RIP3-mediated necroptosis induced by radiation injury in neuronal cells
Yang Songhua, Huang Shixiong, Zeng Biao, Dong Qian, Zhu Xiaocong, Zeng Na, Li Bin, Zhou Guanzhi, Chen Yifang, Yang Huiting, Li Jian, Shi Yingrui
Second Department of Head and Neck Radiotherapy,Hu'nan Cancer Hospital,Department of Radiological Physics and Technology, Hu'nan Cancer Hospital, Changsha 410013, China
Abstract：Objective To observe the presence or absence of necroptosis in PC12 cells after radiation injury, and to detect the expression of receptor-interacting protein 3(RIP3) and evaluate its regulatory effect on necroptosis. Methods PC12 cells were treated with different doses of irradiation and their necroptosis was detected by lactate dehydrogenase (LDH) release at different time points. After pretreatment with necroptosis inhibitor Necrostatin-1(Nec-1), the changes of cell necroptosis were detected by LDH. The expression level of RIP3 after irradiation intervention was detected by Western blot (WB). After pretreatment with the RIP3-specific inhibitor GSK'872, the changes of cell necroptosis were detected by LDH. The best transfection sequence of RIP3 knockout was screened by WB. The cells were divided into the control group, irradiation group, solvent control group, no-load control group and pretreatment group. WB, immunofluorescence staining, MTT, LDH and Annex V-fluorescein Isothiocyanate/Propidium Iodide (AnnexV-FITC/PI) flow cytometry were used for detection and analysis. Results After 4 Gy irradiation, the degree of cell necrosis was the highest after 3 hours of culture, and the expression level of RIP3 protein was up-regulated. The cell necrosis was decreased after Nec-1,GSK'872 and RIP3 gene knockdown pretreatment. Conclusions The radiation injury of 4Gy can induce the necroptosis of PC12 cells, and the most significant effect can be observed when cultured for 3 hours after irradiation. RIP3 is involved in the process of necroptosis of PC12 cells induced by radiation injury, and plays a pivotal positive regulatory role.
Yang Songhua,Huang Shixiong,Zeng Biao et al. RIP3-mediated necroptosis induced by radiation injury in neuronal cells[J]. Chinese Journal of Radiation Oncology, 2020, 29(12): 1124-1129.
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2020, Vol. 29 
