Effect of Glut-1 and HIF-1α double knockout by CRISPR/CAS9 on radiosensitivity in laryngeal carcinoma via the PI3K/Akt/mTOR pathway.

Abstract

Hypoxic resistance is the main obstacle to radiotherapy for laryngeal carcinoma. Our previous study indicated that hypoxia-inducible factor 1α (HIF-1α) and glucose transporter 1 (Glut-1) double knockout reduced tumour biological behaviour in laryngeal carcinoma cells. However, their radioresistance mechanism remains unclear. In this study, cell viability was determined by CCK8 assay. Glucose uptake capability was evaluated by measurement of 18 F-fluorodeoxyglucose radioactivity. A tumour xenograft model was established by subcutaneous injection of Tu212 cells. Tumour histopathology was determined by haematoxylin and eosin staining, immunohistochemical staining, and TUNEL assays. Signalling transduction was evaluated by Western blotting. We found that hypoxia induced radioresistance in Tu212 cells accompanied by increased glucose uptake capability and activation of the PI3K/Akt/mTOR pathway. Inhibition of PI3K/Akt/mTOR activity abolished hypoxia-induced radioresistance and glucose absorption. Mechanistic analysis revealed that hypoxia promoted higher expressions of HIF-1α and Glut-1. Moreover, the PI3K/Akt/mTOR pathway was a positive mediator of HIF-1α and/or Glut-1 in the presence of irradiation. HIF-1α and/or Glut-1 knockout significantly reduced cell viability, glucose uptake and PI3K/Akt/mTOR activity, all of which were induced by hypoxia in the presence of irradiation. In vivo analysis showed that knockout of HIF-1α and/or Glut-1 also inhibited tumour growth by promoting cell apoptosis, more robustly compared with the PI3K inhibitor wortmannin, particularly in tumours with knockout of both HIF-1α and Glut-1. HIF-1α and/or Glut-1 knockout also abrogated PI3K/Akt/mTOR signalling transduction in tumour tissues, in a manner similar to wortmannin. HIF-1α and/or Glut-1 knockout facilitated radiosensitivity in laryngeal carcinoma Tu212 cells by regulation of the PI3K/Akt/mTOR pathway.

• Authors: Yang-Yang Bao, Jiang-Tao Zhong, Li-Fang Shen, Li-Bo Dai, Shui-Hong Zhou, Jun Fan, Hong-Tian Yao, Zhong-Jie Lu
• Journal: Journal of cellular and molecular medicine (J Cell Mol Med) Vol. 26 Issue 10 Pg. 2881-2894 (05 2022) ISSN: 1582-4934 [Electronic] England
• PMID: 35415942 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: © 2022 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.

Chemical References

• Glucose Transporter Type 1
• Hypoxia-Inducible Factor 1, alpha Subunit
• SLC2A1 protein, human
• MTOR protein, human
• Proto-Oncogene Proteins c-akt
• TOR Serine-Threonine Kinases
• Glucose
• Wortmannin

Topics

• Animals
• CRISPR-Cas Systems
• Carcinoma (genetics, metabolism, radiotherapy)
• Cell Line, Tumor
• Glucose
• Glucose Transporter Type 1 (genetics, metabolism)
• Humans
• Hypoxia
• Hypoxia-Inducible Factor 1, alpha Subunit (genetics, metabolism)
• Laryngeal Neoplasms (genetics, metabolism, radiotherapy)
• Phosphatidylinositol 3-Kinases (genetics, metabolism)
• Proto-Oncogene Proteins c-akt (genetics, metabolism)
• Radiation Tolerance (genetics)
• TOR Serine-Threonine Kinases (genetics, metabolism)
• Wortmannin