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下调XBP1s通过Sirt3/SOD2/mtROS轴减轻缺氧/复氧诱导的肾小管上皮细胞衰老

彭宣, 倪海强, 顾世琦, 等. 下调XBP1s通过Sirt3/SOD2/mtROS轴减轻缺氧/复氧诱导的肾小管上皮细胞衰老[J]. 器官移植, 2024, 15(1): 46-54. doi: 10.3969/j.issn.1674-7445.2023186
引用本文: 彭宣, 倪海强, 顾世琦, 等. 下调XBP1s通过Sirt3/SOD2/mtROS轴减轻缺氧/复氧诱导的肾小管上皮细胞衰老[J]. 器官移植, 2024, 15(1): 46-54. doi: 10.3969/j.issn.1674-7445.2023186
Peng Xuan, Ni Haiqiang, Gu Shiqi, et al. Down-regulation of XBP1s alleviates the senescence of renal tubular epithelial cells induced by hypoxia/reoxygenation through Sirt3/SOD2/mtROS signaling pathway[J]. ORGAN TRANSPLANTATION, 2024, 15(1): 46-54. doi: 10.3969/j.issn.1674-7445.2023186
Citation: Peng Xuan, Ni Haiqiang, Gu Shiqi, et al. Down-regulation of XBP1s alleviates the senescence of renal tubular epithelial cells induced by hypoxia/reoxygenation through Sirt3/SOD2/mtROS signaling pathway[J]. ORGAN TRANSPLANTATION, 2024, 15(1): 46-54. doi: 10.3969/j.issn.1674-7445.2023186

下调XBP1s通过Sirt3/SOD2/mtROS轴减轻缺氧/复氧诱导的肾小管上皮细胞衰老

doi: 10.3969/j.issn.1674-7445.2023186
基金项目: 国家自然科学基金(82170772、82370759);湖北陈孝平科技发展基金会青年科学专项基金(CXPJJH122001-2210)
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    作者简介:
    通讯作者:

    宫念樵(ORCID 0000-0001-7634-1440),教授,主任医师,博士研究生导师,研究方向为器官移植、移植免疫、干细胞治疗,Email:nqgong@tjh.tjmu.edu.cn

  • 中图分类号: R617, R692

Down-regulation of XBP1s alleviates the senescence of renal tubular epithelial cells induced by hypoxia/reoxygenation through Sirt3/SOD2/mtROS signaling pathway

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  • 摘要:   目的   探讨剪接型X-盒结合蛋白1(XBP1s)在缺氧/复氧(H/R)诱导的原代肾小管上皮细胞衰老中的作用及机制。  方法   将原代肾小管上皮细胞分为空白对照组(NC组)、H/R组、空载腺病毒阴性对照组(Ad-shNC组)、靶向沉默XBP1s腺病毒组(Ad-shXBP1s组)、空载腺病毒+H/R处理组(Ad-shNC+H/R组)、靶向沉默XBP1s腺病毒+H/R处理组(Ad-shXBP1s+H/R组)。检测NC组、H/R组、Ad-shNC组、Ad-shXBP1s组XBP1s的表达情况。检测Ad-shNC组、Ad-shNC+H/R组、Ad-shXBP1s+H/R组β-半乳糖苷酶染色情况,细胞衰老标志物p53、p21、γH2AX表达情况,氧化应激相关指标活性氧(ROS)、丙二醛(MDA)和超氧化物歧化酶(SOD)水平。采用染色质免疫共沉淀验证XBP1s转录调控沉默信息调节因子3(Sirt3),检测下调XBP1s后Sirt3及下游SOD2表达,采用流式细胞术检测线粒体活性氧簇(mtROS)。  结果   与NC组比较,H/R组XBP1s表达增多;与Ad-shNC组比较,Ad-shXBP1s组XBP1s表达减少(均为P<0.001)。与Ad-shNC组比较,Ad-shNC+H/R组β-半乳糖苷酶染色阳性细胞数增加,p53、p21、γH2AX表达增多,ROS、MDA、mtROS水平升高,SOD活性下降,Sirt3表达量降低,Ac-SOD2/SOD2比值升高;与Ad-shNC+H/R组相比,Ad-shXBP1s+H/R组β-半乳糖苷酶染色阳性细胞数减少,p53、p21、γH2AX表达减少,ROS、MDA、mtROS水平下降,SOD活性升高,Sirt3表达量升高,Ac-SOD2/SOD2比值下降(均为P<0.05)。  结论   下调XBP1s可减轻H/R诱导的原代肾小管上皮细胞衰老,可能是通过Sirt3/SOD2/mtROS信号轴发挥作用。

     

  • 图  1  H/R对肾小管上皮细胞XBP1s表达的影响

    注:A、C图为各组XBP1s mRNA相对表达量;B、D图为蛋白质印迹法检测各组XBP1s蛋白表达。与NC组相比,aP<0.001;与Ad-shNC组比较,bP<0.001。

    Figure  1.  The effect of H/R on XBP1s expression in renal tubular epithelial cells

    图  2  下调XBP1s减轻H/R诱导的细胞衰老

    注:A图为检测各组β-半乳糖苷酶阳性细胞数(×200);B图为蛋白质印迹法检测各组p53、p21、γH2AX蛋白表达。

    Figure  2.  Downregulation of XBP1s mitigates cell senescence induced by H/R

    图  3  下调XBP1s减轻H/R诱发的氧化应激

    注:A图为用DCFH-DA荧光探针检测各组ROS水平(免疫荧光,×200);B图为各组ROS荧光强度统计结果;C图为各组MDA含量检测结果;D图为各组SOD活性检测结果。与Ad-shNC组比较,aP<0.01;与Ad-shNC+H/R组比较,bP<0.05。

    Figure  3.  Downregulation of XBP1s alleviates oxidative stress induced by H/R

    图  4  下调XBP1s对Sirt3/SOD2/mtROS信号轴的影响

    注:A图为ChIP-seq峰图;B图为ChIP-qPCR结果,与IgG组比较,aP<0.01;C图为各组Sirt3 mRNA相对表达量;D图为蛋白质印迹法检测各组Sirt3、Ac-SOD2、SOD2蛋白表达;E图为各组Ac-SOD2/SOD2蛋白相对表达量;F图为流式细胞术分析mtROS变化;G图为各组mtROS荧光强度分析结果。与Ad-shNC组比较,aP<0.01,与Ad-shNC+H/R组比较,bP<0.01。

    Figure  4.  The impact of downregulating XBP1s on the Sirt3/SOD2/mtROS signaling axis

    表  1  引物序列

    Table  1.   Primer sequences

    名称 正义链(5'-3') 反义链(5'-3')
    XBP1s AAGAACACGCTTGGGAATGG CTGCACCTGCTGCGGAC
    Sirt3 ATCCCGGACTTCAGATCCCC CAACATGAAAAAGGGCTTGGG
    β-actin AGGCCAACCGTGAAAGATG TGGCGTGAGGGAGAGCATAG
    引物1 TGGTGACTTATCTACGTCTGCT ATCACTTTGTGCCACGGACT
    引物2 CATCGTCTGAGCTGTCTGGTA TGTGGGACTGAGGGCTTTTTG
    引物3 CAGCGTCAACTCCCACTCT AAGGCTGAAGGCATCCGTTTC
    引物4 CCTGACCCTCCGGACTGAT TTGTTCCCAGCTCTCTGCAAT
    引物5 ACTAAGCAGTCAGAACCGGC GTTACCTTGACGACCGCAAC
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  • [1] PONTICELLI C, REGGIANI F, MORONI G. Delayed graft function in kidney transplant: risk factors, consequences and prevention strategies[J]. J Pers Med, 2022, 12(10): 1557. DOI: 10.3390/jpm12101557.
    [2] GRANATA S, VOTRICO V, SPADACCINO F, et al. Oxidative stress and ischemia/reperfusion injury in kidney transplantation: focus on ferroptosis, mitophagy and new antioxidants[J]. Antioxidants (Basel), 2022, 11(4): 769. DOI: 10.3390/antiox11040769.
    [3] 李晓凤, 张国欣, 杨开银, 等. 核因子E2相关因子2在肾缺血-再灌注损伤中的作用[J]. 器官移植, 2023, 14(5): 656-661. DOI: 10.3969/j.issn.1674-7445.2023124.

    LI XF, ZHANG GX, YANG KY, et al. Effect of nuclear factor E2-related factor 2 on renal ischemia-reperfusion injury[J]. Organ Transplant, 2023, 14(5): 656-661. DOI: 10.3969/j.issn.1674-7445.2023124.
    [4] CHEN C, ZHENG M, HOU H, et al. Cellular senescence in ischemia/reperfusion injury[J]. Cell Death Discov, 2022, 8(1): 420. DOI: 10.1038/s41420-022-01205-z.
    [5] DOCHERTY MH, O'SULLIVAN ED, BONVENTRE JV, et al. Cellular senescence in the kidney[J]. J Am Soc Nephrol, 2019, 30(5): 726-736. DOI: 10.1681/ASN.2018121251.
    [6] 郑博文, 刘华亭, 范晓阳, 等. 肾小管上皮细胞损伤促进肾纤维化的研究进展[J]. 中国医药科学, 2023, 13(7): 54-57. DOI: 10.3969/j.issn.2095-0616.2023.07.015.

    ZHENG BW, LIU HT, FAN XY, et al. Research progress of renal tubular epithelial cells injury in promoting renal fibrosis[J]. China Med Pharm, 2023, 13(7): 54-57. DOI: 10.3969/j.issn.2095-0616.2023.07.015.
    [7] YAN M, SHU S, GUO C, et al. Endoplasmic reticulum stress in ischemic and nephrotoxic acute kidney injury[J]. Ann Med, 2018, 50(5): 381-390. DOI: 10.1080/07853890.2018.1489142.
    [8] 王立堃, 李桃, 徐芬芬. 未折叠蛋白响应的激活机制[J]. 生物化学与生物物理进展, 2023, 50(5): 877-891. DOI: 10.16476/j.pibb.2023.0137.

    WANG LK, LI T, XU FF, et al. The mechanism of the unfolded protein response activation[J]. Prog Biochem Biophys, 2023, 50(5): 877-891. DOI: 10.16476/j.pibb.2023.0137.
    [9] CHEN Y, BRANDIZZI F. IRE1: ER stress sensor and cell fate executor[J]. Trends Cell Biol, 2013, 23(11): 547-555. DOI: 10.1016/j.tcb.2013.06.005.
    [10] PARK SM, KANG TI, SO JS. Roles of XBP1s in transcriptional regulation of target genes[J]. Biomedicines, 2021, 9(7): 791. DOI: 10.3390/biomedicines9070791.
    [11] NI H, OU Z, WANG Y, et al. XBP1 modulates endoplasmic reticulum and mitochondria crosstalk via regulating NLRP3 in renal ischemia/reperfusion injury[J]. Cell Death Discov, 2023, 9(1): 69. DOI: 10.1038/s41420-023-01360-x.
    [12] GAO J, FENG Z, WANG X, et al. Sirt3/SOD2 maintains osteoblast differentiation and bone formation by regulating mitochondrial stress[J]. Cell Death Differ, 2018, 25(2): 229-240. DOI: 10.1038/cdd.2017.144.
    [13] TAO R, VASSILOPOULOS A, PARISIADOU L, et al. Regulation of MnSOD enzymatic activity by Sirt3 connects the mitochondrial acetylome signaling networks to aging and carcinogenesis[J]. Antioxid Redox Signal, 2014, 20(10): 1646-1654. DOI: 10.1089/ars.2013.5482.
    [14] QIU X, BROWN K, HIRSCHEY MD, et al. Calorie restriction reduces oxidative stress by Sirt3-mediated SOD2 activation[J]. Cell Metab, 2010, 12(6): 662-667. DOI: 10.1016/j.cmet.2010.11.015.
    [15] ZHU M, HE J, XU Y, et al. AMPK activation coupling SENP1-Sirt3 axis protects against acute kidney injury[J]. Mol Ther, 2023, 31(10): 3052-3066. DOI: 10.1016/j.ymthe.2023.08.014.
    [16] ELEFTHERIADIS T, PISSAS G, FILIPPIDIS G, et al. The role of indoleamine 2, 3-dioxygenase in renal tubular epithelial cells senescence under anoxia or reoxygenation[J]. Biomolecules, 2021, 11(10): 1522. DOI: 10.3390/biom11101522.
    [17] LUO C, ZHOU S, ZHOU Z, et al. Wnt9a promotes renal fibrosis by accelerating cellular senescence in tubular epithelial cells[J]. J Am Soc Nephrol, 2018, 29(4): 1238-1256. DOI: 10.1681/ASN.2017050574.
    [18] VALENTIJN FA, KNOPPERT SN, MARQUEZ-EXPOSITO L, et al. Cellular communication network 2 (connective tissue growth factor) aggravates acute DNA damage and subsequent DNA damage response-senescence-fibrosis following kidney ischemia reperfusion injury[J]. Kidney Int, 2022, 102(6): 1305-1319. DOI: 10.1016/j.kint.2022.06.030.
    [19] CHEN J, LU H, WANG X, et al. VNN1 contributes to the acute kidney injury-chronic kidney disease transition by promoting cellular senescence via affecting RB1 expression[J]. FASEB J, 2022, 36(9): e22472. DOI: 10.1096/fj.202200496RR.
    [20] LI C, SHEN Y, HUANG L, et al. Senolytic therapy ameliorates renal fibrosis postacute kidney injury by alleviating renal senescence[J]. FASEB J, 2021, 35(1): e21229. DOI: 10.1096/fj.202001855RR.
    [21] ZHANG J, ZHANG J, NI H, et al. Downregulation of XBP1 protects kidney against ischemia-reperfusion injury via suppressing HRD1-mediated NRF2 ubiquitylation[J]. Cell Death Discov, 2021, 7(1): 44. DOI: 10.1038/s41420-021-00425-z.
    [22] ZHANG J, XIANG H, LIU J, et al. Mitochondrial sirtuin 3: new emerging biological function and therapeutic target[J]. Theranostics, 2020, 10(18): 8315-8342. DOI: 10.7150/thno.45922.
    [23] WANG D, CAO L, ZHOU X, et al. Mitigation of honokiol on fluoride-induced mitochondrial oxidative stress, mitochondrial dysfunction, and cognitive deficits through activating AMPK/PGC-1α/Sirt3[J]. J Hazard Mater, 2022, 437: 129381. DOI: 10.1016/j.jhazmat.2022.129381.
    [24] XIAO L, FANG Z, WANG Q, et al. Curcumin ameliorates age-induced tight junction impaired in porcine sertoli cells by inactivating the NLRP3 inflammasome through the AMPK/Sirt3/SOD2/mtROS signaling pathway[J]. Oxid Med Cell Longev, 2023: 1708251. DOI: 10.1155/2023/1708251.
    [25] ELEFTHERIADIS T, PISSAS G, GOLFINOPOULOS S, et al. Inhibition of malate dehydrogenase-2 protects renal tubular epithelial cells from anoxia-reoxygenation-induced death or senescence[J]. Biomolecules, 2022, 12(10): 1415. DOI: 10.3390/biom12101415.
    [26] QIN Z, WANG H, DOU Q, et al. Protective effect of fluoxetine against oxidative stress induced by renal ischemia-reperfusion injury via the regulation of miR-450b-5p/Nrf2 axis[J]. Aging (Albany NY), 2022,DOI: 10.18632/aging.204289[Epub ahead of print
    [27] CABRAL-MIRANDA F, TAMBURINI G, MARTINEZ G, et al. Unfolded protein response IRE1/XBP1 signaling is required for healthy mammalian brain aging[J]. EMBO J, 2022, 41(22): e111952. DOI: 10.15252/embj.2022111952.
    [28] HUANG C, WU S, JI H, et al. Identification of XBP1-u as a novel regulator of the MDM2/p53 axis using an shRNA library[J]. Sci Adv, 2017, 3(10): e1701383. DOI: 10.1126/sciadv.1701383.
    [29] WU QJ, ZHANG TN, CHEN HH, et al. The sirtuin family in health and disease[J]. Signal Transduct Target Ther, 2022, 7(1): 402. DOI: 10.1038/s41392-022-01257-8.
    [30] 白玉杰, 王建辉, 吴东颖. 烟酰胺腺嘌呤二核苷酸和Sirtuins在衰老和疾病中的作用[J]. 中国生物化学与分子生物学报, 2022, 38(10): 1294-1303. DOI: 10.13865/j.cnki.cjbmb.2022.04.1562.

    BAI YJ, WANG JH, WU DY. Roles of NAD+ and sirtuins in aging and disease[J]. Chin J Biochem Mol Biol, 2022, 38(10): 1294-1303. DOI: 10.13865/j.cnki.cjbmb.2022.04.1562.
    [31] SUNG JY, KIM SG, KANG YJ, et al. Metformin mitigates stress-induced premature senescence by upregulating AMPKα at Ser485 phosphorylation induced Sirt3 expression and inactivating mitochondrial oxidants[J]. Mech Ageing Dev, 2022, 206: 111708. DOI: 10.1016/j.mad.2022.111708.
    [32] MIAO HH, LIU Q, WANG N, et al. The effect of Sirt3/Ac-SOD2 mediated oxidative stress and HCN1 channel activity on anesthesia/surgery induced anxiety-like behavior in mice[J]. Front Med (Lausanne), 2022, 9: 783931. DOI: 10.3389/fmed.2022.783931.
    [33] 刘恋, 夏中元, 李冰玉, 等. Sirt3过表达对高糖小鼠海马神经元缺氧复氧损伤的影响: 与SOD2的关系[J]. 中华麻醉学杂志, 2021, 41(5): 621-624. DOI: 10.3760/cma.j.cn131073.20210312.00526.

    LIU L, XIA ZY, LI BY, et al. Effect of Sirt3 overexpression on hypoxia-reoxygenation injury to hippocampal neurons of mice exposed to high glucose: relationship with SOD2[J]. Chin J Anesthesiol, 2021, 41(5): 621-624. DOI: 10.3760/cma.j.cn131073.20210312.00526.
    [34] MA LL, KONG FJ, DONG Z, et al. Hypertrophic preconditioning attenuates myocardial ischaemia-reperfusion injury by modulating Sirt3-SOD2-mROS-dependent autophagy[J]. Cell Prolif, 2021, 54(7): e13051. DOI: 10.1111/cpr.13051.
    [35] NING L, RUI X, GUORUI L, et al. A novel mechanism for the protection against acute lung injury by melatonin: mitochondrial quality control of lung epithelial cells is preserved through Sirt3-dependent deacetylation of SOD2[J]. Cell Mol Life Sci, 2022, 79(12): 610. DOI: 10.1007/s00018-022-04628-0.
    [36] CHEN ML, ZHU XH, RAN L, et al. Trimethylamine-N-oxide induces vascular inflammation by activating the NLRP3 inflammasome through the Sirt3-SOD2-mtROS signaling pathway[J]. J Am Heart Assoc, 2017, 6(9): e006347. DOI: 10.1161/JAHA.117.006347.
    [37] PI H, XU S, REITER RJ, et al. Sirt3-SOD2-mROS-dependent autophagy in cadmium-induced hepatotoxicity and salvage by melatonin[J]. Autophagy, 2015, 11(7): 1037-1051. DOI: 10.1080/15548627.2015.1052208.
    [38] LI Q, LIAO J, CHEN W, et al. NAC alleviative ferroptosis in diabetic nephropathy via maintaining mitochondrial redox homeostasis through activating Sirt3-SOD2/Gpx4 pathway[J]. Free Radic Biol Med, 2022, 187: 158-170. DOI: 10.1016/j.freeradbiomed.2022.05.024.
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  • 收稿日期:  2023-09-20
  • 网络出版日期:  2023-11-29
  • 刊出日期:  2024-01-11

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