title: Poly(ADP-ribose) polymerase-1 regulates the mechanism of irradiation-induced CNE-2 human nasopharyngeal carcinoma cell autophagy and inhibition of autophagy contributes to the radiation sensitization of CNE-2 cells.
abstract: The aim of the present study was to investigate the role of autophagy in response to ionizing radiation (IR) in CNE-2 human nasopharyngeal carcinoma cells and to demonstrate the function of poly(ADP-ribose) polymerase-1 (PARP-1) in the regulation of IR-induced autophagy. Microtubule-associated protein 1 light chain 3 (LC3) and poly(ADP-ribose) (PAR) were assessed using western blotting. Ultrastructural analysis was performed using transmission electron microscopy (TEM). The percentage of apoptotic cells was assessed by flow cytometry. The MTT method was used to detect cell viability of CNE-2 cells at different time points after IR. Clonogenic survival assays were used to evaluate the radiosensitivity of nasopharyngeal carcinoma cells treated with IR and IR combined with autophagy inhibitor (chloroquine phosphate), with autophagy inducer (rapamycin) or with PARP-1 inhibitor 3-amino benzamide (3AB). IR induced a massive accumulation of autophagosomes detected by TEM and intensified the conversion of cytosolic LC3-I to LC3-II. PARP-1 activation was accompanied by strong upregulation of PAR and LC3-II expression in CNE-2 cells. Compared with radiation alone, chloroquine phosphate (CDP) or 3AB combined with IR significantly decreased cell viability, as well as the autophagic ratio and LC3-II protein levels. Inhibition of autophagy increased radiation-induced apoptosis; rapamycin (RAPA) significantly decreased cell viability as well, but RAPA increased the autophagic ratio and LC3-II protein levels; induction of autophagy increased radiation-induced apoptosis. To conclude, PARP-1 regulates IR-induced autophagy, and PARP-1 inhibitor contributes to the radiation sensitization of CNE-2 cells. Blockade of autophagy with CDP enhanced the cytotoxicity of radiotherapy in CNE-2 cells. This suggests that inhibition of autophagy or PARP-1 may be used as an adjuvant therapy to treat nasopharyngeal carcinoma.
authors.affiliation: Department of Radiation Oncology, Cancer Hospital of Guangxi Medical University and Cancer Institute of Guangxi Zhuang Autonomous Region, Nanning 530021, PR China.
authors.forename: Zhi-Rui
authors.initials: ZR
authors.lastname: Zhou
authors.forename: Xiao-Dong
authors.initials: XD
authors.lastname: Zhu
authors.forename: Wei
authors.initials: W
authors.lastname: Zhao
authors.forename: Song
authors.initials: S
authors.lastname: Qu
authors.forename: Fang
authors.initials: F
authors.lastname: Su
authors.forename: Shi-Ting
authors.initials: ST
authors.lastname: Huang
authors.forename: Jia-Ling
authors.initials: JL
authors.lastname: Ma
authors.forename: Xiao-Yu
authors.initials: XY
authors.lastname: Li
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis; D001343:Autophagy; D001549:Benzamides / Q000494:pharmacology; D002277:Carcinoma; D045744:Cell Line, Tumor / Q000528:radiation effects; D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects; D002738:Chloroquine / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D000077274:Nasopharyngeal Carcinoma; D009303:Nasopharyngeal Neoplasms / Q000532:radiotherapy*; D010588:Phagosomes / Q000378:metabolism / Q000528:radiation effects; D000071137:Poly (ADP-Ribose) Polymerase-1; D011064:Poly Adenosine Diphosphate Ribose / Q000378:metabolism; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011065:Poly(ADP-ribose) Polymerases / Q000502:physiology*; D011836:Radiation Tolerance; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology; D020123:Sirolimus / Q000494:pharmacology
chemical_list: D000970:Antineoplastic Agents; D001549:Benzamides; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D011838:Radiation-Sensitizing Agents; D011064:Poly Adenosine Diphosphate Ribose; C023676:chloroquine diphosphate; D002738:Chloroquine; C025160:3-aminobenzamide; C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D020123:Sirolimus