在啮齿类动物的肠动力及胃肠壁的炎症分析功能评价一个标准化的模式肠道操作
Summary
术后肠梗阻（POI）是腹部外科手术的并发症导致的发病率和住院时间延长。由于缺乏预防或治疗策略加强研究是必要的。因此，我们建立了一个标准化的和可行的小鼠模型的病理生理POI调查和研究潜在的治疗方案。
Abstract
胃肠道的炎症，是一种常见的人类疾病的各种原因。在调查复杂的细胞和分子的肠道病理动物研究模型是至关重要的。虽然经常在许多炎症性疾病，最近的作品表现出兴趣的器官的粘膜，肌层（ME）是也是一个高度免疫功能的器官，港口密集的居民免疫网络。1,2这些作品内进行标准化肠道操纵模式（IM），导致炎症的肠壁，主要限于向ME。临床炎症导致长期的肠道运动功能障碍，术后肠梗阻（POI），这是一个频繁和不可避免的腹部手术后的并发症。炎症的特点是解放促炎症介质，如IL-6和IL-1β或抑制性神经递质像nitriÇ氧化氮（NO）。随后，免疫细胞的数量极大外渗到ME，中性粒细胞（PMN）和单核细胞为主的终于保持POI。2天为期，肠麻痹导致风险增加的愿望，细菌易位和感染并发症脓毒症和多器官功能衰竭，并导致较高的经济负担。
在这个手稿中，我们证明了IM标准化的模式，并在体内胃肠道过境评估（GIT）和结肠运输。此外，我们表现出的方法分离ME粘膜免疫分析，这是非常重要的区别，在这些具有高度的免疫活性肠壁车厢之间的炎症反应。所有的分析很容易转移到任何其他的研究模型，影响肠胃功能。
Protocol
1。动物男C57BL6 / J小鼠的平均重量为25g，得到从Harlan Winkelmann的（Borchen，德国）。所有的实验都按照联邦法律有关保护动物的。实验动物管理的原则得到遵守。动物均保持在12小时光/暗周期，并提供使用市售的啮齿动物食物和自来水自由采食 。动物应该被安置在动物实验前至少7天到达后，所有实验都可以很容易地适应老鼠或其他物种，到大动物模型稍作修改。 </…
Discussion
分析不同水平的IM（小肠膨出10 min比轻轻用的两个棉花喷头与IM的小肠进入盲肠内容的疏散的小肠进行检查）的比较研究显示手术操作和POI的程度之间的相关性。与其他组相比，IM显示出最高的白细胞数量在ME（PNMS，巨噬细胞和肥大细胞），导致长期和严重的POI 8。
在建立模型的IM，肠操作的规范运作和强度的关键步骤。触摸肠系膜在此过程中，必须严格避免，以防止?…
Divulgazioni
The authors have nothing to disclose.
Acknowledgements
这项工作是支持的补助金从德意志研究联合会（KA1270/3-1/2）“和BONFOR（O-112.0040）。
Materials
| Name of the reagent/Equipment | Company | Catalogue number |
| Ethilon 5/0 | Ethicon | 1666H |
| Cotton applicators | MaiMed | 71010 |
| arterial catheter | Vygon | 115-798 |
| 96 well plate (black) | Greiner Bio One | 655096 |
| glass ball (2 mm diameter) | Worf | available on request |
| Insect pins | Fine Science Tools | 26000-25 |
| Hanker Yates Reagent | Polyscience | 560694 |
| Aquatek | Merck | HC109850 |
| DMEM | Lonza | BE12-604 F |
| FITC-dextran (MW 70000) | Sigma Aldrich | 46945-500MG-F |
Riferimenti
- Wehner, S., Behrendt, F. F., Lyutenski, B. N., Lysson, M., Bauer, A. J., Hirner, A., Kalff, J. C. Inhibition of macrophage function prevents intestinal inflammation and postoperative ileus in rodents. Gut. 56, 176-185 (2007).
- Kalff, J. C., Schwarz, N. T., Walgenbach, K. J., Schraut, W. H., Bauer, A. J. Leukocytes of the intestinal muscularis: their phenotype and isolation. J. Leukoc. Biol. 63, 683-691 (1998).
- Kehlet, H. Postoperative ileus. Gut. 47, iv85-iv86 (2000).
- Wehner, S., Schwarz, N. T., Hundsdoerfer, R., Hierholzer, C., Tweardy, D. J., Billiar, T. R., Bauer, A. J., Kalff, J. C. Induction of IL-6 within the rodent intestinal muscularis after intestinal surgical stress. Surgery. 137, 436-446 (2005).
- Kalff, J. C., Schraut, W. H., Billiar, T. R., Simmons, R. L., Bauer, A. J. Role of inducible nitric oxide synthase in postoperative intestinal smooth muscle dysfunction in rodents. Gastroenterology. 118, 316-327 (2000).
- Iyer, S., Saunders, W. B., Stemkowski, S. Economic burden of postoperative ileus associated with colectomy in the United States. J. Manag. Care Pharm. 15, 485-494 (2009).
- Miller, M. S., Galligan, J. J., Burks, T. F. Accurate measurement of intestinal transit in the rat. J. Pharmacol. Methods. 6, 211-217 (1981).
- Kalff, J. C., Schraut, W. H., Simmons, R. L., Bauer, A. J. Surgical manipulation of the gut elicits an intestinal muscularis inflammatory response resulting in postsurgical ileus. Ann. Surg. 228, 652-663 (1998).
- Wehner, S., Straesser, S., Vilz, T. O., Pantelis, D., Sielecki, T., de lC, V., Hirner, A., Kalff, J. C. Inhibition of p38 mitogen-activated protein kinase pathway as prophylaxis of postoperative ileus in mice. Gastroenterology. 136, 619-629 (2009).
- Konigsrainer, I., Turck, M. H., Eisner, F., Meile, T., Hoffmann, J., Kuper, M., Zieker, D., Glatzle, J. The Gut is not only the Target but a Source of Inflammatory Mediators Inhibiting Gastrointestinal Motility During Sepsis. Cell Physiol. Biochem. 28, 753-760 (2011).
- Schwarz, N. T., Kalff, J. C., Turler, A., Speidel, N., Grandis, J. R., Billiar, T. R., Bauer, A. J. Selective jejunal manipulation causes postoperative pan-enteric inflammation and dysmotility. Gastroenterology. 26, 159-169 (2004).
- Engel, D. R., Koscielny, A., Wehner, S., Maurer, J., Schiwon, M., Franken, L., Schumak, B., Limmer, A., Sparwasser, T., Hirner, A. T helper type 1 memory cells disseminate postoperative ileus over the entire intestinal tract. Nat. Med. , (2010).
- Stoffels, B., Schmidt, J., Nakao, A., Nazir, A., Chanthaphavong, R. S., Bauer, A. J. Role of interleukin 10 in murine postoperative ileus. Gut. 58, 648-660 (2009).
Tags
Functional AssessmentIntestinal MotilityGut Wall InflammationRodentsStandardized ModelIntestinal ManipulationGastrointestinal TractHuman DiseasesAnimal Research ModelsCellular And MolecularIntestinal PathologyTunica MucosaMuscularis ExternaImmunocompetent OrganResident ImmunocytesInflammation Of The Bowel WallPostoperative IleusAbdominal SurgeryProinflammatory MediatorsIL-6IL-1βNitric Oxide (NO)Immunocytes ExtravasatePolymorphonuclear Neutrophils (PMN)MonocytesIntestinal ParalysisIncreased Risk Of AspirationBacterial TranslocationInfectious ComplicationsSepsisMulti Organ Failure
Citazione di questo articolo
Vilz, T. O., Overhaus, M., Stoffels, B., Websky, M. v., Kalff, J. C., Wehner, S. Functional Assessment of Intestinal Motility and Gut Wall Inflammation in Rodents: Analyses in a Standardized Model of Intestinal Manipulation. J. Vis. Exp. (67), e4086, doi:10.3791/4086 (2012).