一种改进的袖带技术用于小鼠颈椎异位心脏移植模型
Summary
在本协议中，使用小鼠心脏移植模型来研究心脏同种异体移植排斥的机制。在这种异位心脏移植模型中，提高了手术效率，并使用改进的袖带技术对心脏植入的颈椎端到端吻合确保了心脏移植物的存活。
Abstract
心脏同种异体移植排斥反应限制了心脏移植后患者的长期生存。小鼠心脏移植模型是临床前研究中研究心脏同种异体移植排斥反应机制的理想选择，因为它们与人类基因具有高度同源性。这种理解将有助于开发独特的方法来改善用心脏同种异体移植物治疗的患者的长期生存率。在小鼠模型中，腹部供体心脏植入通常使用缝线对受体的主动脉和下腔静脉进行端到端吻合。在该模型中，通过改良袖带技术通过端到端吻合将供体的心脏植入受体的颈动脉和颈静脉。移植手术无需缝合即可进行，因此可以增加接受者的生存，因为不会干扰下半身的血液供应和静脉反流。该小鼠模型将有助于研究心脏同种异体移植物的免疫和病理（急性/慢性）排斥反应的机制。
Introduction
心脏移植已成为终末期心力衰竭的标准治疗方法。每年有超过5，500例心脏移植在国际心肺移植学会注册的组织中进行。在同种异体心脏移植受者中，1 年排斥率仍为 >10%，而 3 年排斥率上升至 36%1，2。然而，对于心脏同种异体移植排斥反应患者缺乏有效的预防性治疗。因此，有必要进行动物模型研究，阐明心脏同种异体移植物免疫和病理排斥反应的生理机制。这些研究将有助于研究开发有效药物所需的新靶点，这将有助于防止心脏同种异体移植排斥反应并提高这些患者群体的存活率。
最近在异位心脏移植的小鼠模型研究中提出了心脏同种异体移植排斥反应的一些潜在的免疫和病理生理机制3，4，5。因此，小鼠异位心脏移植因其与人类基因的高度同源性而成为研究心脏移植后心脏同种异体移植物发生免疫排斥和病理损伤机制的理想临床前模型。普遍的概念是在小鼠模型中使用缝线在受体主动脉和下腔静脉中进行腹部端到端吻合，类似于正常的人体解剖结构。但是，此过程可能会干扰接受者的血液供应和下半身的静脉反流6。因此，本文提出了一种改良的小鼠模型中异位心脏移植程序。
捐赠者的心脏通过使用改进的袖带技术通过端到端颈吻合术植入接受者的颈动脉和颈静脉。这种改良的程序促进了手术的可行性，并确保了心脏移植物的存活，而不会干扰下半身的血液供应和静脉反流。
Protocol
所有动物实验均按照国家研究委员会（美国）2011年《实验动物护理和使用指南》第八版进行。涉及动物受试者的程序已获得中国重庆大学肿瘤医院动物护理和使用委员会的批准。从商业来源获得的重20-30g的雄性BALB / c和C57BL / 6小鼠用于同种异体心脏移植研究。C57BL/6小鼠作为供体和同系受体，BALB/c小鼠作为同种异体受体。该协议的原理图如图1所示。注意：手?…
Representative Results
在该小鼠宫颈异位心脏移植模型中，受体小鼠的存活率约为95.2%（21只小鼠中有20只存活）。死亡的主要原因是术后出血。具有规律节律的快速心跳可作为植入供体心脏存活的指标。 C57BL/6和BALB/c小鼠在该模型中分别为MHC（H-2b）和MHC（H-2d）类型，分别为7，8。这两种菌株因H-2而异，H-2导致急性T细胞介导的排斥反应9?…
Discussion
小鼠心脏移植模型有助于研究心脏移植后的排斥机制，有助于开发独特的方法来改善心脏同种异体移植受者的长期生存率。然而，小鼠心脏移植是一项复杂且具有挑战性的任务，需要高水平的显微外科技术，特别是在血管吻合术中11，12，13。小鼠腹部异位心脏移植模型是通过供体主动脉和肺动脉与受体主动脉和下腔静脉吻合缝…
Disclosures
The authors have nothing to disclose.
Acknowledgements
这项工作得到了中国国家自然科学基金（81870304）对李军的支持。
Materials
| 5-0 Polyamide Monofilament suture | B.Braun Medical Inc. | C3090954 | |
| 8-0 Polyamide Monofilament suture | B.Braun Medical Inc. | C2090880 | |
| 10-0 Polyamide Monofilament suture | B.Braun Medical Inc. | G0090781 | |
| 22 G polyurethane cuff | B.Braun Medical Inc. | 4251628-02 | |
| 26 G polyurethane cuff | Suzhou Linhua Medical Instrument Co., LTD | REF383713 | |
| Anesthesia induction chamber | RWD Life Science Co., LTD | V100 | |
| Atraumatic microvascular clamp | Beyotime | FS500 | |
| BALB/c and C57BL/6 mice (20–30 g) | Centre of Experimental Animals (Army Medical University, Chongqing, China) | ||
| Buprenorphine | US Biological life Sciences | 352004 | |
| Electrocoagulator | Guangzhou Runman Medical Instrument Co., LTD | ZJ1099 | |
| Gauze | Henan piaoan group Co., LTD | 10210402 | |
| Heating pad | Guangzhou Dewei Biological Technology Co., LTD | DK0032 | |
| Heparin | North China Pharmaceutical Co., LTD | 2101131-2 | |
| HTK solution | Shenzhen Changyi Pharmaceutical Co., LTD | YZB/Min8263-2013 | |
| Injection syringe (10 mL) | Shandong weigao group medical polymer Co., LTD | 20211001 | |
| Isoflurane | RWD Life Science Co., LTD | 21070201 | |
| Physiological saline | Southwest pharmaceutical Co., LTD | H50021610 | |
| Scalp needle | Hongyu Medical Group | 20183150210 | |
| Shaver | Beyotime | FS600 | |
| Small animal anesthesia machine | RWD Life Science Co., LTD | R500 | |
| Surgical operation microscope | Tiannuoxiang Scientific Instrument Co. , Ltd, Beijing, China | SZX-6745 | |
| Swab | Yubei Medical Materials Co., LTD | 21080274 |
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Cite This Article
Mao, X., Xian, P., You, H., Huang, G., Li, J. A Modified Cuff Technique for Mouse Cervical Heterotopic Heart Transplantation Model. J. Vis. Exp. (180), e63504, doi:10.3791/63504 (2022).