文章摘要
环泊酚通过上调应激诱导蛋白2表达减轻肾脏细胞氧糖剥夺/复糖复氧损伤
Ciprofol alleviates injury of oxygen-glucose deprivation/reglucose and reoxygenation in renal cells by up-regulating sestrin 2 expression
  
DOI：10.12089/jca.2025.07.013
中文关键词: 应激诱导蛋白2，人类  铁死亡  人肾皮质近曲小管上皮细胞  氧糖剥夺/复糖复氧  环泊酚
英文关键词: Sestrin 2 protein, human  Ferroptosis  Human kidney-2 cells  Oxygen-glucose deprivation/reoxygenation  Ciprofol
基金项目:黔西南州医学科研联合项目（2023-3-16）
作者单位E-mail
刘美言 550004,贵阳市,贵州医科大学附属医院麻醉科  
宋涛 550004,贵阳市,贵州医科大学附属医院麻醉科  
闵洁煜 550004,贵阳市,贵州医科大学附属医院麻醉科  
应流念 贵州医科大学麻醉学院  
刘磊 550004,贵阳市,贵州医科大学附属医院麻醉科  
钟毅 550004,贵阳市,贵州医科大学附属医院麻醉科 490173559@qq.com 
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中文摘要:
      
目的：探究环泊酚调控应激诱导蛋白2（SESN2）表达量及其在氧糖剥夺/复糖复氧（OGD/R）诱导的HK-2细胞铁死亡中的作用。
方法：选择野生型及SESN2基因敲低的人肾皮质近曲小管上皮细胞（HK-2细胞株），将所选细胞分为五组：正常培养组（C组）、OGD/R组（O组）、OGD/R+环泊酚50 μmol/L组（OC组）、SESN2基因敲低+OGD/R组（SO组）和SESN2敲低+OGD/R+环泊酚50 μmol/L组（SOC组）。其中O组、OC组、SO组和SOC组建立OGD/R细胞损伤模型。采用CCK-8法检测细胞存活率，荧光酶标仪检测ROS荧光强度。收集细胞，采用微板法检测丙二醛（MDA）含量，采用比色法检测谷胱甘肽（GSH）、半胱氨酸（Cys）含量；收集细胞培养上清液，采用ELISA法检测白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）、乳酸脱氢酶（LDH）浓度。提取细胞蛋白和细胞RNA，分别采用Western blot和qRT-PCR法检测SESN2、GPX4、FSP1蛋白含量和mRNA表达量。
结果：与C组比较，O组、OC组、SO组和SOC组细胞存活率、GSH、Cys含量、GPX4、FSP1蛋白含量和mRNA表达量明显降低（P＜0.05），ROS水平、MDA含量、细胞培养上清液IL-6、TNF-α、LDH浓度、SESN2蛋白含量和mRNA表达量明显升高（P＜0.05）。与O组比较，OC组细胞存活率、GSH、Cys含量、SESN2、GPX4、FSP1蛋白含量和mRNA表达量明显升高（P＜0.05），ROS水平、MDA含量、细胞培养上清液IL-6、TNF-α、LDH浓度明显降低（P＜0.05）；SO组细胞存活率、GSH、Cys含量、SESN2、GPX4、FSP1蛋白含量和mRNA表达量明显降低（P＜0.05），ROS水平、MDA含量、细胞培养上清液IL-6、TNF-α、LDH浓度明显升高（P＜0.05）。与OC组比较，SOC组细胞存活率、GSH、Cys含量、SESN2、GPX4、FSP1蛋白含量和mRNA表达量明显降低（P＜0.05），ROS水平、MDA含量、细胞培养上清液IL-6、TNF-α、LDH浓度明显升高（P＜0.05）。与SO组比较，SOC组细胞存活率、GSH、Cys含量、SESN2、GPX4、FSP1蛋白含量和mRNA表达量明显升高（P＜0.05），ROS水平、MDA含量、细胞培养上清液IL-6、TNF-α、LDH浓度明显降低（P＜0.05）。
结论：环泊酚可以减轻HK-2细胞OGD/R损伤，降低铁死亡水平和细胞氧化应激反应，可能与上调SESN2表达量有关。
英文摘要:
      
Objective: To explore the regulation of sestrin 2 (SESN2) expression by ciprofol and its role in ferroptosis in HK-2 cells induced by oxygen-glucose deprivation/reglucose and reoxygenation (OGD/R).
Methods: The wild type and SESN2 knockdown HK-2 cell line were selected and divided into five groups: normal culture group (group C), OGD/R group (group O), OGD/R + ciprofol 50 μmol/L group (OC group), and SESN2 knockdown + OGD/R + ciprofol 50 μmol/L group (SO group), and SESN2 knockdown + OGD/R + ciprofol 50 μmol/L group (SOC group). OGD/R damage model was established in groups O, OC, SO and SOC by OGD/R method. The cell survival rate was detected by CCK-8 method, and the fluorescence intensity of ROS was measured using a fluorescence microplate reader. The cells were collected, and the MDA content was detected by microplate method, and the GSH and Cys contents were detected by colorimetric method. The supernatant of the cell culture was collected, and the contents of IL-6, TNF-α, and LDH were detected by ELISA. After extracting cellular proteins and cellular RNA, the protein contents and mRNA expression levels of SESN2, GPX4, and FSP1 were detected by Western blot and qRT-PCR, respectively.
Results: Compared with group C, the cell survival rate, contents of GSH and Cys, and the protein contents and mRNA expression levels of GPX4 and FSP1 in groups O, OC, SO, and SOC were significantly decreased (P < 0.05), while the levels of ROS, the content of MDA, the concentrations of IL-6, TNF-α, and LDH in the cell culture supernatant, the protein contents and mRNA expression levels of SESN2 were significantly increased (P < 0.05). Compared with group O, the cell survival rate, the contents of GSH and Cys, the protein contents and mRNA expression levels of SESN2, GPX4, and FSP1 in group OC were significantly increased (P < 0.05), while the ROS level, the content of MDA, and the concentrations of IL-6, TNF-α, and LDH in the cell culture supernatant were significantly decreased (P < 0.05). Compared with group O, the cell survival rate, the contents of GSH and Cys, protein contents and mRNA expression levels of SESN2, GPX4, and FSP1 in group SO were significantly decreased (P < 0.05), while the ROS level, the content of MDA, and the concentrations of IL-6, TNF-α, and LDH in the cell culture supernatant were significantly increased (P < 0.05). Compared with group OC, the cell survival rate, the contents of GSH and Cys, the protein contents and mRNA expression levels of SESN2, GPX4, and FSP1 in group SOC were significantly decreased (P < 0.05), while the ROS level, the content of MDA, and the concentrations of IL-6, TNF-α, and LDH in the cell culture supernatant were significantly increased (P < 0.05). Compared with group SO, the cell survival rate, the contents of GSH and Cys, the protein contents and mRNA expression levels of SESN2, GPX4, and FSP1 in group SOC were significantly increased (P < 0.05), while the ROS level, the content of MDA, and the concentrations of IL-6, TNF-α, and LDH in the cell culture supernatant were significantly decreased (P < 0.05).
Conclusion: Cyclopofol can alleviate OGD/R injury in HK-2 cells, reduce the level of ferroptosis and cellular oxidative stress response, which may be related to the upregulation of SESN2 expression.
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