The role of cGAS STING pathway regulation of NCOA4 mediated iron autophagy in hypoxia reoxygenation injury of HT22 cells
Zhao Lan
Li Zhenwen
Zhong Zhentong
Zhan Liying
Gao Wenwei
Abstract：Objective To investigate the role of cGAS interferon gene stimulating protein(STING)pathway regula-tion of nuclear receptor co activator 4(NCOA4)mediated iron autophagy in hypoxia reoxygenation injury of mouse hipp-ocampal neurons(HT22).Methods From September 2020 to December 2022,experiments were conducted in the Central La-boratory of People's Hospital of Wuhan University.HT22 cell lines were randomly divided into four groups:control group(Ctrl group),hypoxia reoxygenation group(HR group),hypoxia reoxygenation + cGAS inhibitor(RU.521)group(HR + RU group),and hypoxia reoxygenation + RU.521 + NCOA4 overexpression group(HR + RU + NCOA4 group).The HR group was cultured for 6 hours under sugar free hypoxia conditions and then re cultured for 24 hours to construct a hypoxia reox-ygenation model.The HR+RU group and HR+RU+NCOA4 group were given 3.0 in advance μ After 24 hours of treat-ment with RU.521 at a concentration of mol/L,a hypoxia reoxygenation model was constructed.The HR + RU + NCOA4 group was transfected with NCOA4 lentivirus 5 days before hypoxia reoxygenation,while the control group was cultured routinely.Immunofluorescence was used to detect ROS in each group,ELISA was used to detect CCK8,LDH,SOD,MDA,and ferrous ions,Western blot was used to detect cGAS,STING,NCOA4,LC3B,and ferritin.Results Compared with the Ctrl group,the cell viability CCK8 and SOD in the HR group decreased,while LDH,ROS,MDA,cGAS,STING,NCOA4,LC3B,ferritin,and ferrous ions increased(P<0.05);Compared with the HRgroup,the HR+RUgroup showed an increase in cell viability CCK8,SOD,and ferritin,while LDH,ROS,MDA,cGAS,STING,NCOA4,LC3B,and ferrous ions decreased(P<0.05);Compared with the HR + RU group,the cell viability CCK8,SOD,and ferritin in the HR + RU + NCOA4 group de-creased(P<0.05),while LDH,ROS,MDA,NCOA4,LC3B,and ferrous ions increased.There was no statistically significant difference in cGAS and STING(P>0.05).Conclusion After hypoxia and reoxygenation,the cGAS STINGpathway is up-regulated,and NCOA4 mediated excessive activation of iron autophagy exacerbates cell damage.
Keywords：Iron autophagyHypoxia/reoxygenationHypoxia reoxygenation cGAS-STING pathwayNuclear recep-tor co activator 4Mice
Publication Date：2024-03-18
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：5( 360-364 )