高级检索

  • ISSN 1006-3080
  • CN 31-1691/TQ

AAV介导的TNFR关节内基因治疗对血友病性关节病预防和治疗的实验研究

林振洋 王冶凡 张飞旭 周新月 宗晓莹 吴侠 华宝来 肖啸 孙君江

林振洋, 王冶凡, 张飞旭, 周新月, 宗晓莹, 吴侠, 华宝来, 肖啸, 孙君江. AAV介导的TNFR关节内基因治疗对血友病性关节病预防和治疗的实验研究[J]. 华东理工大学学报(自然科学版). doi: 10.14135/j.cnki.1006-3080.20210429004
引用本文: 林振洋, 王冶凡, 张飞旭, 周新月, 宗晓莹, 吴侠, 华宝来, 肖啸, 孙君江. AAV介导的TNFR关节内基因治疗对血友病性关节病预防和治疗的实验研究[J]. 华东理工大学学报(自然科学版). doi: 10.14135/j.cnki.1006-3080.20210429004
LIN Zhenyang, WANG Yefan, ZHANG Feixu, ZHOU Xinyue, ZONG Xiaoying, WU Xia, HUA Baolai, XIAO Xiao, SUN Junjiang. Adeno-associated Virus Mediated Tumor Necrosis Factor Receptor for the Prevention and Treatment of Hemophilic Arthropathy in Hemophilic Mice[J]. Journal of East China University of Science and Technology. doi: 10.14135/j.cnki.1006-3080.20210429004
Citation: LIN Zhenyang, WANG Yefan, ZHANG Feixu, ZHOU Xinyue, ZONG Xiaoying, WU Xia, HUA Baolai, XIAO Xiao, SUN Junjiang. Adeno-associated Virus Mediated Tumor Necrosis Factor Receptor for the Prevention and Treatment of Hemophilic Arthropathy in Hemophilic Mice[J]. Journal of East China University of Science and Technology. doi: 10.14135/j.cnki.1006-3080.20210429004

AAV介导的TNFR关节内基因治疗对血友病性关节病预防和治疗的实验研究

doi: 10.14135/j.cnki.1006-3080.20210429004
基金项目: 国家自然科学基金资助项目(81970171,82070139),上海市自然科学基金项目(20ZR1415000),中央高校基本科研业务费专项资金资助
详细信息
    作者简介:

    林振洋(1996—),男,硕士生,研究方向为基因治疗。E-mail:reigetsu@163.com

    通讯作者:

    张飞旭,E-mail:zhangfeixu2009@126.com

    孙君江,E-mail:sunjunjiang@ecust.edu.cn

  • 中图分类号: R554+.1

Adeno-associated Virus Mediated Tumor Necrosis Factor Receptor for the Prevention and Treatment of Hemophilic Arthropathy in Hemophilic Mice

  • 摘要: 血友病性关节病(HA)的发病机制与关节内炎性细胞因子过表达有关,TNFα是最重要的炎性因子之一。sTNFR作为TNFα的受体可以特异性地与之结合拮抗其炎性效应。为探讨关节腔内持续表达sTNFR对HA关节的保护作用,将表达TNFR的质粒(pAAV-TNFR:Fc)包装为rAAV5-TNFR:Fc载体,注射到血友病小鼠膝关节内。结果显示炎症持续6周后,关节内仍能表达TNFR:Fc,两个给药组中TNFα和IL-1β的表达不同程度地下降,关节病变均显著减轻,表明该关节内基因治疗对血友病性关节病有预防和治疗效果。

     

  • 图  1  B型血友病小鼠右膝关节进行穿刺处理4周后右膝关节TNFα表达水平的变化

    Figure  1.  Expression level change of TNFα in right knee joints of Hemophilia B mice 4 weeks after the puncture * indicates significance between values of this group and NC(*p<0.05).

    图  2  质粒酶切后电泳结果及rAAV5-TNFR:Fc感染HFLS后TNFR表达的Western Blot结果

    Figure  2.  Electrophoresis of digested plasmid and Western Blot of TNFR in HFLS infected with rAAV5-TNFR:Fc

    图  3  向B型血友病小鼠右膝关节内注射rAAV5-TNFR:Fc 4周后右膝关节组织内的TNFR的表达水平变化

    Figure  3.  Expression level change of TNFR in right knee joints after Hemophilia B mice were intraarticularly injected with rAAV5-TNFR:Fc into right knee joints. * indicates significance between values of this group and NC (**p<0.01)

    图  4  主要动物实验的小鼠分组

    Figure  4.  Mice grouping in the study

    图  5  关节内注射rAAV5-TNFR:Fc对B型血友病小鼠关节内出血后细胞因子的表达水平的影响

    Figure  5.  Effect of intraarticular injection of rAAV5-TNFR:Fc on cytokine expression levels in hemarthrosis joints of Hemophilia B mice. * indicates significance between values of this group and the NC group or between values of two groups (*p<0.05, **, p<0.01, ***p<0.001)

    图  6  关节内注射rAAV5-TNFR:Fc治疗B型血友病小鼠HA,关节组织的H&E染色结果以及滑膜炎评分

    Figure  6.  H&E staining of joint tissue and synovitis scoring after Hemophilia B mice were intraarticular injected with rAAV5-TNFR:Fc to treat HA. * indicates significance between values of this group and the PBS group (*p<0.05)

    图  7  关节内注射rAAV5-TNFR:Fc治疗B型血友病小鼠HA,关节组织的针对F4/80抗原的组化染色结果以及巨噬细胞浸润评分

    Figure  7.  Anti-F4/80 antigen IHC of joint tissue and macrophage infiltration scoring after Hemophilia B mice were intraarticular injected with rAAV5-TNFR:Fc to treat HA. * indicates significance between values of this group and the PBS group (*p<0.05)

    图  8  关节内注射rAAV5-TNFR:Fc治疗B型血友病小鼠HA,关节组织的针对vWF抗原的组化染色结果以及巨噬细胞浸润评分

    Figure  8.  Anti-vWF antigen IHC of joint tissue and macrophage infiltration scoring after Hemophilia B mice were intraarticular injected with rAAV5-TNFR:Fc to treat HA. * indicates significance between values of this group and the PBS group(*p<0.05, **p<0.01).

  • [1] SRIVASTAVA A, SANTAGOSTINO E, DOUGALL A, et al. WFH Guidelines for the Management of Hemophilia, 3rd edition[J]. Haemophilia, 2020, 26(Suppl 6): 1-158. doi: 10.1111/hae.14046
    [2] CHRISTENSEN K R, KJELGAARD-HANSEN M, NIELSEN L N, et al. Rapid inflammation and early degeneration of bone and cartilage revealed in a time-course study of induced haemarthrosis in haemophilic rats[J]. Rheumatology (Oxford), 2019, 58(4): 588-599. doi: 10.1093/rheumatology/key186
    [3] PRATT K P, ARRUDA V R, LACROIX-DESMAZES S. Inhibitors-Recent insights[J]. Haemophilia, 2021, 27(Suppl 3): 28-36. doi: 10.1111/hae.14077
    [4] WOJDASIEWICZ P, PONIATOWSKI L A, NAUMAN P, et al. Cytokines in the pathogenesis of hemophilic arthropathy[J]. Cytokine Growth Factor Rev, 2018, 39: 71-91. doi: 10.1016/j.cytogfr.2017.11.003
    [5] MANETTI M, LINARI S, ROMANO E, et al. TNF-alpha/TNF-R System May Represent a Crucial Mediator of Proliferative Synovitis in Hemophilia A[J]. J Clin Med, 2019, 8(7). doi: 10.3390/jcm8070939
    [6] BRENNAN F M, GIBBONS D L, COPE A P, et al. TNF inhibitors are produced spontaneously by rheumatoid and osteoarthritic synovial joint cell cultures: evidence of feedback control of TNF action[J]. Scand J Immunol, 1995, 42(1): 158-165. doi: 10.1111/j.1365-3083.1995.tb03639.x
    [7] ZHOU X, SHEN L, LIU L, et al. Preclinical safety evaluation of recombinant adeno-associated virus 2 vector encoding human tumor necrosis factor receptor-immunoglobulin Fc fusion gene[J]. Hum Vaccin Immunother, 2016, 12(3): 732-739. doi: 10.1080/21645515.2015.1090070
    [8] WANG F, YU J, WANG Y, et al. Combination therapy with TNFR-Fc and CTLA4-FasL using the recombinant adeno-associated virus potently suppresses adjuvant-induced arthritis in rats[J]. Appl Microbiol Biotechnol, 2015, 99(15): 6327-6337. doi: 10.1007/s00253-015-6459-7
    [9] ZHOU X, GAO K, SHEN L, et al. Modulation of immune and inflammatory responses on experimental arthritis following intraarticular gene transfer of tumor necrosis factor receptor-immunoglobulin Fc[J]. Rheumatol Int, 2012, 32(9): 2605-2614. doi: 10.1007/s00296-011-1974-z
    [10] KHOURY M, ADRIAANSEN J, VERVOORDELDONK M J, et al. Inflammation-inducible anti-TNF gene expression mediated by intra-articular injection of serotype 5 adeno-associated virus reduces arthritis[J]. J Gene Med, 2007, 9(7): 596-604. doi: 10.1002/jgm.1053
    [11] BLOQUEL C, BESSIS N, BOISSIER M-C. Gene Therapy of Collagen-Induced Arthritis by Electrotransfer of Human Tumor Necrosis Factor-a Soluble Receptor I Variants[J]. Hum Gene Ther, 2004, 15: 189-201. doi: 10.1089/104303404772679995
    [12] ZHANG H G, XIE J, YANG P, et al. Adeno-associated virus production of soluble tumor necrosis factor receptor neutralizes tumor necrosis factor alpha and reduces arthritis[J]. Hum Gene Ther, 2000, 11(17): 2431-2442. doi: 10.1089/104303400750038525
    [13] MANTRAVADI S, OGDIE A, KRAFT W K. Tumor necrosis factor inhibitors in psoriatic arthritis[J]. Expert Rev Clin Pharmacol, 2017, 10(8): 899-910. doi: 10.1080/17512433.2017.1329009
    [14] CROFT M, BENEDICT C A, WARE C F. Clinical targeting of the TNF and TNFR superfamilies[J]. Nat Rev Drug Discov, 2013, 12(2): 147-168. doi: 10.1089/hgtb.2012.243
    [15] ZHANG F, XU M, YANG Q, et al. A Translational Study of TNF-Alpha Antagonists as an Adjunctive Therapy for Preventing Hemophilic Arthropathy[J]. J Clin Med, 2019, 9(1). doi: 10.3390/jcm9010075
    [16] VALENTINO L A, HAKOBYAN N. Histological changes in murine haemophilic synovitis: a quantitative grading system to assess blood-induced synovitis[J]. Haemophilia, 2006, 12(6): 654-662. doi: 10.1111/j.1365-2516.2006.01348.x
    [17] HOFFMAN M, HARGER A, LENKOWSKI A, et al. Cutaneous wound healing is impaired in hemophilia B[J]. Blood, 2006, 108(9): 3053-3060. doi: 10.1182/blood-2006-05-020495
    [18] HAXAIRE C, HAKOBYAN N, PANNELLINI T, et al. Blood-induced bone loss in murine hemophilic arthropathy is prevented by blocking the iRhom2/ADAM17/TNF-alpha pathway[J]. Blood, 2018, 132(10): 1064-1074. doi: 10.1182/blood-2017-12-820571
    [19] ZHONG C, SZOLLOSI D, SUN J, et al. Novel Piperazino-Enaminones Decrease Pro-inflammatory Cytokines Following Hemarthrosis in a Hemophilia Mouse Model[J]. Inflammation, 2019, 42(5): 1719-1729. doi: 10.1007/s10753-019-01032-y
    [20] OVLISEN K, KRISTENSEN A T, JENSEN A L, et al. IL-1 beta, IL-6, KC and MCP-1 are elevated in synovial fluid from haemophilic mice with experimentally induced haemarthrosis[J]. Haemophilia, 2009, 15(3): 802-810. doi: 10.1111/j.1365-2516.2008.01973.x
    [21] MATTHIAS G, WAJANT H, ZIMMERMANN G. The type 1 receptor (CD120a) is the high-affinity receptor for soluble tumor necrosis factor[J]. Proc Natl Acad Sci, 1998, 95: 570-575. doi: 10.1073/pnas.95.2.570
    [22] APPARAILLY F, KHOURY M, VERVOORDELDONK M J, et al. Adeno-associated virus pseudotype 5 vector improves gene transfer in arthritic joints[J]. Hum Gene Ther, 2005, 16(4): 426-434. doi: 10.1089/hum.2005.16.426
    [23] LOUIS JEUNE V, JOERGENSEN J A, HAJJAR R J, et al. Pre-existing anti-adeno-associated virus antibodies as a challenge in AAV gene therapy[J]. Hum Gene Ther Methods, 2013, 24(2): 59-67. doi: 10.1089/hgtb.2012.243
    [24] SEN D, CHAPLA A, WALTER N, et al. Nuclear factor (NF)-kappaB and its associated pathways are major molecular regulators of blood-induced joint damage in a murine model of hemophilia[J]. J Thromb Haemost, 2013, 11(2): 293-306. doi: 10.1111/jth.12101
    [25] PULLES A E, MASTBERGEN S C, SCHUTGENS R E, et al. Pathophysiology of hemophilic arthropathy and potential targets for therapy[J]. Pharmacol Res, 2017, 115: 192-199. doi: 10.1016/j.phrs.2016.11.032
    [26] VALENTINO L A, HAKOBYAN N, KAZARIAN T, et al. Experimental haemophilic synovitis: rationale and development of a murine model of human factor VIII deficiency[J]. Haemophilia, 2004, 10(3): 280-287. doi: 10.1111/j.1365-2516.2004.00899.x
    [27] NICHOLS W L, HULTIN M B, JAMES A H, et al. von Willebrand disease (VWD): evidence-based diagnosis and management guidelines, the National Heart, Lung, and Blood Institute (NHLBI) Expert Panel report (USA)[J]. Haemophilia, 2008, 14(2): 171-232. doi: 10.1111/j.1365-2516.2007.01643.x
  • 加载中
图(8)
计量
  • 文章访问数:  57
  • HTML全文浏览量:  38
  • PDF下载量:  5
  • 被引次数: 0
出版历程
  • 收稿日期:  2021-04-29
  • 网络出版日期:  2021-07-27

目录

    /

    返回文章
    返回