留言板

尊敬的读者、作者、审稿人, 关于本刊的投稿、审稿、编辑和出版的任何问题, 您可以本页添加留言。我们将尽快给您答复。谢谢您的支持!

姓名
邮箱
手机号码
标题
留言内容
验证码

人脐带间充质干细胞来源的细胞外囊泡增强纤维化肝脏再生能力

雷耘果, 姚嘉, 郑俊, 等. 人脐带间充质干细胞来源的细胞外囊泡增强纤维化肝脏再生能力[J]. 器官移植, 2023, 14(3): 379-388. doi: 10.3969/j.issn.1674-7445.2023.03.009
引用本文: 雷耘果, 姚嘉, 郑俊, 等. 人脐带间充质干细胞来源的细胞外囊泡增强纤维化肝脏再生能力[J]. 器官移植, 2023, 14(3): 379-388. doi: 10.3969/j.issn.1674-7445.2023.03.009
Lei Yunguo, Yao Jia, Zheng Jun, et al. Human umbilical cord mesenchymal stem cell-derived extracellular vesicles enhance the regenerative capability of fibrotic liver[J]. ORGAN TRANSPLANTATION, 2023, 14(3): 379-388. doi: 10.3969/j.issn.1674-7445.2023.03.009
Citation: Lei Yunguo, Yao Jia, Zheng Jun, et al. Human umbilical cord mesenchymal stem cell-derived extracellular vesicles enhance the regenerative capability of fibrotic liver[J]. ORGAN TRANSPLANTATION, 2023, 14(3): 379-388. doi: 10.3969/j.issn.1674-7445.2023.03.009

人脐带间充质干细胞来源的细胞外囊泡增强纤维化肝脏再生能力

doi: 10.3969/j.issn.1674-7445.2023.03.009
基金项目: 

广东省自然科学基金面上项目 2019A1515011698

详细信息
    作者简介:
    通讯作者:

    姚嘉(ORCID:0000-0003-1457-7241),博士,副主任医师,研究方向为肝移植排斥反应、肝移植缺血-再灌注损伤、肝再生,Email:yaojia6@mail.sysu.edu.cn

    郑俊(ORCID:0000-0002-4283-0403),博士,住院医师,研究方向为肝移植排斥反应、肝移植缺血-再灌注损伤、肝再生,Email:zhengj67@mail2.sysu.edu.cn

  • 中图分类号: R329.2, R575

Human umbilical cord mesenchymal stem cell-derived extracellular vesicles enhance the regenerative capability of fibrotic liver

More Information
  • 摘要:   目的  探讨人脐带间充质干细胞来源的细胞外囊泡(hUC-MSC-EV)在纤维化肝脏再生中的作用。  方法  将C57BL/6小鼠随机分为正常肝脏70%肝切除组(Oil+PHx组)、肝纤维化70%肝切除组(CCl4+PHx组)、肝纤维化70%肝切除+间充质干细胞来源的细胞外囊泡(MSC-EV)治疗组(CCl4+PHx+MSC-EV组),每组8只。将LX-2细胞分为磷酸盐缓冲液(PBS)组、转化生长因子(TGF)-β组、TGF-β+MSC-EV组。检测各组小鼠肝部分切除术后丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)、乳酸脱氢酶(LDH)水平,分析各组小鼠肝组织纤维化及增殖相关指标的表达情况。检测各组LX-2细胞表皮细胞生长因子(EGF)、成纤维母细胞生长因子(FGF)、血管内皮生长因子(VEGF)、肝细胞生长因子(HGF)信使RNA(mRNA)的表达水平。观察对小鼠肝脏HGF表达的影响。  结果  与Oil+PHx组比较,CCl4+PHx组小鼠血清AST、ALT、LDH水平升高,纤维化程度较高,天狼星红及α-平滑肌肌动蛋白(α-SMA)染色阳性区域面积增大,α-SMA蛋白表达水平升高;与CCl4+PHx组比较,CCl4+PHx+MSC-EV组小鼠血清AST、ALT、LDH水平下降,纤维化程度较轻,天狼星红及α-SMA染色阳性区域面积缩小,α-SMA蛋白表达水平下降,差异均有统计学意义(均为P < 0.05)。与Oil+PHx组比较,CCl4+PHx组Ki67、增殖细胞核抗原(PCNA)蛋白表达水平降低;与CCl4+PHx组比较,CCl4+PHx+MSC-EV组Ki67、PCNA蛋白表达水平升高,差异均有统计学意义(均为P < 0.05)。与PBS组比较,TGF-β组LX-2细胞内CollagenⅠmRNA表达水平升高,α-SMA蛋白表达水平升高,HGF蛋白表达水平下降;与TGF-β组比较,TGF-β+MSC-EV组LX-2细胞内CollagenⅠmRNA表达水平下降,HGF mRNA和蛋白表达水平升高,α-SMA蛋白表达水平降低,差异均有统计学意义(均为P < 0.05)。CCl4+PHx组HGF蛋白表达水平较Oil+PHx组下降,但差异无统计学意义(P > 0.05);CCl4+PHx+MSC-EV组HGF蛋白表达水平较CCl4+PHx组上升,差异有统计学意义(P < 0.05)。  结论  纤维化肝脏再生能力较正常肝脏减弱,hUC-MSC-EV可以减轻肝纤维化,并可能通过促进活化的肝星状细胞分泌HGF,有效改善纤维化肝脏的肝再生能力。

     

  • 图  1  hUC-MSC细胞及hUC-MSC-EV鉴定

    注:A图为hUC-MSC的形态(×100);B图为hUC-MSC茜素红染色图(×100);C图为hUC-MSC油红O染色图(×100);D~F图为hUC-MSC表面免疫表型的流式细胞图;G图为蛋白质印迹法检测结果。

    Figure  1.  Identification of hUC-MSC and hUC-MSC-EV

    图  2  小鼠肝部分切除术后各组血清AST、ALT、LDH水平变化

    注:与Oil+PHx组比较,aP < 0.05;与CCl4+PHx组比较,bP < 0.05。

    Figure  2.  Changes of serum AST, ALT and LDH of mice among each group after partial hepatectomy

    图  3  各组小鼠部分肝切除术后纤维化相关指标比较

    注:A~C图为小鼠肝组织病理学染色结果(×100);D、E图为蛋白质印迹法检测结果。与Oil+PHx组比较,aP < 0.05;与CCl4+PHx组比较,bP < 0.05。

    Figure  3.  Comparison of fibrosis related indexes of mice among each group after partial hepatectomy

    图  4  各组小鼠部分肝切除术后增殖相关指标比较

    注:A图小鼠肝组织免疫组织化学染色(×100);B图为蛋白质印迹法检测结果。与Oil+PHx组比较,aP < 0.05;与CCl4+PHx组比较,bP < 0.05。

    Figure  4.  Comparison of proliferation-related indexes of mice among each group after partial hepatectomy

    图  5  各组细胞生长因子比较

    注:A图红色为PKH-26染料标记的MSC-EV,绿色为LX-2细胞的标志性抗原α-SMA,蓝色为细胞核(免疫荧光,×400);B~F图为RT-qPCR检测结果;G~I图为蛋白质印迹法检测结果;J图为ELISA检测结果。与PBS组比较,aP < 0.05;与TGF-β组比较,bP < 0.05。

    Figure  5.  Comparison of cell growth factor among each group

    图  6  各组小鼠肝脏HGF表达情况

    注:与CCl4+PHx组比较,aP < 0.05。

    Figure  6.  HGF expression in liver of mice among each group

  • [1] 谢闰鹏, 谷明旗, 张凤博, 等. 肝移植手术技术的现状和展望[J]. 器官移植, 2022, 13(1): 105-110. DOI: 10.3969/j.issn.1674-7445.2022.01.016.

    XIE RP, GU MQ, ZHANG FB, et al. Current status and prospect of surgical technique of liver transplantation[J]. Organ Transplant, 2022, 13(1): 105-110. DOI: 10.3969/j.issn.1674-7445.2022.01.016.
    [2] 夏强, 沙朦. 活体肝移植的进展与展望[J]. 中华消化外科杂志, 2022, 21(1): 39-42. DOI: 10.3760/cma.j.cn115610-20211205-00622.

    XIA Q, SHA M. Progress and prospect of living donor liver transplantation[J]. Chin J Dig Surg, 2022, 21(1): 39-42. DOI: 10.3760/cma.j.cn115610-20211205-00622.
    [3] AHMED O, DOYLE MBM. Liver transplantation: expanding the donor and recipient pool[J]. Chin Clin Oncol, 2021, 10(1): 6. DOI: 10.21037/cco-20-212.
    [4] GAO SL, MOU BY, DAI DS, et al. Marginal donor liver versus standard donor liver: a single center-observational study[J]. Hepatobiliary Pancreat Dis Int, 2022, DOI: 10.1016/j.hbpd.2022.10.005[Epubaheadofprint].
    [5] 郭义威, 庞盼姣, 孙永琨. miR-217负调控转化生长因子β受体Ⅱ抑制肝纤维化发生及发展的研究[J]. 中华肝脏病杂志, 2022, 30(7): 752-757. DOI: 10.3760/cma.j.cn501113-20200203-00026.

    GUO YW, PANG PJ, SUN YK. Study of the negative regulation of transforming growth factor beta type Ⅱ receptor to inhibit the occurrence and development of liver fibrosis with miR-217[J]. Chin J Hepatol, 2022, 30(7): 752-757. DOI: 10.3760/cma.j.cn501113-20200203-00026.
    [6] 刘世恒, 赵素贤, 张玉果. 肝纤维化发生机制及其中药逆转治疗的研究进展[J]. 临床肝胆病杂志, 2022, 38(10): 2373-2377. DOI: 10.3969/j.issn.1001-5256.2022.10.033.

    LIU SH, ZHAO SX, ZHANG YG. Recent research progress and mechanisms on traditional Chinese medicine reversal therapy of liver fibrosis[J]. J Clin Hepatol, 2022, 38(10): 2373-2377. DOI: 10.3969/j.issn.1001-5256.2022.10.033.
    [7] KISSELEVA T, BRENNER D. Molecular and cellular mechanisms of liver fibrosis and its regression[J]. Nat Rev Gastroenterol Hepatol, 2021, 18(3): 151-166. DOI: 10.1038/s41575-020-00372-7.
    [8] ZHAO Y, YE W, WANG YD, et al. HGF/c-Met: a key promoter in liver regeneration[J]. Front Pharmacol, 2022, 13: 808855. DOI: 10.3389/fphar.2022.808855.
    [9] RIZVI F, EVERTON E, SMITH AR, et al. Murine liver repair via transient activation of regenerative pathways in hepatocytes using lipid nanoparticle-complexed nucleoside-modified mRNA[J]. Nat Commun, 2021, 12(1): 613. DOI: 10.1038/s41467-021-20903-3.
    [10] 李璇, 吴敏超, 段伟娜, 等. 肝细胞生长因子与肝再生的研究进展[J]. 吉林医学, 2021, 42(4): 983-985. https://www.cnki.com.cn/Article/CJFDTOTAL-JLYX202104088.htm

    LI X, WU MC, DUAN WN, et al. Research progress on hepatocyte growth factor and liver regeneration[J]. Jilin Med J, 2021, 42(4): 983-985. https://www.cnki.com.cn/Article/CJFDTOTAL-JLYX202104088.htm
    [11] 李淑萍, 许晓燕, 孙箴, 等. 黄芪多糖和黄芪总甙调控LX-2细胞因子分泌[J]. 浙江大学学报(医学版), 2007, 36(6): 543-548. DOI: 10.3785/j.issn.1008-9292.2007.06.005.

    LI SP, XU XY, SUN Z, et al. Astragalus polysaccharides and astragalosides regulate cytokine secretion in LX-2 cell line[J]. J Zhejiang Univ (Med Sci), 2007, 36(6): 543-548. DOI: 10.3785/j.issn.1008-9292.2007.06.005.
    [12] ZHAO R, WANG L, WANG T, et al. Inhalation of MSC-EVs is a noninvasive strategy for ameliorating acute lung injury[J]. J Control Release, 2022, 345: 214-230. DOI: 10.1016/j.jconrel.2022.03.025.
    [13] ZHAO M, LIU S, WANG C, et al. Mesenchymal stem cell-derived extracellular vesicles attenuate mitochondrial damage and inflammation by stabilizing mitochondrial DNA[J]. ACS Nano, 2021, 15(1): 1519-1538. DOI: 10.1021/acsnano.0c08947.
    [14] PSARAKI A, NTARI L, KARAKOSTAS C, et al. Extracellular vesicles derived from mesenchymal stem/stromal cells: the regenerative impact in liver diseases[J]. Hepatology, 2022, 75(6): 1590-1603. DOI: 10.1002/hep.32129.
    [15] 周志鸿, 彭立辉. 间充质干细胞来源的细胞外囊泡治疗创伤性脑损伤的研究进展[J/CD]. 中华细胞与干细胞杂志(电子版), 2021, 11(4): 251-255. DOI: 10.3877/cma.j.issn.2095-1221.2021.04.009.

    ZHOU ZH, PENG LH. Research progress of extracellular vesicles derived from mesenchymal stem cells in the treatment of traumatic brain injury[J/CD]. Chin J Cell Stem Cell (Electr Edit), 2021, 11(4): 251-255. DOI: 10.3877/cma.j.issn.2095-1221.2021.04.009.
    [16] MITCHELL C, WILLENBRING H. A reproducible and well-tolerated method for 2/3 partial hepatectomy in mice[J]. Nat Protoc, 2008, 3(7): 1167-1170. DOI: 10.1038/nprot.2008.80.
    [17] DONG S, CHEN QL, SONG YN, et al. Mechanisms of CCl4-induced liver fibrosis with combined transcriptomic and proteomic analysis[J]. J Toxicol Sci, 2016, 41(4): 561-572. DOI: 10.2131/jts.41.561.
    [18] SHRESTHA N, CHAND L, HAN MK, et al. Glutamine inhibits CCl4 induced liver fibrosis in mice and TGF-β1 mediated epithelial-mesenchymal transition in mouse hepatocytes[J]. Food Chem Toxicol, 2016, 93: 129-137. DOI: 10.1016/j.fct.2016.04.024.
    [19] YAGI S, HIRATA M, MIYACHI Y, et al. Liver regeneration after hepatectomy and partial liver transplantation[J]. Int J Mol Sci, 2020, 21(21): 8414. DOI: 10.3390/ijms21218414.
    [20] LEE J, GARCIA V, NAMBIAR SM, et al. Pregnancy facilitates maternal liver regeneration after partial hepatectomy[J]. Am J Physiol Gastrointest Liver Physiol, 2020, 318(4): G772-G780. DOI: 10.1152/ajpgi.00125.2019.
    [21] MU M, ZUO S, WU RM, et al. Ferulic acid attenuates liver fibrosis and hepatic stellate cell activation via inhibition of TGF-β/Smad signaling pathway[J]. Drug Des Devel Ther, 2018, 12: 4107-4115. DOI: 10.2147/DDDT.S186726.
    [22] XU L, HUI AY, ALBANIS E, et al. Human hepatic stellate cell lines, LX-1 and LX-2: new tools for analysis of hepatic fibrosis[J]. Gut, 2005, 54(1): 142-151. DOI: 10.1136/gut.2004.042127.
    [23] MICHALOPOULOS GK, BHUSHAN B. Liver regeneration: biological and pathological mechanisms and implications[J]. Nat Rev Gastroenterol Hepatol, 2021, 18(1): 40-55. DOI: 10.1038/s41575-020-0342-4.
    [24] MAO SA, GLORIOSO JM, NYBERG SL. Liver regeneration[J]. Transl Res, 2014, 163(4): 352-562. DOI: 10.1016/j.trsl.2014.01.005.
    [25] HU C, WU Z, LI L. Mesenchymal stromal cells promote liver regeneration through regulation of immune cells[J]. Int J Biol Sci, 2020, 16(5): 893-903. DOI: 10.7150/ijbs.39725.
    [26] FAN S, GAO Y, QU A, et al. YAP-TEAD mediates PPAR α-induced hepatomegaly and liver regeneration in mice[J]. Hepatology, 2022, 75(1): 74-88. DOI: 10.1002/hep.32105.
    [27] HAGIWARA K, HARIMOTO N, YAMANAKA T, et al. A new liver regeneration molecular mechanism involving hepatic stellate cells, Kupffer cells, and glucose-regulated protein 78 as a new hepatotrophic factor[J]. J Hepatobiliary Pancreat Sci, 2023, 30(2): 165-176. DOI: 10.1002/jhbp.1183.
    [28] KAMM DR, MCCOMMIS KS. Hepatic stellate cells in physiology and pathology[J]. J Physiol, 2022, 600(8): 1825-1837. DOI: 10.1113/JP281061.
    [29] SIAPATI EK, ROUBELAKIS MG, VASSILOPOULOS G. Liver regeneration by hematopoietic stem cells: have we reached the end of the road?[J]. Cells, 2022, 11(15): 2312. DOI: 10.3390/cells11152312.
    [30] ZHANG W, CONWAY SJ, LIU Y, et al. Heterogeneity of hepatic stellate cells in fibrogenesis of the liver: insights from single-cell transcriptomic analysis in liver injury[J]. Cells, 2021, 10(8): 2129. DOI: 10.3390/cells10082129.
    [31] GE JY, ZHENG YW, TSUCHIDA T, et al. Hepatic stellate cells contribute to liver regeneration through galectins in hepatic stem cell niche[J]. Stem Cell Res Ther, 2020, 11(1): 425. DOI: 10.1186/s13287-020-01942-x.
    [32] 鲁苏日古嘎, 刘霆, 朱单单, 等. 肝纤维化相关信号通路及其相应的抗肝纤维化药物研究进展[J]. 临床肝胆病杂志, 2022, 38(5): 1161-1164. DOI: 10.3969/j.issn.1001-5256.2022.05.039.

    LU SRGG, LIU T, ZHU DD, et al. Research advances in hepatic fibrosis related signal pathways and anti-hepatic fibrosis drugs[J]. J Clin Hepatol, 2022, 38(5): 1161-1164. DOI: 10.3969/j.issn.1001-5256.2022.05.039.
    [33] MA L, WEI J, ZENG Y, et al. Mesenchymal stem cell-originated exosomal circDIDO1 suppresses hepatic stellate cell activation by miR-141-3p/PTEN/AKT pathway in human liver fibrosis[J]. Drug Deliv, 2022, 29(1): 440-453. DOI: 10.1080/10717544.2022.2030428.
    [34] KIM J, LEE C, SHIN Y, et al. sEVs from tonsil-derived mesenchymal stromal cells alleviate activation of hepatic stellate cells and liver fibrosis through miR-486-5p[J]. Mol Ther, 2021, 29(4): 1471-1486. DOI: 10.1016/j.ymthe.2020.12.025.
    [35] CHENG N, KIM KH, LAU LF. Senescent hepatic stellate cells promote liver regeneration through IL-6 and ligands of CXCR2[J]. JCI Insight, 2022, 7(14): e158207. DOI: 10.1172/jci.insight.158207.
  • 加载中
图(7)
计量
  • 文章访问数:  406
  • HTML全文浏览量:  210
  • PDF下载量:  47
  • 被引次数: 0
出版历程
  • 收稿日期:  2023-01-20
  • 刊出日期:  2023-05-15

目录

    /

    返回文章
    返回