南方医科大学学报 ›› 2024, Vol. 44 ›› Issue (7): 1345-1354.doi: 10.12122/j.issn.1673-4254.2024.07.14

• • 上一篇    

STING高表达通过调控TLR4/NF-κB/NLRP3通路和影响炎症与凋亡水平促进小鼠肾脏缺血再灌注损伤

陶怀祥1,2(), 骆金光1,2, 闻志远1, 虞亘明1,2, 苏萧1, 王鑫玮1, 关翰1, 陈志军1()   

  1. 1.蚌埠医学院第一附属医院泌尿外科,安徽 蚌埠 233004
    2.蚌埠医学院慢性疾病免疫学基础与临床安徽省重点实验室,安徽 蚌埠 233030
  • 收稿日期:2023-12-18 出版日期:2024-07-20 发布日期:2024-07-25
  • 通讯作者: 陈志军 E-mail:2240489402@qq.com;byczj@bbmc.edu.cn
  • 作者简介:陶怀祥,在读硕士研究生,E-mail: 2240489402@qq.com
  • 基金资助:
    安徽省自然科学基金重点项目(2008085QH358);慢性疾病免疫学基础与临床安徽省重点实验室开放课题基金(AHIAI2022K01)

High STING expression exacerbates renal ischemia-reperfusion injury in mice by regulating the TLR4/NF-κB/NLRP3 pathway and promoting inflammation and apoptosis

Huaixiang TAO1,2(), Jinguang LUO1,2, Zhiyuan WEN1, Genming YU1,2, Xiao SU1, Xinwei WANG1, Han GUAN1, Zhijun CHEN1()   

  1. 1.Department of Urology, First Affiliated Hospital of Bengbu Medical College, Bengbu 233004, China
    2.Anhui Provincial Key Laboratory of Immunology in Chronic Disease, Bengbu Medical College, Bengbu 233030, China
  • Received:2023-12-18 Online:2024-07-20 Published:2024-07-25
  • Contact: Zhijun CHEN E-mail:2240489402@qq.com;byczj@bbmc.edu.cn

摘要:

目的 探讨STING在肾缺血再灌注损伤(IRI)中的表达水平以及相关作用机制。 方法 在体内水平,将24只C57BL/6小鼠分为假手术组(Sham)、IRI组、IRI+药物溶剂组(IRI+DMSO)、IRI+SN-011组,6只/组。通过肾动脉夹闭方法建立IRI模型,通过血清肌酐和尿素氮检测、PAS染色检测肾组织损伤变化,采用RT-qPCR、ELISA、Western blotting和IHC法检测肾组织中STING、KIM-1、Bcl-2、Bax、caspase-3、TLR4、P65、NLRP3、caspase-1、CD68、MPO、 IL-1β、IL-6、TNF-α的水平。在体外水平,将HK-2细胞分为对照组、缺氧复氧(H/R)组、H/R+药物溶剂组(H/R+DMSO)、H/R+SN-011组,用厌氧包模拟缺氧环境,RT-qPCR和Western blotting法检测STING表达水平,流式细胞术检测各组细胞凋亡率。 结果 在体内水平,与Sham组相比,IRI组的PAS染色显示组织损伤增加(P<0.05),小鼠血清肌酐、尿素氮含量以及组织KIM-1、STING、TLR4、P65、NLRP3、caspase-1、caspase-3、Bax、CD68、MPO、IL-1β、IL-6、TNF-α表达水平升高(P<0.05),Bcl-2水平降低(P<0.05),SN-011抑制STING表达后,逆转了上述结果(P<0.05)。在体外水平,与对照组相比,H/R组STING的mRNA与蛋白水平升高(P<0.05),流式细胞仪检测显示细胞凋亡率上升(P<0.05),SN-011抑制STING表达,细胞凋亡率下降(P<0.05)。 结论 STING在肾脏IRI中表达水平上升,且可通过作用于TLR4/NF-κB/NLRP3通路以及影响炎症与凋亡水平促进肾损伤。

关键词: STING, TLR4, NF-κB, NLRP3, 肾缺血再灌注, 炎症, 凋亡

Abstract:

Objective To investigate renal expression level of STING in mice with renal ischemia-reperfusion injury (IRI) and its regulatory role in IRI. Methods C57BL/6 mice were divided into sham operation group, IRI (induced by clamping the renal artery) model group, IRI+DMSO treatment group, and IRI+SN-011 treatment group. Serum creatinine and blood urea nitrogen of the mice were analyzed, and pathological changes in the renal tissue were assessed with PAS staining. RT-qPCR, ELISA, Western blotting, and immunohistochemistry were used to detect the expression levels of STING, KIM-1, Bcl-2, Bax, caspase-3, TLR4, P65, NLRP3, caspase-1, CD68, MPO, IL-1β, IL-6, and TNF-α in the renal tissues. In the cell experiment, HK-2 cells exposed to hypoxia-reoxygenation (H/R) were treated with DMSO or SN-011, and cellular STING expression levels and cell apoptosis were analyzed using RT-qPCR, Western blotting or flow cytometry. Results In C57BL/6 mice, renal IRI induced obvious renal tissue damage, elevation of serum creatinine and blood urea nitrogen levels and renal expression levels of KIM-1, STING, TLR4, P65, NLRP3, caspase-1, caspase-3, Bax, CD68, MPO, IL-1β, IL-6, and TNF-α, and reduction of Bcl-2 expression level. Treatment of the mouse models with SN-011 for inhibiting STING expression significantly alleviated these changes. In HK-2 cells, H/R exposure caused significant elevation of cellular STING expression and obviously increased cell apoptosis rate, which was significantly lowered by treatment with SN-011. Conclusion Renal STING expression is elevated in mice with renal IRI to exacerbate renal injury by regulating the TLR4/NF-κB/NLRP3 pathway and promoting inflammation and apoptosis in the renal tissues.

Key words: STING, TLR4, NF-κB, NLRP3, ischemia-reperfusion, inflammation, apotosis