目前的研究是由美国国家研究院​​以R01-HL092188卫生补助金，以R01-DK083385和R01HL098294，麻醉教育与研究基金会的支持（发耳）授予李鹏和1东风集团（德意志研究联合会）研究奖学金（GR2121/1- 1）和美国心脏协会的资助到AG。

大会发言。所有操作应执行下一个堂堂正正的解剖显微镜（Leica），并通过使用手术凝固剂。在实验和假手术组的小鼠应在年龄和体重相匹配，以确保结果的可比性。应该是稳定的体温，血压，麻醉和流体管理和监控整个实验过程中。 1。麻醉和手术<ol><li> C57BL / 6小鼠应该是10至16周的年龄范围内，他们应该在各自的群体性别相匹配的。 </li><li>麻醉小鼠戊巴比妥钠（70毫克/公斤体重IP），约维持麻醉。 10毫克/千克/小时戊巴比妥钠。过量可显着降低血压，从而可以改变的结果。 </li><li>广场上的温度控制加热表（RT，埃芬博格，德国慕尼黑）小鼠直肠温度计探头连接到热反馈控制器，以保持体温在37°C。 </li><li>地方小鼠在手术台上仰卧位，上肢和下肢表附有可拆卸的磁带。 </li><li>在生存实验的情况下，适当的无菌技术，包括裁剪头发，皮肤消毒，用碘伏（如优碘），蒸压仪器的无菌处理，并使用无菌手套口罩医生是必需的。 </li></ol> 2。门户三合会闭塞<ol><li>用矿物油覆盖切口区，以防止吸入鼠标头发，皮肤切割，从而防止鼠标毛过敏的风险。执行白线和中线剖腹切口上端，使您可以轻松地暴露肝脏。如果可能的话，执行，以防止出血与凝血电极（ERBE，ICC50，图宾根，德国）的切口。 </li><li>将加热表，用鼠标在一个位置，尾巴是方向泰德医生。这一立场提供本地化的黑社会，并放置缝合的最佳设置。 </li><li>取代用湿棉棒棉签揭露入肝右侧肠。 </li><li>稍微转向对膜片的中位数和左叶和右叶对肠道。这会给你一个明确的说法入肝右，左，中位数/叶分叉以上。 </li><li>仔细检查入肝举行平淡钳右叶，轻轻放下。这会给你一个很好的左侧入肝。 </li><li>一旦视觉识别（7/0尼龙线;爱惜康，Norderstedt的，德国）与缝合针持针器通过左入肝下的，包括肝动脉，肝静脉和胆管从左侧到右侧鼠标。 </li><li>十字绣是黑社会下密切合作。针可以伤害更深位于下腔巴卡劣质黑社会。因此保持与右侧肝叶钳查明前腔静脉放置针。 </li><li>以镊子取针右侧的一角，轻轻地不伤肝，肝区拉出。 </li><li>附加到每个缝合Eppendorf管（约3克，装满水），将两者缝合超过缝合持有结束。缝合持有人将远端的针，使缝合不正确的黑社会干扰。 </li><li>黑社会所采用的权重将立即闭塞，引起肝脏的左，中位数叶的血液供应中断。成功闭塞，可以很容易地证实缺血叶苍白热烫的视觉检查。相比之下，颜色变化立即消失，吊重时从两极释放和再灌注肝脏。这部分肝缺血模型，避免了维护布卢肠系膜充血正确的肝叶的三维流。 </li><li>取代肠道进入腹腔。保持肝脏和小肠温暖的湿棉签用清水浸泡在37.0°C。此外，以保持体温稳定的市售食品保鲜膜覆盖完成手术后的小鼠。 </li><li>执行缺血为实验方案的要求。定义的缺血时间释放后的重量和再灌注损伤的中位数和左肝叶将立即开始。这项技术使生产可逆性缺血小鼠。如果没有进一步的缺血时间将小心地取出缝合。 </li><li>进入前15分钟和再灌注后直接腹腔加入约400μL氯化钠。继续在100μL，以弥补在再灌注时间的流体损失的金额来代替液每隔30分钟。 </li><li>生存实验外科伤口闭合用连续缝合肌肉壁和皮肤的Wi第4/0缝线。 </li></ol> 3。肝脏灌注的变化建议确定以下肝损伤肝缺血以下结果参数: 确定肝酶（如血清ALT，AST）和肝组织病理（H＆E染色）以下的肝脏缺血再灌注3,9。 <img alt="图1" src="/files/ftp_upload/2550/2550fig1.jpg" /> 图1对肝脏缺血悬重系统型号:吊重系统已经建立，诱发肝缺血的左，中叶3。 （一）因此，尾和肝右叶，左叶轻轻分开。右叶再稍微转移到清楚地查看上面的右，左，中位数和叶分叉入肝。 8.0尼龙线被划归左肝黑社会。 （二）3克离心涂在小，重量轻（每个缝合结束装满水的BES）重视和应用。肝脏缺血再由左中叶从红色到浅的颜色的变化是可见的。这个实验设计，可以精确的闭塞和再灌注左肝黑社会的申请和释放负重。 <img alt="图2" src="/files/ftp_upload/2550/2550fig2.jpg" /> 图2。不同的肝缺血时间对丙氨酸（ALT）和天门冬氨酸（AST）在小鼠转氨酶。局部的入肝缺血诱导指示（0 - 60分钟缺血）。再灌注2小时后，谷丙转氨酶（A）和助攻（二）进行了测量。结果表示为平均值±SE 6 - 7小鼠/组。 *与0 min缺血（假手术对照组）相比，P &lt;0.05。

<ol>
	<li>Rook, M., Rand, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Predictors+of+long-term+outcome+after+liver+transplant.">Predictors of long-term outcome after liver transplant.</a> Curr. Opin. Organ. Transplant. 16, 499-504 (2011).</li><li>Henrion, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ischemia/reperfusion+injury+of+the+liver:+pathophysiologic+hypotheses+and+potential+relevance+to+human+hypoxic+hepatitis.">Ischemia/reperfusion injury of the liver: pathophysiologic hypotheses and potential relevance to human hypoxic hepatitis.</a> Acta. Gastroenterol. Belg. 63, 336-347 (2000).</li><li>Hart, M. L., Much, C., Kohler, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+a+hanging-weight+system+for+liver+ischemic+preconditioning+in+mice.">Use of a hanging-weight system for liver ischemic preconditioning in mice.</a> Am. J. Physiol. Gastrointest. Liver Physiol. 294, 1431-1440 (2008).</li><li>Schneider, M., Geyte, K., Fraisl, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Loss+or+silencing+of+the+PHD1+prolyl+hydroxylase+protects+livers+of+mice+against+ischemia/reperfusion+injury.">Loss or silencing of the PHD1 prolyl hydroxylase protects livers of mice against ischemia/reperfusion injury.</a> Gastroenterology. 138, 1143-1154 (2010).</li><li>Coriat, R., Leconte, M., Kavian, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mangafodipir+Protects+against+Hepatic+Ischemia-Reperfusion+Injury+in+Mice.">Mangafodipir Protects against Hepatic Ischemia-Reperfusion Injury in Mice.</a> PLoS One. 6, e27005 (2011).</li><li>Sakai, N., Van Sweringen, H. L., Schuster, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Receptor+activator+of+NF-kappaB+ligand+(RANKL)+protects+against+hepatic+ischemia/reperfusion+injury+in+mice.">Receptor activator of NF-kappaB ligand (RANKL) protects against hepatic ischemia/reperfusion injury in mice.</a> Hepatology. , (2011).</li><li>Zhang, J., Hu, W., Xing, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Protective+Role+of+CD59+and+Pathogenic+Role+of+Complement+in+Hepatic+Ischemia+and+Reperfusion+Injury.">The Protective Role of CD59 and Pathogenic Role of Complement in Hepatic Ischemia and Reperfusion Injury.</a> Am. J. Pathol. 179, 2876-2884 (2011).</li><li>Beldi, G., Banz, Y., Kroemer, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deletion+of+CD39+on+natural+killer+cells+attenuates+hepatic+ischemia/reperfusion+injury+in+mice.">Deletion of CD39 on natural killer cells attenuates hepatic ischemia/reperfusion injury in mice.</a> Hepatology. 51, 1702-1711 (2010).</li><li>Hart, M. L., Much, C., Gorzolla, I. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+adenosine+production+by+ecto-5'-nucleotidase+protects+during+murine+hepatic+ischemic+preconditioning.">Extracellular adenosine production by ecto-5'-nucleotidase protects during murine hepatic ischemic preconditioning.</a> Gastroenterology. 135, 1739-1750 (2008).</li><li>Beraza, N., Ludde, T., Assmus, U., Roskams, T., Vander Borght, S., Trautwein, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hepatocyte-specific+IKK+gamma/NEMO+expression+determines+the+degree+of+liver+injury.">Hepatocyte-specific IKK gamma/NEMO expression determines the degree of liver injury.</a> Gastroenterology. 132, 2504-2517 (2007).</li><li>Lentsch, A. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Activation+and+function+of+hepatocyte+NF-kappaB+in+postischemic+liver+injury.">Activation and function of hepatocyte NF-kappaB in postischemic liver injury.</a> Hepatology. 42, 216-218 (2005).</li><li>Luedde, T., Assmus, U., Wustefeld, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deletion+of+IKK2+in+hepatocytes+does+not+sensitize+these+cells+to+TNF-induced+apoptosis+but+protects+from+ischemia/reperfusion+injury.">Deletion of IKK2 in hepatocytes does not sensitize these cells to TNF-induced apoptosis but protects from ischemia/reperfusion injury.</a> J. Clin. Invest. 115, 849-859 (2005).</li><li>Kapitsinou, P. P., Liu, Q., Unger, T. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hepatic+HIF-2+regulates+erythropoietic+responses+to+hypoxia+in+renal+anemia.">Hepatic HIF-2 regulates erythropoietic responses to hypoxia in renal anemia.</a> Blood. , (2010).</li></ol>

没有利益冲突的声明。

本研究中描述的执行缺血小鼠模型中，由闭塞的左肝入肝使用吊重系统的技术。虽然这是一个简单的方法来学习，应考虑以下的陷阱。在开始时很常见的错误是一种深深的缝，其中包括入肝和腔静脉。在15分钟内，由于阻力增加和减少血液返回​​到心脏造成小鼠的早期死亡的呼吸和心脏的速度将放慢。最好的办法，以防止包括腔静脉定位针前腔静脉右叶用钳子。另一个重要问题是，权重不重，这可能会导致太多压力也停止血流到右叶的左边和右边的门户试验。因此，在离心瓶的水的量应调整​​鼠标的重量和Suture持有人应放在远端缝合位置。相反，如果权重低的血压没有完全停止。这两种错误是不易察觉的一个额外的中位数和左叶，右叶或颜色变化不足苍白的颜色变化，分别。因此，肝的颜色变化，运用权重后，应控制在每次10分钟。另一种常见的手术并发症是肝针受伤。因此，应按住右叶一个平淡的镊子，针尖，这样可以很容易看到，并与持针器。如果这些缺陷在开始考虑为肝缺血悬重系统提供了一个高度可重复性，减少变异肝损伤夹紧入肝缺血损伤。使用吊重系统，入肝仅操作一次，在整个手术过程中，造成肝瓣显着减少伤害。此外，无肝，肠充血，发生这种技术。发生更多的组织创伤血管钳，拆卸和更换，尤其是在多个缺血周期，如在缺血预处理钳补办。使用挂在肝组织的远程位置的权重，根据我们的观察，提供了可靠的闭塞的优势，同时防止因操纵钳补办肝叶组织创伤。两者合计，目前的研究提供了入肝闭塞缺血悬重系统的可行性，尽量减少与夹紧的变异和相关的限制。因此，这种技术可能会感兴趣的其他研究者考虑研究肝缺血。

<table border="1"><tbody><tr><td> 试剂名称 </td><td> 公司 </td><td> 目录编号 </td><td> 评论 </td></tr><tr><td>戊巴比妥钠（致命加） </td><td> LS，vortech制药有限公司</td><td>机VPL 9372 </td><td> 4毫克/毫升生理盐水中</td></tr><tr><td>缝合，普理灵8.0 </td><td>美国爱惜康， </td><td> M8739 </td><td>可重复使用</td></tr><tr><td>解剖显微镜</td><td>徕卡</td><td> N / A </td><td>考虑慷慨的工作距离</td></tr><tr><td>暖气表</td><td> RT，Effenberger德国</td><td> N / A </td><td>仅与单一供应商</td></tr><tr><td>手术器械</td><td> WPI的，杜蒙</td><td></td><td>钳，持针器</td></tr><tr><td>凝结剂</td><td> ERBE，ICC50，德国图宾根， </td><td> N / A </td><td>不同供应商的凝结剂</td></tr></tbody></table> 表1。特定的试剂和设备。

use of a hanging-weight system for liver ischemia in mice

因缺血的急性肝损伤过程中可能出现一些如肝移植，肝肿瘤切除或创伤修复的临床程序，并可能导致肝功能衰竭，具有较高的死亡率1-2。因此小鼠肝缺血的研究已成为一个重要的研究领域提供了机会，利用药理学和遗传学研究3-9。具体来说，有条件组织特定基因缺失小鼠（CRE，FLOX系统）提供了的蛋白质在特定组织的作用10-13见解。由于手动夹紧小鼠入肝，与相 ​​关的技术难度，我们进行了系统评价，使用悬重系统已先前所描述的3入肝闭塞。使用吊重系统，我们将不会造成任何损害肝瓣缝合周围的左支入肝，因为还最好的夹子可用可引起肝组织的损伤，由于肝组织的船只密切位置。此外，右支，肝黑社会仍然从而灌注血流量，肝右叶保留，没有肠道淤血发生这种技术。此外，入肝只操纵在整个手术过程一次。因此，程序，如空调前，缺血再灌注短的时间，可以很容易地进行。这个模型通过执行不同的缺血再灌注时间的系统评价发现与肝损伤的肝缺血时间密切相关，由丙氨酸（ALT）和天门冬氨酸（AST），血清转氨酶水平3,9。两者合计，这些研究证实，高重现肝损伤，肝缺血和间歇性灌注悬重系统时使用。因此，这种技术可能是有用的其他研究者的兴趣在小鼠肝脏缺血研究。因此，视频剪辑，这种技术提供了一个详细的一步一步的描述。

Watch this Scientific Journal Video about Use of a Hanging-weight System for Liver Ischemia in Mice at JoVE.com

Use of a Hanging-weight System for Liver Ischemia in Mice

我们建立了一个门户黑社会闭塞的吊重系统的一种新的小鼠模型。这项技术可能有助于为今后的调查，在小鼠肝模型缺血。

使用吊重系统在小鼠肝缺血

Acute liver injury due to ischemia can occur during several clinical procedures e.g. liver transplantation, hepatic tumor resection or trauma repair and ...

use-of-a-hanging-weight-system-for-liver-ischemia-in-mice

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

15.9K 次观看.  University of Colorado, Denver. 我们建立了一个门户黑社会闭塞的吊重系统的一种新的小鼠模型。这项技术可能有助于为今后的调查，在小鼠肝模型缺血。

视频: Use of a Hanging-weight System for Liver Ischemia in Mice

Use of a Hanging Weight System for Coronary Artery Occlusion in Mice

Murine studies of acute injury are an area of intense investigation, as knockout mice for different genes are becoming increasingly available 1-38. Cardioprotection by ischemic preconditioning (IP) remains an area of intense investigation. To further elucidate its molecular basis, the use of knockout mouse studies is particularly important 7, 14, 30, 39. Despite the fact that previous studies have already successfully performed cardiac ischemia and reperfusion in mice, this model is technically very challenging. Particularly, visual identification of the coronary artery, placement of the suture around the vessel and coronary occlusion by tying off the vessel with a supported knot is technically difficult. In addition, re-opening the knot for intermittent reperfusion of the coronary artery during IP without causing surgical trauma adds additional challenge. Moreover, if the knot is not tied down strong enough, inadvertent reperfusion due to imperfect occlusion of the coronary may affect the results. In fact, this can easily occur due to the movement of the beating heart.
Based on potential problems associated with using a knotted coronary occlusion system, we adopted a previously published model of chronic cardiomyopathy based on a hanging weight system for intermittent coronary artery occlusion during IP 39. In fact, coronary artery occlusion can thus be achieved without having to occlude the coronary by a knot. Moreover, reperfusion of the vessel can be easily achieved by supporting the hanging weights which are in a remote localization from cardiac tissues.
We tested this system systematically, including variation of ischemia and reperfusion times, preconditioning regiments, body temperature and genetic backgrounds39. In addition to infarct staining, we tested cardiac troponin I (cTnI)
as a marker of myocardial infarction in this model. In fact, plasma levels of cTnI correlated with infarct sizes (R2=0.8). Finally, we could show in several studies that this technique yields highly reproducible infarct sizes during murine IP and myocardial infarction6, 8, 30, 40, 41. Therefore, this technique may be helpful for researchers who pursue molecular mechanisms involved in cardioprotection by IP using a genetic approach in mice with targeted gene deletion. Further studies on cardiac IP using transgenic mice may consider this technique.

A murine model for myocardial ischemia and ischemic preconditioning is an important tool study cardioprotective mechanisms in vivo. Here, we report an easy applicable in situ model for cardiac IP using a hanging-weight system for coronary artery occlusion.

悬重系统的使用在小鼠冠状动脉闭塞

Murine studies of acute injury are an area of intense investigation, as knockout mice for different genes are becoming increasingly available 1-38. ...

科研

医学

Use of a Hanging-weight System for Isolated Renal Artery Occlusion

In hospitalized patients, over 50% of cases of acute kidney injury (AKI) are caused by renal ischemia 1-3. A recent study of hospitalized patients revealed that only a mild increase in serum creatinine levels (0.3 to 0.4 mg/dl) is associated with a 70% greater risk of death than in persons without any increase 1. Along these lines, surgical procedures requiring cross-clamping of the aorta and renal vessels are associated with a renal failure rates of up to 30% 4. Similarly, AKI after cardiac surgery occurs in over 10% of patients under normal circumstances and is associated with dramatic increases in mortality. AKI are also common complications after liver transplantation. At least 8-17% of patients end up requiring renal replacement therapy 5. Moreover, delayed graft function due to tubule cell injury during kidney transplantation is frequently related to ischemia-associated AKI 6. Moreover, AKI occurs in approximately 20% of patients suffering from sepsis 6.The occurrence of AKI is associated with dramatic increases of morbidity and mortality 1. Therapeutic approaches are very limited and the majority of interventional trials in AKI have failed in humans. Therefore, additional therapeutic modalities to prevent renal injury from ischemia are urgently needed 3, 7-9. 
To elucidate mechanisms of renal injury due to ischemia and possible therapeutic strategies murine models are intensively required 7-13. Mouse models provide the possibility of utilizing different genetic models including gene-targeted mice and tissue specific gene-targeted mice (cre-flox system). However, murine renal ischemia is technically challenging and experimental details significantly influence results. We performed a systematic evaluation of a novel model for isolated renal artery occlusion in mice, which specifically avoids the use of clamping or suturing the renal pedicle 14. This model requires a nephrectomy of the right kidney since ischemia can be only performed in one kidney due to the experimental setting. In fact, by using a hanging-weight system, the renal artery is only instrumented once throughout the surgical procedure. In addition, no venous or urethral obstruction occurs with this technique. We could demonstrate time-dose-dependent and highly reproducible renal injury with ischemia by measuring serum creatinine. Moreover, when comparing this new model with conventional clamping of the whole pedicle, renal protection by ischemic preconditioning is more profound and more reliable. Therefore his new technique might be useful for other researchers who are working in the field of acute kidney injury.

A precise murine model for acute kidney injury (AKI) due to ischemia is an important tool to investigate acute kidney injury and possibly find therapeutic tools to treat renal injury. The hanging weight system offers a tool for immediate and reliable renal artery occlusion and reperfusion without causing renal congestion.

一个悬重系统使用隔离肾动脉闭塞

In hospitalized patients, over 50% of cases of acute kidney injury (AKI) are caused by renal ischemia 1-3. A recent study of hospitalized patients ...

Use of Animal Model of Sepsis to Evaluate Novel Herbal Therapies

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several techniques, including infusion of exogenous bacterial toxin (endotoxemia) or bacteria (bacteremia), as well as surgical perforation of the cecum by cecal ligation and puncture (CLP)1-3. CLP allows bacteria spillage and fecal contamination of the peritoneal cavity, mimicking the human clinical disease of perforated appendicitis or diverticulitis. The severity of sepsis, as reflected by the eventual mortality rates, can be controlled surgically by varying the size of the needle used for cecal puncture2. In animals, CLP induces similar, biphasic hemodynamic cardiovascular, metabolic, and immunological responses as observed during the clinical course of human sepsis3. Thus, the CLP model is considered as one of the most clinically relevant models for experimental sepsis1-3.
Various animal models have been used to elucidate the intricate mechanisms underlying the pathogenesis of experimental sepsis. The lethal consequence of sepsis is attributable partly to an excessive accumulation of early cytokines (such as TNF, IL-1 and IFN-&gamma;)4-6 and late proinflammatory mediators (e.g., HMGB1)7. Compared with early proinflammatory cytokines, late-acting mediators have a wider therapeutic window for clinical applications. For instance, delayed administration of HMGB1-neutralizing antibodies beginning 24 hours after CLP, still rescued mice from lethality8,9, establishing HMGB1 as a late mediator of lethal sepsis. The discovery of HMGB1 as a late-acting mediator has initiated a new field of investigation for the development of sepsis therapies using Traditional Chinese Herbal Medicine. In this paper, we describe a procedure of CLP-induced sepsis, and its usage in screening herbal medicine for HMGB1-targeting therapies.

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection, and can be simulated by a surgical technique termed cecal ligation and puncture (CLP). Here we describe a method to use CLP-induced animal model to screen medicinal herbs for therapeutic agents.

脓毒症动物模型，用于评估新型草药疗法

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several ...

Intraoperative Detection of Subtle Endometriosis: A Novel Paradigm for Detection and Treatment of Pelvic Pain Associated with the Loss of Peritoneal Integrity

Endometriosis is a common disease affecting 40 to 70% of reproductive-aged women with chronic pelvic pain (CPP) and/or infertility. The purpose of this study was to demonstrate the use of a blue dye (methylene blue) to stain peritoneal surfaces during laparoscopy (L/S) to detect the loss of peritoneal integrity in patients with pelvic pain and suspected endometriosis. Forty women with CPP and 5 women without pain were evaluated in this pilot study. During L/S, concentrated dye was sprayed onto peritoneal surfaces, then aspirated and rinsed with Lactated Ringers solution. Areas of localized dye uptake were evaluated for the presence of visible endometriotic lesions. Areas of intense peritoneal staining were resected and some fixed in 2.5% buffered gluteraldehyde and examined by scanning (SEM) electron microscopy. Blue dye uptake was more common in women with endometriosis and chronic pelvic pain than controls (85% vs. 40%). Resection of the blue stained areas revealed endometriosis by SEM and loss of peritoneal cell-cell contact compared to normal, non-staining peritoneum. Affected peritoneum was associated with visible endometriotic implants in most but not all patients. Subjective pain relief was reported in 80% of subjects. Based on scanning electron microscopy, we conclude that endometrial cells extend well beyond visible implants of endometriosis and appear to disrupt the underlying mesothelium. Subtle lesions of endometriosis could therefore cause pelvic pain by disruption of peritoneal integrity, allowing menstrual or ovulatory blood and associated pain factors access to underlying sensory nerves. Complete resection of affected peritoneum may provide a better long-term treatment for endometriosis and CPP. This simple technique appears to improve detection of subtle or near invisible endometriosis in women with CPP and minimal visual findings at L/S and may serve to elevate diagnostic accuracy for endometriosis at laparoscopy.

Loss of peritoneal integrity provides a new paradigm to understand and treat chronic pelvic pain in women with mild forms of endometriosis and can be easily detected using intraoperative instillation of dye at the time of laparoscopy.

术中的微妙的子宫内膜异位症的检测:A新型范式为相关联的与的的腹膜的完整性之亏损盆腔疼痛的的的的检测和治疗的

Endometriosis is a  common disease affecting 40 to 70% of reproductive-aged women with chronic  pelvic pain (CPP) and/or infertility. The purpose of this ...

Carotid Artery Infusions for Pharmacokinetic and Pharmacodynamic Analysis of Taxanes in Mice

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study model. As the use of mouse models are often a vital step in preclinical drug discovery and drug development1-8, it is necessary to design a system to introduce drugs into mice in a uniform, reproducible manner. Ideally, the system should permit the collection of blood samples at regular intervals over a set time course. The ability to measure drug concentrations by mass-spectrometry, has allowed investigators to follow the changes in plasma drug levels over time in individual mice1, 9, 10. In this study, paclitaxel was introduced into transgenic mice as a continuous arterial infusion over three hours, while blood samples were simultaneously taken by retro-orbital bleeds at set time points. Carotid artery infusions are a potential alternative to jugular vein infusions, when factors such as mammary tumors or other obstructions make jugular infusions impractical. Using this technique, paclitaxel concentrations in plasma and tissue achieved similar levels as compared to jugular infusion. In this tutorial, we will demonstrate how to successfully catheterize the carotid artery by preparing an optimized catheter for the individual mouse model, then show how to insert and secure the catheter into the mouse carotid artery, thread the end of the catheter out through the back of the mouse&#8217;s neck, and hook the mouse to a pump to deliver a controlled rate of drug influx. Multiple low volume retro-orbital bleeds allow for analysis of plasma drug concentrations over time.

This method was developed with the goal of delivering a steady drug solution via the carotid artery, to assess the pharmacokinetics of novel drugs in mouse models.

颈动脉输液的紫杉烷类化合物在小鼠体内的药代动力学和药效学分析

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study ...

Photothrombosis-induced Focal Ischemia as a Model of Spinal Cord Injury in Mice

Spinal cord injury (SCI) is a devastating clinical condition causing permanent changes in sensorimotor and autonomic functions of the spinal cord (SC) below the site of injury. The secondary ischemia that develops following the initial mechanical insult is a serious complication of the SCI and severely impairs the function and viability of surviving neuronal and non-neuronal cells in the SC. In addition, ischemia is also responsible for the growth of lesion during chronic phase of injury and interferes with the cellular repair and healing processes. Thus there is a need to develop a spinal cord ischemia model for studying the mechanisms of ischemia-induced pathology. Focal ischemia induced by photothrombosis (PT) is a minimally invasive and very well established procedure used to investigate the pathology of ischemia-induced cell death in the brain. Here, we describe the use of PT to induce an ischemic lesion in the spinal cord of mice. Following retro-orbital sinus injection of Rose Bengal, the posterior spinal vein and other capillaries on the dorsal surface of SC were irradiated with a green light resulting in the formation of a thrombus and thus ischemia in the affected region. Results from histology and immunochemistry studies show that PT-induced ischemia caused spinal cord infarction, loss of neurons and reactive gliosis. Using this technique a highly reproducible and relatively easy model of SCI in mice can be achieved that would serve the purpose of scientific investigations into the mechanisms of ischemia induced cell death as well as the efficacy of neuroprotective drugs. This model will also allow exploration of the pathological changes that occur following SCI in live mice like axonal degeneration and regeneration, neuronal and astrocytic Ca2+ signaling using two-photon microscopy.

Photothrombosis is a minimally invasive and highly reproducible procedure to induce focal ischemia in the spinal cord and serves as a model of spinal cord injury in mice.

Photothrombosis引起的局部脑缺血脊髓损伤的小鼠模型

Spinal cord injury (SCI) is a devastating clinical condition causing permanent changes in sensorimotor and autonomic functions of the spinal cord (SC) ...

Use of a Low-flow Digital Anesthesia System for Mice and Rats

A traditional vaporizer depends on flowing gas and atmospheric pressure for passive anesthetic vaporization. Newly developed direct injection vaporizers utilize a syringe pump to directly administer volatile anesthetics into a gas stream. Unlike a traditional vaporizer, it can be used at very low flow rates, making it ideal for use on mice and rats. 
The equipment's capability to use low flow rates could result in a substantial cost savings due to the reduced need for anesthetic agents, compressed gas, and charcoal scavenging filters1. A lower flow rate means less waste of anesthetic gas and likely reduces the risk of anesthetic exposure to laboratory personnel. Thus, the high levels of precision and safety associated with direct injection vaporizers, along with a reduced need for anesthetic agents, compressed gas, and charcoal filters are beneficial for research requiring small animal anesthesia. 
The goal of this protocol is to demonstrate the use of a syringe-driven direct injection vaporizer as part of a digital, low-flow anesthesia system. The direct injection vaporizer is capable of accurately delivering anesthesia at very low flow rates compared to a traditional vaporizer, making it a promising alternative for controlled gas anesthetic delivery to rodents.

Here, we present a protocol to more safely and efficiently administer anesthetic gas to mice using a digital, low flow anesthesia system utilizing a syringe-driven direct injection vaporizer.

低流量数字麻醉系统的小鼠和大鼠的用途

A traditional vaporizer depends on flowing gas and atmospheric pressure for passive anesthetic vaporization. Newly developed direct injection vaporizers ...

Confirmation of Myocardial Ischemia and Reperfusion Injury in Mice Using Surface Pad Electrocardiography

Many animal models have been established for the study of myocardial remodeling and heart failure due to its status as the number one cause of mortality worldwide. In humans, a pathologic occlusion forms in a coronary artery and reperfusion of that occluded artery is considered essential to maintain viability of the myocardium at risk. Although essential for myocardial recovery, reperfusion of the ischemic myocardium creates its own tissue injury. The physiologic response and healing of an ischemia/reperfusion injury is different from a chronic occlusion injury. Myocardial ischemia/reperfusion injury is gaining recognition as a clinically relevant model for myocardial infarction studies. For this reason, parallel animal models of ischemia/reperfusion are vital in advancing the knowledge base regarding myocardial injury. Typically, ischemia of the mouse heart after left anterior descending (LAD) coronary artery occlusion is confirmed by visible pallor of the myocardium below the occlusion (ligature). However, this offers only a subjective way of confirming correct or consistent ligature placement, as there are multiple major arteries that could cause pallor in different myocardial regions. A method of recording electrocardiographic changes to assess correct ligature placement and resultant ischemia as well as reperfusion, to supplement observed myocardial pallor, would help yield consistent infarct sizes in mouse models. In turn, this would help decrease the number of mice used. Additionally, electrocardiographic changes can continue to be recorded non-invasively in a time-dependent fashion after the surgery. This article will demonstrate a method of electrocardiographically confirming myocardial ischemia and reperfusion in real time.

During murine myocardial ischemia/reperfusion surgery, correct placement of the occluding ligature is typically confirmed by visible observation of myocardial pallor. Herein, a method of electrocardiographically confirming ischemia and reperfusion, to supplement observed myocardial pallor, is demonstrated in male C57Bl/6 mice.

小鼠使用表面垫心电图心肌缺血再灌注损伤的确认

Many animal models have been established for the study of myocardial remodeling and heart failure due to its status as the number one cause of mortality ...

Ultrasound-guided Intracardiac Injection of Human Mesenchymal Stem Cells to Increase Homing to the Intestine for Use in Murine Models of Experimental Inflammatory Bowel Diseases

Crohn&#39;s disease (CD) is a common chronic inflammatory disease of the small and large intestines. Murine and human mesenchymal stem cells (MSCs) have immunosuppressive potential and have been shown to suppress inflammation in mouse models of intestinal inflammation, even though the route of administration can limit their homing and effectiveness 1,3,4,5. Local application of MSCs to colonic injury models has shown greater efficacy at ameliorating inflammation in the colon. However, there is paucity of data on techniques to enhance the localization of human bone marrow-derived MSCs (hMSCs) to the small intestine, the site of inflammation in the SAMP-1/YitFc (SAMP) model of experimental Crohn&#39;s disease. This work describes a novel technique for the ultrasound-guided intracardiac injection of hMSCs in SAMP mice, a well-characterized spontaneous model of chronic intestinal inflammation. Sex- and age-matched, inflammation-free AKR/J (AKR) mice were used as controls. To analyze the biodistribution and the localization, hMSCs were transduced with a lentivirus containing a triple reporter. The triple reporter consisted of firefly luciferase (fl), for bioluminescent imaging; monomeric red fluorescent protein (mrfp), for cell sorting; and truncated herpes simplex virus thymidine kinase (ttk), for positron emission tomography (PET) imaging. The results of this study show that 24 h after the intracardiac administration, hMSCs localize in the small intestine of SAMP mice as opposed to inflammation-free AKR mice. This novel, ultrasound-guided injection of hMSCs in the left ventricle of SAMP mice ensures a high success rate of cell delivery, allowing for the rapid recovery of mice with minimal morbidity and mortality. This technique could be a useful method for the enhanced localization of MSCs in other models of small-intestinal inflammation, such as TNF&#916;RE6. Future studies will determine if the increased localization of hMSCs by intra-arterial delivery can lead to increased therapeutic efficacy.

Murine studies in models of colonic inflammation have demonstrated that a small percentage (1 - 5%) of mesenchymal stem cells (MSC) injected intravenously or intraperitoneally home to the inflamed colon1,2. This study shows that ultrasound-guided intracardiac injections of MSCs result in increased localization to the intestine.

超声引导下人骨髓间充质干细胞在实验性肠炎小鼠模型中的应用

Crohn&#39;s disease (CD) is a common chronic inflammatory disease of the small and large intestines. Murine and human mesenchymal stem cells (MSCs) have ...

Murine Precision-Cut Liver Slices as an Ex Vivo Model of Liver Biology

Understanding the mechanisms of liver injury, hepatic fibrosis, and cirrhosis that underlie chronic liver diseases (i.e., viral hepatitis, non-alcoholic fatty liver disease, metabolic liver disease, and liver cancer) requires experimental manipulation of animal models and in vitro cell cultures. Both techniques have limitations, such as the requirement of large numbers of animals for in vivo manipulation. However, in vitro cell cultures do not reproduce the structure and function of the multicellular hepatic environment. The use of precision-cut liver slices is a technique in which uniform slices of viable mouse liver are maintained in laboratory tissue culture for experimental manipulation. This technique occupies an experimental niche that exists between animal studies and in vitro cell culture methods. The presented protocol describes a straightforward and reliable method to isolate and culture precision-cut liver slices from mice. As an application of this technique, ex vivo liver slices are treated with bile acids to simulate cholestatic liver injury and ultimately assess the mechanisms of hepatic fibrogenesis.

This protocol provides a simple and reliable method for the production of viable precision-cut liver slices from mice. The ex vivo tissue samples can be maintained under laboratory tissue culture conditions for multiple days, providing a flexible model to examine liver pathobiology.

毛利精密切肝片作为肝脏生物学的Ex Vivo模型

Understanding the mechanisms of liver injury, hepatic fibrosis, and cirrhosis that underlie chronic liver diseases (i.e., viral hepatitis, non-alcoholic ...