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下调XBP1s通过抑制ITPR介导的线粒体功能障碍改善肾小管上皮细胞缺氧/复氧损伤

倪海强, 彭宣, 顾世琦, 等. 下调XBP1s通过抑制ITPR介导的线粒体功能障碍改善肾小管上皮细胞缺氧/复氧损伤[J]. 器官移植. doi: 10.3969/j.issn.1674-7445.2023198
引用本文: 倪海强, 彭宣, 顾世琦, 等. 下调XBP1s通过抑制ITPR介导的线粒体功能障碍改善肾小管上皮细胞缺氧/复氧损伤[J]. 器官移植. doi: 10.3969/j.issn.1674-7445.2023198
Ni Haiqiang, Peng Xuan, Gu Shiqi, et al. Down-regulating XBP1s alleviates hypoxia/reoxygenation injury of renal tubular epithelial cells by inhibiting ITPR-mediated mitochondrial dysfunction[J]. ORGAN TRANSPLANTATION. doi: 10.3969/j.issn.1674-7445.2023198
Citation: Ni Haiqiang, Peng Xuan, Gu Shiqi, et al. Down-regulating XBP1s alleviates hypoxia/reoxygenation injury of renal tubular epithelial cells by inhibiting ITPR-mediated mitochondrial dysfunction[J]. ORGAN TRANSPLANTATION. doi: 10.3969/j.issn.1674-7445.2023198

下调XBP1s通过抑制ITPR介导的线粒体功能障碍改善肾小管上皮细胞缺氧/复氧损伤

doi: 10.3969/j.issn.1674-7445.2023198
基金项目: 国家自然科学基金(82170772),湖北陈孝平科技发展基金会青年科学专项基金(CXPJJH122001-2210)
详细信息
    作者简介:
    通讯作者:

    宫念樵(ORCID 0000-0001-7634-1440),教授,主任医师,研究方向为器官移植,Email: nqgong@tjh.tjmu.edu.cn

  • 中图分类号: R617, R692

Down-regulating XBP1s alleviates hypoxia/reoxygenation injury of renal tubular epithelial cells by inhibiting ITPR-mediated mitochondrial dysfunction

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  • 摘要:   目的  探讨剪接型X盒结合蛋白1(XBP1s)对小鼠肾小管上皮细胞缺氧/复氧(H/R)损伤的影响及其作用机制。  方法  将小鼠肾小管上皮细胞分为腺病毒阴性对照组(Ad-shNC组)、靶向沉默XBP1s腺病毒组(Ad-shXBP1s组)、Ad-shNC+H/R组、Ad-shXBP1s+H/R组。检测各组细胞凋亡水平、线粒体活性氧活性、线粒体膜电位及线粒体钙离子水平。使用染色质免疫共沉淀测序(ChIP-seq)分析XBP1s调控肌醇1,4,5-三磷酸受体(ITPR)家族的结合位点。检测各组XBP1s和ITPR家族信使RNA(mRNA)和蛋白表达水平。  结果  与Ad-shNC组比较,Ad-shNC+H/R组细胞凋亡水平更高,线粒体活性氧产生增多,线粒体膜电位降低,线粒体钙离子水平升高;与Ad-shNC+H/R组比较,Ad-shXBP1s+H/R组细胞凋亡水平较低,线粒体活性氧产生减少,线粒体膜电位升高,线粒体钙离子水平降低(均为P<0.05)。与Ad-shNC组比较,Ad-shXBP1s组XBP1s、ITPR1、ITPR2和ITPR3 mRNA和蛋白相对表达量降低(均为P<0.05)。与Ad-shNC组相比,Ad-shNC+H/R组XBP1s、ITPR1、ITPR2和ITPR3蛋白相对表达量升高;与Ad-NC+H/R组相比,Ad-shXBP1s+H/R组XBP1s、ITPR1、ITPR2和ITPR3蛋白相对表达量下降(均为P<0.05)。ChIP-seq结果显示,XBP1s能够结合ITPR1的启动子和外显子、ITPR2外显子和ITPR3外显子。  结论  XBP1s可能通过直接调控ITPR转录和翻译而影响线粒体相关的内质网膜结构功能,下调XBP1s能够抑制ITPR表达,改善线粒体损伤。

     

  • 图  1  各组细胞形态学变化(×100,×200)

    Figure  1.  Cell morphological changes in each group

    图  2  各组细胞凋亡水平

    注:与Ad-shNC组比较,aP>0.05;与Ad-shNC+H/R组比较,bP<0.05。

    Figure  2.  Apoptosis levels of cells in each group

    图  3  各组细胞线粒体活性氧水平

    注:与Ad-shNC组比较,aP<0.05;与Ad-shNC+H/R组比较,bP<0.05。

    Figure  3.  Mitochondrial reactive oxygen species levels of cells in each group

    图  4  各组细胞线粒体膜电位水平

    注:与Ad-shNC组比较,aP<0.05;与Ad-shNC+H/R组比较,bP<0.05。

    Figure  4.  Mitochondrial membrane potential levels of cells in each group

    图  5  各组细胞线粒体钙离子水平

    注:与Ad-shNC组比较,aP<0.05,与Ad-shNC+H/R组比较,bP<0.05。

    Figure  5.  Mitochondrial calcium levels of cells in each group

    图  6  染色质免疫共沉淀测序峰图

    Figure  6.  Chromatin immunoprecipitation sequencing peak maps

    图  7  各组XBP1s和ITPR家族mRNA和蛋白表达水平

    注:A图为各组mRNA表达水平;B、C图为各组蛋白表达水平。与Ad-shNC组比较,aP<0.05;与Ad-shNC+H/R组比较,bP<0.05。

    Figure  7.  The mRNA and protein expression levels of XBP1s and ITPR family in each group

    表  1  引物序列

    Table  1.   Primer sequences

    基因 序列(5’→ 3’)
    XBP1s 正义链 AAGAACACGCTTG GGAATGG
    反义链 CTGCACCTGCTGCGGAC
    ITPR1 正义链 CTCTGTATGCGGAGGGATCTAC
    反义链 CAGGATACTTAGCTATGAGGCG
    ITPR2 正义链 CTTCCTCTACATTGGGGACATC
    反义链 GGGATACTTAGCTATGAGACGG
    ITPR3 正义链 CGACCTCATCAAGGCTCTCC
    反义链 TCCCCATCTCGTTGTTCTGC
    β-actin 正义链 AGGCCAACCGTGAAAGATG
    反义链 TGGCGTGAGGGAGAGCATAG
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  • 收稿日期:  2023-10-07
  • 网络出版日期:  2024-01-05

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