UBC9介导SUMO化修饰在同型半胱氨酸诱导巨噬细胞焦亡中的作用研究
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1.宁夏医科大学总医院老年与特需医学科银川;2.国家卫生健康委员会代谢性心血管疾病研究重点实验室银川;3.宁夏医科大学基础医学院银川;4.宁夏医科大学检验学院银川;5.宁夏医科大学检验学院

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] 国家自然科学基金青年项目(81900273),项目负责人:马胜超;国家自然科学基金面上项目(82060139,82270492),项目负责人:马胜超;宁夏回族自治区重点研发计划项目(2019BEG03006),项目负责人:马胜超;宁夏青年科技拔尖人才工程(马胜超),宁夏自然科学基金优秀青年项目(2023AAC05035),项目负责人:马胜超;宁夏医科大学校级重点项目 (XZ2022004),项目负责人:彭红建


The role of UBC9 mediated SUMO modification in homocysteine-induced pyrodeath of macrophages
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1.Department of geriatrics and special needs medicine,General Hospital of Ningxia Medical University;2.Basic school of medicine, Ningxia Medical University;3.Basic school of medicine,Ningxia Medical University;4.School of Clinical Medicine,Ningxia Medical University;5.School of Clinical Medicine, Ningxia Medical University

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    摘要:

    目的 探讨泛素结合酶9(ubiquitin Conjugating Enzyme 9, UBC9)介导SUMO化修饰在同型半胱氨酸诱导巨噬细胞焦亡中的作用。方法 首先采用细胞计数试剂 (cell counting kit-8,CCK-8) 法和Western blot检测不同浓度(用 0μmol/L、50μmol/L、100μmol/L、150μmol/L、200μmol/L)同型半胱氨酸对小鼠巨噬细胞(RAW264.7)的活力及焦亡的影响;采用Western blot检测不同组细胞中UBC9、SUMO化修饰蛋白SUMO-1、炎症因子IL-1β的表达水平;qRT-PCR检测干扰前后UBC9的mRNA表达,同时检测干扰UBC9后,UBC9、焦亡相关蛋白、IL-1β及SUMO-1相关表达。结果 当采用100μmol/L的Hcy刺激后,巨噬细胞的活力不受影响且焦亡蛋白NLRP3、caspase-1表达最明显( P<0.001);与Control组相比,Hcy组IL-1β的表达水平升高( P<0.01),SUMO-1表达升高( P<0.01);与Control组相比,Hcy组中UBC9蛋白水平及mRNA水平表达均增高( P<0.05);转染si-UBC9后,与si-NC+Hcy组相比si-UBC9+Hcy组NLRP3、caspase-1、IL-1β 、UBC9、SUMO-1表达降低P<0.01)。结论 高同型半胱氨诱导巨噬细胞焦亡的发生,其机制与上调泛素结合酶9发生SUMO化修饰有关。

    Abstract:

    Objective To study the role of ubiquitin Conjugating Enzyme 9 (UBC9) in the pyrodeath of homocysteine-induced macrophages mediated by SUMO modification. Methods Firstly, The effects of homocysteine at different concentrations (0μmol/L, 50μmol/L, 100μmol/L, 150μmol/L, 200μmol/L) on the viability and pyrodeath of mouse macrophages (RAW264.7) were detected by CCK-8 and Western blot ;Western blot was used to detect the expression levels of UBC9, SUMO-modified protein SUMO-1 and inflammatory cytokine IL-1β in different groups of cells. qRT-PCR was used to detect the mRNA expression of UBC9 before and after interference, and to detect the expression of UBC9, pyrogen-related protein and SUCO-1 after interference.Results After stimulation with 100μmol/L Hcy, the activity of macrophages was not affected, and the expressions of NLRP3 and caspase-1 were the most obvious (P < 0.05).Compared with Control group, the expression levels of and IL-1β were increased in Hcy group (P < 0.01), and the expression of SUMO-1 was increased (P < 0.01). Compared with Control group, UBC9 protein level and mRNA level in Hcy group were increased (P < 0.05). After transfection with si-UBC9, the expressions of NLRP3, caspase-1, IL-1β, UBC9 and SUMO-1 in si-UBC9+Hcy group were decreased compared with si-NC+Hcy group (P < 0.01). Conclusion Hyperhomocysteamine (Hcy) induces pyrodeath in macrophages, and its mechanism is related to the up-regulation of ubiquitin binding enzyme 9 to SUMO modification.

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  • 收稿日期:2024-02-27
  • 最后修改日期:2024-03-28
  • 录用日期:2024-06-07
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