<0.05). At 1 h after irradiation, the protein expression of γ-H2AX in the CTR1 knockdown group was obviously lower than that in the control group ( t=7.50, 4.29, P<0.05). The expressions of Nrf2 and HO-1 were increased after irradiation, which could be further increased after CTR1 silencing. In addition, cuproptosis associated protein DLAT, LIAS and FDX1 were reduced post-irradiation, which were recovered after CTR1 silencing. Conclusions:The radioresistance of HIEC and IEC-6 cells was enhanced after CTR1 silencing, possibly through the intracellular ROS and cuproptosis pathway.
