小鼠异位心脏移植

Biology

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Summary

小鼠异位心脏移植模型已被证明许多研究者排斥反应和免疫反应机制研究的重要方法。然而,所涉及的技术仍然充满挑战。通过修改标准的技术,我们已与超过1000移植成功。

Cite this Article

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Liu, F., Kang, S. M. Heterotopic Heart Transplantation in Mice. J. Vis. Exp. (6), e238, doi:10.3791/238 (2007).

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Abstract

小鼠异位心脏移植已被广泛使用,因为它是由博士介绍。科里和罗素在1973年。它特别适用于研究排斥反应和免疫反应,现在,新的转基因和基因敲除小鼠可宝贵的,已经开发和大量的免疫试剂。心脏移植模型是严格小于皮肤移植模型,虽然在技术上更具挑战性。我们已经开发出一种改性技术,并已完成超过1000对小鼠异位心脏移植成功的病例。升主动脉和腹主动脉吻合时,两呆缝线放置捐助小号升主动脉与受体腹主动脉的近端和远端的顶点,然后使用主动脉两侧11-0继续缝线缝合吻合。留缝线吻合更容易和11-0是一个理想的缝合的大小,以避免出血和血栓形成。

肺动脉和下腔静脉吻合时,两呆缝线捐助小号肺动脉近端顶点与收件人小号下腔静脉远端心尖。下腔静脉和捐助小号肺动脉左侧壁与下腔静脉内继续缝合封闭后,与近端顶点住宿结缝合下腔静脉的右侧壁和捐助小号肺动脉外下腔洞穴,用10-0缝线缝合继续封闭。这种方法更容易执行,因为只是一方的下腔静脉和10-0缝线吻合,是正确的大小,以避免出血和血栓形成。在这篇文章中,我们提供的技术,以补充视频的细节。

Protocol

捐助者的心脏的制备及收获:

  1. 该鼠标是与腹腔注射巴比妥和手术视野放在仰卧麻醉。
  2. 是一个长期的中线腹部切口。以暴露腹主动脉和下腔静脉,腹腔内容物用纱布缩回到左。
  3. 1毫升肝素(10u/ml)注入下腔静脉肝素。
  4. 开胸脊柱两侧的肋骨上存在分歧,前胸壁杠杆暴露心脏。
  5. 下腔静脉,上腔静脉和奇静脉静脉结扎6-0丝,分为优越的连字。
  6. 升主动脉和肺动脉的分离,分为远端尽可能远。
  7. 肺静脉结扎,分为远端的关系。
  8. 心脏灌注4 C O盐溶液。
  9. 心脏被保存在冰冷的生理盐水溶液。

收件人的制备及移植

  1. 该鼠标是与腹腔注射巴比妥和手术视野放在仰卧麻醉。
  2. 是一个长期的中线腹部切口。缩回腹腔内容外,用纱布腹部暴露腹主动脉和下腔静脉。
  3. 暴露腹主动脉和下腔静脉的分支,并用10-0缝线结扎。
  4. 主动脉和下腔静脉的近端和远端连字周围放置。
  5. venotomy是在用30号针头的收件人的下腔静脉。开幕式,然后再扩展到一个长度等于捐助的微型剪刀肺主动脉。
  6. aortotomy是在30号针头与腹主动脉的收件人。
  7. 打开腹主动脉和下腔静脉盐水灌溉。
  8. 捐赠者的心放在收件人的腹部左侧,并用纱布覆盖。
  9. 留缝线捐助的升主动脉与受体腹主动脉的近端和远端的顶点。
  10. 收件人的腹主动脉和捐助者的升主动脉的右侧吻合完成后继续缝合。
  11. 第一个捐赠者的捐赠者的肺动脉和收件人的下腔静脉吻合肺动脉近端顶点与收件人的下腔静脉远端心尖两呆缝线。
  12. 左墙的下腔静脉和捐赠者的肺动脉与下腔静脉内继续缝合关闭。
  13. 与近端先端留缝合后打结,下腔静脉和捐助的肺动脉的右墙以外的下腔静脉缝合关闭。
  14. 捐赠者的心交给收件人的腹部右侧。收件人的腹主动脉和捐赠者的升主动脉吻合的右墙封闭,继续缝合。
  15. 吻合后,远端结扎删除,检查吻合口出血。
  16. 如果有一点没有出血,吻合口近端结扎删除。捐赠者的心立即充满血液,心的颜色变成红色,并开始再次击败。
  17. 肠子都返回到腹部,这是连续用6-0缝线封闭。
  18. 收件人鼠标放在一个温暖的地方。一小时后,收件人的复苏。

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Discussion

结果

我们已经成功地在小鼠异位心脏移植1000余例,并取得90%以上的成功率在不同品系小鼠,包括野生型,转基因和基因敲除小鼠。

讨论

由于博士。科里和罗素介绍,1973年在小鼠异位心脏移植模型,该模型已被证明是一个特别有价值的研究免疫排斥反应和开发新的免疫策略。这种模式可以更广泛地使用,因为现在出现了大量的转基因和基因敲除小鼠研究排斥/宽容的机制提供了新的途径。血管心脏移植模型,可更切合临床实体器官移植。虽然皮肤移植或新生儿心脏的皮下移植在技术上是简单的,是这些移植的移植物的存活需要的新生血管的同种异体反应特别敏感。因此,听到皮肤及皮下移植可以为一种宽容的严格测试,但不发展或免疫或耐受的诱导协议的优化非常有用。

我们修改了几种方法,以提高效率和成功率血管心脏移植技术:

  1. 捐赠者的心脏应该是从升主动脉盐水灌溉,直到心成为收获后的光色。
  2. 用10-0缝线,防止在操作过程中和手术后血栓形成的出血应结扎腹主动脉和下腔静脉的收件人的分支机构。
  3. 腹主动脉腔和下腔静脉的收件人应灌溉用生理盐水溶液,直至无血离开。这可以防止手术后的血栓形成。
  4. 吻合时,应避免气泡内的船只。
  5. 动脉吻合应该用11-0缝线,以帮助防止从吻合口出血。
  6. 腹主动脉和下腔静脉收件人的切口应该是平等的捐赠者的心脏肺动脉和主动脉开放。
  7. 重要的是有一个清晰和广阔的经营领域,因此使用的夹具6-0缝合,而不是暂时的主动脉和下腔静脉闭塞。

结论

虽然在小鼠异位心脏移植技术是在技术上具有挑战性,这是一个重要的技术研究alloresponses。实践和技术改进,导致在我们的实验室在90%以上的成功率。我们相信,视频和所附的手稿将有助于减轻希望使用这项技术的实验室学习曲线。

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References

  1. Corry, R. obert J., Winn, H. enry J., Russell, P. aul S. Primarily vascularized allografts of heart in mice. Transplantation. 16, 343-350 (1973).

Comments

23 Comments

  1. Hello Mr.Kang, My name is Vered Gigi and i am a graduste student in Israel for transplanted immunology. I am volunteering in a program (The NIR School of the Heart)  that expose young teenage student to the world of science, focusing on the cardiology field. As such i would love to show them this movie which is very nicely done. However, u was unable to download the movie. Is there a way that i can get the movie? Is it possible for you to e-mial me a copy for teaching porposes only? Thank you very much for your attention, Vered Gigi  my e-mail: vered.gigi@gmail.com

    Reply
    Posted by: Anonymous
    December 22, 2008 - 4:12 AM
  2. Hi, The video was produced by JOVE. We don’t have any copies. Sorry about that. Thank you very much for your interests.   Fengchun Liu

    Reply
    Posted by: Anonymous
    April 18, 2009 - 12:45 AM
  3. Dear Mr Kang, your video and technique is very interesting. I'm really interesting in getting also a copy of this video because I would like to perform this surgery. Please, could you please send me a copy of this video by email? if it is not possible, you can maybe send me it by normal mail. My email address is svielc07@gmail.com. Thanks a lot.   Maria

    Reply
    Posted by: Anonymous
    January 22, 2009 - 8:22 AM
  4. Hi there. Thank you for posting this video. Do you ever see paralysis of the hind end after this surgery? How about swelling of the hind feet? What is the maximum time for the vessels to be clamped or tied off?   Thank you for your time. Sheri

    Reply
    Posted by: Anonymous
    February 4, 2009 - 4:14 PM
  5. The maximum time for the vessels to be clamped is 40 minutes, the paralysis or swelling of the hind end may appear if the vessels are clamped over 40 minutes. Fengchun Liu

    Reply
    Posted by: Anonymous
    April 19, 2009 - 2:59 PM
  6. What a wonderful job well done!!! Highly admired your technique Dr. Liu!!!

    Reply
    Posted by: Anonymous
    February 18, 2009 - 9:34 PM
  7. Thank you very much! This vedio is useful for me. I'm Li Zhoubin from Ruijin hospital, shanghai, China. jove.com is wonderful!!

    Reply
    Posted by: Anonymous
    March 7, 2009 - 5:21 AM
  8. &#²1016;²769;&#²407²;: &#²4744;&#²²909;A²81;&#²5105;&#²9616;&#²²31²;&#²7491;&#²²31²;&#²0570;腹腔²²7;&#²6893;的&#²7169;&#²²411;A²9²;&#²59²8;&#²65²4;不&#²6159;&#²4456;&#²²909;A²9²;&#²1487;不&#²1487;&#²0197;&#²5²²6;&#²4744;的video&#²1457;²473;&#²5105;一&#²0²²1;&#²1834;A²9²;非&#²41²0;&#²4863;谢A²81; 谢艾&#²²958;  

    Reply
    Posted by: Anonymous
    April 5, 2009 - 8:27 AM
  9. Great job! Could you tell me the applicable mouse body weight in this transplant surgery? In the video, the mouse looked like 50 g big.
    Thank you.

    Reply
    Posted by: Anonymous
    July 30, 2009 - 12:16 PM
  10. Mice weights are ²5-30g. Thanks.

    Reply
    Posted by: Anonymous
    July 30, 2009 - 1:22 PM
  11. Dear Dr. Kang,
    I enjoyed the procedure and watched it several times. My qyestion is regarding the instrument you were using as the needle holder. Can I know what that is, the brand and specifications?

    thanks,
    A. E. Nia

    Reply
    Posted by: Anonymous
    August 17, 2009 - 5:21 PM
  12. Thanks for the great procedure...My question for you is, what is the type of needle holder you are using? can I get the brand and specifications please...

    thanks
    A.E. Nia

    Reply
    Posted by: Abbas E.
    August 18, 2009 - 2:05 PM
  13. assi needle holder B-13-8. Thanks.

    Reply
    Posted by: Anonymous
    August 18, 2009 - 7:41 PM
  14. Hello,
    Do you heparinize the recepient?
    In my experiments, teh transplanted heart dŒs not start beating...I am suspecting clot formation at the anastomosis site. I heparinize the donor with 0.1 hep plus 0.9 saline... Any thoughts?

    thanks
    Abb

    Reply
    Posted by: Reza N.
    September 18, 2009 - 7:22 PM
  15. We do not heparinize the recepient.

    Reply
    Posted by: Anonymous
    May 20, 2010 - 12:31 PM
  16. Dr. Liu; Great job! I have two more question for you. Firstly, in the step 8 of your Donor Preparation and Heart Harvest, where do you perfuse heart using 4 Co saline solution? Secondly, is the 10-0 suture the nylon suture?
    Thanks a lot!
    Guangming
    gcheng@kumc.edu


    G

    Reply
    Posted by: Guangming C.
    May 19, 2010 - 1:08 AM
  17. Saline solution is injected into the aorta. The 10-0 sutures is nylon suture.

    Reply
    Posted by: Anonymous
    May 20, 2010 - 12:29 PM
  18. Congratulations to this nice video. However I have a question.
    Can you explain why the heart is still beating quite normal at the end of the explant, if you performed a perfusion over the aorta.
    One would think that after a perfusion with cold saline a heart should stop beating.
    Cheers Werner

    Reply
    Posted by: Anonymous
    July 24, 2011 - 4:04 PM
  19. &#²1016;²769;&#²407²;:

    &#²4744;&#²²909;A²81;&#²4744;精&#²8²51;的&#²5²16;ഌ²;&#²0196;&#²0154;&#²5²40;&#²6381;A²9²;不知&#²6159;&#²154²;Ŋ²1;&#²4471;&#²1040;该#²70;频的邮&#²0²14;&#²6469;Ű²7;一&#²7493;&#²3398;&#²0064;?
    谢谢
    &#²647²;

    Reply
    Posted by: arthur y.
    December 22, 2012 - 12:27 AM
  20. Dr. Liu,

    Very decent surgical technique, great job! Just curious how long it will normally take to complete the whole surgical procedure for professional level guys like you? I haven't tried it, but I was informed that it is very time consuming.

    Jian Wu (B.M., M.S.)
    Ph.D Candidate in Biomedical Science (Cardiovascular & Metabolic Diseases Track)
    University of Toledo, College of Medicine and Life Sciences

    Reply
    Posted by: JIAN W.
    May 16, 2013 - 12:00 AM
  21. Impressive idea . but i found couple of things here:
    1- A narration would be very useful!
    2- Apparently sterility is not a goal of this surgery as i saw multiple violations
    3- Harvesting the donor heart takes long , Isn't it better to intubate the donor and have it ventilated during harvest?

    Reply
    Posted by: hadi j.
    March 20, 2014 - 5:47 PM
  22. thanks for your video, could you please tell what micro potts scissors you use for aortotomy and venotomy? as i need to get suitable scissors for my experiments. thanks

    Reply
    Posted by: Anne S.
    June 6, 2015 - 11:36 AM
  23. Dr. Liu,
    Your superb skill is very impressive! Amazing! Thank you for your video!
    But why should we post comment using in English? Would you please email me, we can communicate in Chinese, that would be wonderful! zzh24085@creighton.edu

    Reply
    Posted by: zefu z.
    June 10, 2016 - 4:32 PM

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