肾缺血再灌注损伤：损伤的小鼠模型和再生
Summary
The mouse model of renal ischaemia reperfusion injury described here comprises of a right nephrectomy that provides control tissue and clamping of the left renal pedicle to induce ischaemia that results in acute kidney injury. This model uses a midline laparotomy approach with all steps performed via one incision.
Abstract
肾移植术中肾缺血再灌注损伤（IRI）是急性肾损伤（AKI）患者和肾血流量的阻塞的常见原因是不可避免的。实验模型，准确和可重复地概括肾IRI在解剖AKI的病理生理和新的治疗药物的发展是至关重要的。这里介绍的是肾IRI小鼠模型，结果在重复性的AKI。这是通过对与一个切口允许两个右肾切除术，它提供控制组织和左肾蒂夹紧诱导左肾缺血的手术中线剖腹手术的方法来实现。通过在缺血期间仔细监测夹紧位置和体温的这种模式实现了可重复的功能和结构的损伤。小鼠处死24小时以下手术示范肾功能的丧失与血清或血浆肌酸酐水平的升高，以及结构郭晓丽肾损害与急性肾小管坏死明显。肾功能改善和肾IRI这样，该模型可用于研究肾再生7天期间的急性组织损伤消退。已动用肾IRI的这个模型，研究AKI的分子和细胞病理生理学，以及随后的肾再生的分析。
Introduction
缺血再灌注损伤（IRI）是受伤的多个器官，包括肾脏，心脏和大脑的共同模式。肾IRI可能导致急性肾损伤（AKI）患者，并没有具体的治疗方法。 AKI作为IRI的结果具有复杂的发病机理涉及两个先天和适应性免疫应答1。肾IRI的实验模型提供解剖的关键细胞，并参与了AKI的发病机制，以及随后的肾再生随之而来的是在随后的日子里调解员的潜力。此外后疾病进程的新的治疗剂的效果进行评估。
这里所描述的肾IRI的实验模型的总体目标是诱导急性功能和结构的肾损伤。一些研究者已经利用涉及双边IRI 2的诱导的模型。虽然双侧肾IRI模型的使用，单方面仁人IRI模型有一个右肾切除正在开展在手术时间的优势。右侧肾切除的组织可作为有价值的对照组织中的研究涉及要么诱导或抑制的基因或蛋白质的表达的预处理步骤。例如，我们用这个模型来评估血红素精氨酸的预处理效果（HA）注射肾IRI 3 24小时前交回。成功诱导细胞保护酶血红素加氧酶-1（HO-1）由医管局IRI之前在右肾切除对照组织4中得到证实。房委会通过的HO-1依赖的机制减少肾IRI在老年小鼠中的一部分。类似地，我们已经使用了模型中巨噬细胞的耗竭研究，探讨巨噬细胞在肾IRI 5的作用。右侧肾切除组织的免疫组织化学分析可用于确定消融方法的疗效。右侧肾切除的组织，因此可以用来同时确认和量化i的平nduction或抑制在每个个体实验动物感兴趣分子的。这个模型将是感兴趣的谁正在使用的药物或其他化合物来调节肾IRI的诱导前基因或蛋白质等的表达研究。
其他研究已经使用侧面切口访问肾脏。这里所描述的模型使用一个单一的中线腹部手术同时执行的右肾切除，诱导左肾缺血再灌注损伤。这种手术方式提供了出色的外科领域，包括肾蒂和颜色的变化遵循肾蒂夹紧的可视化。我们的出版经验与模型4-6表明，小鼠很快从手术中恢复了接近100％的存活率。
最后，该模型在一个为期7天的动力学分析表明，该模型具有两个恢复肾功能和肾小管完整性，用显著肾小管细胞增殖。
Protocol
Representative Results
Discussion
肾IRI是AKI的一个重要原因，没有提供具体的治疗。肾脏IRI的实验研究已经在以前的工作表明巨噬细胞，树突细胞，淋巴细胞，调节性T细胞以及其他细胞和两者的急性损伤和愈合阶段5,8中感应介质的作用非常翔实– 16。此外，实验肾脏IRI已被用于评估各种治疗剂4,17-19的效果。
肾IRI这里详述的模型使用一个中线剖腹手术的方法来进行右肾切除，并用夹子?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
The present study was supported by grants from Kidney Research UK (ST4/2011), the Cunningham Trust (CT11/14) and the Mrs EA Hogg’s Charitable Trust.
Materials
| Name of Material/ Equipment | Company | Catalog Number | Comments/Description |
| Tissue scissors | Fine Science Tools | 14072 – 10 | |
| Micro-Adson forceps (Rat toothed) | Fine Science Tools | 11019 – 12 | |
| S&T JFA-5bTC Forceps – SuperGrip Angled | Fine Science Tools | 00649-11 | |
| Colibri retractor | Fine Science Tools | 17000 – 04 | |
| Micro clip applicator | Fine Science Tools | 18057-14 | |
| Micro serrafines | Fine Science Tools | 18055-04 | |
| Olsen-Hegar needle holder | Fine Science Tools | 12002 – 12 | |
| Hemoclip Plus Ligating Clips Small | Weck | 533837 | |
| Autoclip Wound Clip System, 9mm | Harvard Apparatus | PY2 52-3748 | |
| Silk Black Braided Suture, Size 6-0 | Harvard Apparatus | 723288 | |
| Standard Heat Matt | |||
| Homeothermic Blanket & Control Unit | Harvard Apparatus | ||
| Lacri-Lube | Allergan | ||
| Vetasept Chlorhexidine | AnimalCare | ||
| Vetalar : Ketamine hydrochloride | 100mg/ml solution | ||
| Domitor : medetomidine hydrochloride | 1mg/ml | ||
| Vetergesic : Buprenorphine hydrochloride | 0.3mg/ml | ||
| Antisedan : Atipamezole hydrochoride | 5mg/ml | ||
References
- Kinsey, G. R., Li, L., Okusa, M. D. Inflammation in acute kidney injury. Nephron Exp Nephrol. 109, (2008).
- Wei, Q., Dong, Z. Mouse model of ischemic acute kidney injury: technical notes and tricks. Am J Physiol Renal Physiol. 303, (2012).
- Ferenbach, D. A., Kluth, D. C., Hughes, J. Hemeoxygenase-1 and renal ischaemia-reperfusion injury. Nephron Exp Nephrol. 115, (2010).
- Ferenbach, D. A., et al. The induction of macrophage hemeoxygenase-1 is protective during acute kidney injury in aging mice. Kidney Int. 79, 966-976 (2011).
- Ferenbach, D. A., et al. Macrophage/monocyte depletion by clodronate, but not diphtheria toxin, improves renal ischemia/reperfusion injury in mice. Kidney Int. 82, 928-933 (2012).
- Ferenbach, D. A., et al. Macrophages expressing heme oxygenase-1 improve renal function in ischemia/reperfusion injury. Mol Ther. 18, 1706-1713 (2010).
- Keppler, A., et al. Plasma creatinine determination in mice and rats: an enzymatic method compares favorably with a high-performance liquid chromatography assay. Kidney Int. 71, 74-78 (2007).
- Vinuesa, E., et al. Macrophage involvement in the kidney repair phase after ischaemia/reperfusion injury. J Pathol. 214, 104-113 (2008).
- Friedewald, J. J., Rabb, H. Inflammatory cells in ischemic acute renal failure. Kidney Int. 66, 486-491 (2004).
- Gandolfo, M. T., et al. Foxp3+ regulatory T cells participate in repair of ischemic acute kidney injury. Kidney Int. 76, 717-729 (2009).
- Burne, M. J., et al. Identification of the CD4+ T cell as a major pathogenic factor in ischemic acute renal failure. J Clin Invest. , 1283-1290 (2001).
- Burne-Taney, M. J., et al. B cell deficiency confers protection from renal ischemia reperfusion injury. J Immunol. 171, 3210-3215 (2003).
- Kinsey, G. R., et al. Regulatory T cells suppress innate immunity in kidney ischemia-reperfusion injury. J Am Soc Nephrol. 20, 1744-1753 (2009).
- Li, L., Okusa, M. D. Macrophages, dendritic cells, and kidney ischemia-reperfusion injury. Semin Nephrol. 30, 268-277 (2010).
- Lin, S. L., et al. Macrophage Wnt7b is critical for kidney repair and regeneration. Proc Natl Acad Sci U S A. 107, 4194-4199 (2010).
- Lee, S., et al. Distinct macrophage phenotypes contribute to kidney injury and repair. J Am Soc Nephrol. 22, 317-326 (2011).
- Kumar, S., et al. Dexamethasone ameliorates renal ischemia-reperfusion injury. J Am Soc Nephrol. 20, 2412-2425 (2009).
- Sharples, E. J., et al. Erythropoietin protects the kidney against the injury and dysfunction caused by ischemia-reperfusion. J Am Soc Nephrol. 15, 2115-2124 (2004).
- Gueler, F., et al. Statins attenuate ischemia-reperfusion injury by inducing heme oxygenase-1 in infiltrating macrophages. Am J Pathol. 170, 1192-1199 (2007).
- Delbridge, M. S., Shrestha, B. M., Raftery, A. T., ElNahas, A. M., Haylor, J. L. The effect of body temperature in a rat model of renal ischemia-reperfusion injury. Transplant Proc. 39, 2983-2985 (2007).
