Journal of Shandong University (Health Sciences) ›› 2021, Vol. 59 ›› Issue (4): 28-34.doi: 10.6040/j.issn.1671-7554.0.2020.1295

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Atg7-siRNA interferes with radiosensitivity of esophageal cancer ECA109 cells by regulating arginine circulation

ZHANG Xiaohong1*, ZHOU Yun2*, DU Qiuying1, REN Huixin1, WANG Chaoqun1   

  1. 1. Laboratory of Clinical and Experimental Pathology, Xuzhou Medical University, Xuzhou 221004, Jiangsu, China;
    2. Department of Radiotherapy, Xuzhou Clinical School of Xuzhou Medical University, Xuzhou 221004, Jiangsu, China
  • Published:2021-04-30

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Abstract: Objective To investigate the effects of siRNA interference with autophagy related gene 7(Atg7)on the radiosensitivity of ECA109 cells and the regulation of arginine circulation-related pathways. Methods ECA109 cells were divided into four groups: Ctrl group, siRNA group, Ctrl+R8Gy group(ECA109 cells transfected with Atg7-Ctrl at 8Gy radiation dose)and siRNA+R8Gy group(ECA109 cells transfected with Atg7-siRNA at 8Gy radiation dose). The protein expressions of Atg7, Beclin-1, microtubule associated protein 1 light chain 3 Ⅱ(LC3 Ⅱ), arginine succinate synthetase1(ASS1)and arginine succinic acid lyase(ASL)in each group were detected with Western blotting. The effects of arginine were detected with CCK8 assay. The cell survival curves were drawn at different concentrations of arginine(0, 8, 16, 32, 64, 128 mmol/L). IC50 and IC10 were calculated, and IC10 was set as the action concentration in the subsequent experiments. Cells were treated with arginine and siRNA respectively, and irradiated with 8Gy. After 24 h, the cell mortality was assessed by counting the number of cells. Results siRNA interference down-regulated Atg7 before and after radiotherapy(P<0.01). Compared with the Ctrl group, the Ctrl+R8Gy group had up-regulated levels of Beclin-1 and LC3Ⅱ(P<0.01). Compared with the Ctrl+R8Gy group, the siRNA+R8Gy group had down-regulated levels of Beclin-1 and LC3Ⅱ(P<0.01). Compared with the Ctrl+R8Gy group, the siRNA+R8Gy group had up-regulated levels of ASS1 and ASL(P<0.01). Arginine inhibited the proliferation of ECA109 cells in a concentration and time-dependent manner. Cell death analysis showed that siRNA interference(F=73.59, P<0.001)and arginine addition(F=158.72, P<0.001)increased cell mortality. The cell mortality was at the highest level when siRNA and arginine were used in combination(F=7.88, P=0.02). Conclusion Radiotherapy can induce autophagy in ECA109 cells. The Atg7-siRNA interference effectively antagonizes the radiotherapy-induced increase in autophagy, increases the radiotherapy-induced ECA109 cell death, and enhances the radiosensitivity of ECA109 cells. Atg7 knockdown can regulate the arginine circulation-related pathway proteins of ECA109 cells after radiotherapy, and increasing arginine concentration can enhance radiosensitivity of ECA109 cells.

Key words: autophagy related gene 7-siRNA, Arginine succinate synthetase1, Arginine succinic acid lyase, Esophageal cancer, Arginine, Radiosensitivity

CLC Number: 

  • R310.31
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