Name: intraductal delivery to the rabbit mammary gland
Uploaded: 2017-03-09T13:00:00.000+00:00
Duration: 6 min 30 s
Description: Watch this Scientific Journal Video about Intraductal Delivery to the Rabbit Mammary Gland at JoVE.com

The authors acknowledge support from a Translational Breast Cancer Research Grant (14-60-26-BROC to AB) from the Breast Cancer Research Foundation and the American Association for Cancer Research.

使用动物对象的程序已通过得克萨斯大学奥斯汀分校的机构动物护理和使用委员会。 1.术前准备<ol><li>记录体重每只兔。如同所有的临床前研究，定期监测动物的重量来评估潜在的毒性。 </li><li>前麻醉兔子，填充的50mL锥形管与可商购的超声凝胶并在500 xg离心旋转30秒;应在离心结束后的凝胶无可见的气泡。 </li><li>在剂量为0.75毫克/公斤给药格隆皮下注射，剂量为0.1毫克/千克和乙酰丙嗪肌注。等待15-20分钟的镇静，同时监测生命体征和行为才能生效。 注:格隆溴铵是防止心动过缓和减少呼吸道和胃肠道分泌物的抗胆碱能剂。乙酰丙嗪是作为一个premedicati镇静剂对麻醉。 </li><li>施用35毫克/千克氯胺酮和5mg / kg的甲苯噻嗪的皮下作为麻醉剂。但是，随着操作者变得更有经验，并且可以更迅速地完成导管内输送，降低药物剂量以15mg / kg的氯胺酮和3mg / kg的甲苯噻嗪皮下;这会缩短麻醉时间，并为动物从麻醉中恢复所需的时间。 </li><li>检查并记录心脏率，SPO 2，温度，呼吸频率和粘液膜的颜色，每15分钟。敷眼润滑剂双眼。 注:只有谁曾接受适当的培训，并已通过其机构的IACUC应该管理或监督人员的麻醉。使用兽医麻醉监视器可以在获取生命体征和推荐帮助。兽医的麻醉监测，但是，并不能取代需要手动检查动物每隔15分钟。请参阅制造商的说明˚F或正确使用兽医显示器。 </li><li>验证麻醉用温柔的脚趾捏发作;兔子应该在继续之前无响应。 </li><li>仔细剃兔子的尾腹部周围的第三和第四对腹股沟奶嘴的面积。 </li><li>与大多数除去毛发，施加过度的计数器脱毛膏到剃区域。删除使用用温水浸湿水的湿纸巾申请后，奶油10分钟。 注:兔子的皮肤是非常脆弱性/敏感性。脱毛膏应使用不超过10分钟。实际上，这是更安全的在5分钟以尝试"测试点"，如果测试点表示需要一段较长的时间仅在再离开。 </li><li>用酒精擦拭浸泡的纱布垫清洁注射部位的区域。 </li><li>放置在其背部的兔子在具有循环温水毯和吸收垫衬一个V形槽。 </li></ol> 2.镨造影剂的eparation <ol><li>根据制造商的说明重组的非靶向对照试剂。移液器上下轻轻混合。 注意:在这个协议中所使用的造影剂是在室温下搅拌4稳定 - 重构后6小时。轻轻摇动每个检索之间的小瓶。 注意:被注入的，将取决于管道的数目所需溶液的体积。兔子具有每奶头4导管开口，和0.2毫升的溶液是足以填满一个成年新西兰白兔（Oryctolaguscuniculus）之一导管树。因此，为0.8毫升总体积可以被传递给一个乳腺。 </li></ol> 3.乳腺导管内交货<ol><li>找到合适的奶嘴被注入;推荐腹股沟奶嘴的3 次和第 4 次对，如当动物被定位在其背部它们容易可视化。 </li><li>无菌的0.9％生理盐水的负载0.2毫升1毫升鲁尔锁结核菌素注射器用22克针。妥善处置了22克针的一次生理盐水是在注射器，并用无菌25号针头更换。轻轻地用一个纱布垫85％异丙醇的区域。 </li><li>用针上并平行于动物的身体的注射器的斜角，插入针的斜角到乳头的侧面和缓缓注入0.1-0.2毫升盐水;这将允许对导管开口的更好的可视化。 </li><li>负载0.2毫升注射溶液到1毫升鲁尔锁结核菌素注射器。 </li><li>用拇指和食指轻轻按住乳头，将其抬起稍微将它定位为导管内注射;可穿戴眼放大镜可在可视化导管开口帮助。 </li><li>在保持乳头的位置抬起，仔细导管插入的使用兴趣将25g钝尖的针管道。 </li><li>插管后，轻轻一拧就不客气-T的集线器鲁尔锁注射器IP输液针，直到它被锁定就位。 </li><li>提起乳头并注入溶液慢慢地减少因快速移动的管道内流体的潜在损害;在任何时候注入解决方案时，应该有阻力。 </li></ol> 4.超声成像<ol><li>应用离心超声凝胶的一个自由量的感兴趣的区域的皮肤上。确保有在凝胶中无气泡，因为这些将损害图像质量。 </li><li>成像深度设置为6毫米。放置在与凝胶接触的21-MHz的换能器和扫描的B模式感兴趣的区域。观察包括导管开放和整个管道的扫描区域的造影剂。 注意:这些设置已针对与特定光声超声机使用而开发的。请参阅材料表了解更多详情。可能需要调整的发射功率和其它成像系统的成像深度以优化乳腺可视化。 </li><li>从动物的皮肤用纱布垫除去超声波凝胶。 </li></ol> 5.术后护理<ol><li>观察注射部位:应该没有创伤乳头区域或周围组织的迹象，并在包围乳头区域肿胀可能表示一个乳房脂肪垫注射，而不是一个成功的导管内注射。 </li><li>将兔子在胸骨位置。如果需要的话，得到0.2毫克/公斤的育亨宾静脉内边缘耳静脉;这将扭转赛拉嗪的影响，并允许动物从麻醉中恢复得更快。 </li><li>监控兔子整个恢复期每15分钟，直到动物警觉，反应灵敏，并保持胸骨位置。 注:此过程不应导致组织损伤或肿胀。如果观察到发红或肿胀，管理一个剂量的美洛昔康0.1-0.2毫克/公斤PO一旦动物警觉，并能服用药物口服。为进一步指导联系该机构的兽医人员。 </li></ol>

<ol>
	<li>Flanagan, M., Love, S., Hwang, E. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Status+of+Intraductal+Therapy+for+Ductal+Carcinoma+in+Situ.">Status of Intraductal Therapy for Ductal Carcinoma in Situ.</a> Curr Breast Cancer Rep. 2 (2), 75-82 (2010).</li><li>Love, S. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Feasibility+Study+of+the+Intraductal+Administration+of+Chemotherapy.">A Feasibility Study of the Intraductal Administration of Chemotherapy.</a> Cancer Prevention Research. 6 (1), 51-58 (2013).</li><li>Stearns, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preclinical+and+Clinical+Evaluation+of+Intraductally+Administered+Agents+in+Early+Breast+Cancer.">Preclinical and Clinical Evaluation of Intraductally Administered Agents in Early Breast Cancer.</a> Sci Transl Med. 3 (106), 106ra108 (2011).</li><li>Zhang, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Safety+Parameters+of+the+Study+on+Intraductal+Cytotoxic+Agent+Delivery+to+the+Breast+before+Mastectomy.">The Safety Parameters of the Study on Intraductal Cytotoxic Agent Delivery to the Breast before Mastectomy.</a> Chin J Cancer Res. 26 (5), 579-587 (2014).</li><li>Mahoney, M. E., Gordon, E. J., Rao, J. Y., Jin, Y., Hylton, N., Love, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intraductal+Therapy+of+Ductal+Carcinoma+In+Situ:+a+Presurgery+Study.">Intraductal Therapy of Ductal Carcinoma In Situ: a Presurgery Study.</a> Clin Breast Cancer. 13 (4), 280-286 (2013).</li><li>Murata, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ductal+Access+for+Prevention+and+Therapy+of+Mammary+Tumors.">Ductal Access for Prevention and Therapy of Mammary Tumors.</a> Cancer Res. 66 (2), 638-645 (2006).</li><li>Chun, Y. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intraductally+Administered+Pegylated+Liposomal+Doxorubicin+Reduces+Mammary+Stem+Cell+Function+In+the+Mammary+Gland+but+in+the+Long+Term,+Induces+Malignant+Tumors.">Intraductally Administered Pegylated Liposomal Doxorubicin Reduces Mammary Stem Cell Function In the Mammary Gland but in the Long Term, Induces Malignant Tumors.</a> Breast Cancer Res Treat. 135 (1), 201-208 (2012).</li><li>Brock, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Silencing+HoxA1+by+Intraductal+Injection+of+siRNA+Lipidoid+Nanoparticles+Prevents+Mammary+Tumor+Progression+in+Mice.">Silencing HoxA1 by Intraductal Injection of siRNA Lipidoid Nanoparticles Prevents Mammary Tumor Progression in Mice.</a> Sci Trans Med. 6 (217), 2172a2 (2014).</li><li>Krause, S., Brock, A., Ingber, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intraductal+Injection+For+Localized+Drug+Delivery+To+The+Mouse+Mammary+Epithelium.">Intraductal Injection For Localized Drug Delivery To The Mouse Mammary Epithelium.</a> J Vis Exp. (80), e50692 (2013).</li><li>Brock, A., Goh, H. T., Yang, B., Yu, L., Li, H., Loh, Y. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cellular+Reprogramming:+A+New+Technology+Frontier+In+Pharmaceutical+Research.">Cellular Reprogramming: A New Technology Frontier In Pharmaceutical Research.</a> Pharm Res. 29 (1), 35-52 (2012).</li><li>Silverstein, M. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ductal+Carcinoma+In+Situ+of+the+Breast.">Ductal Carcinoma In Situ of the Breast.</a> Annu. Rev Med. 51, 17-32 (2000).</li><li>Love, S. M., Barsky, S. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anatomy+of+the+Nipple+And+Breast+Ducts+Revisited.">Anatomy of the Nipple And Breast Ducts Revisited.</a> Cancer. 101 (9), 1947-1957 (2004).</li><li>Virnig, B. A., Shamliyan, T., Tuttle, T. M., Kane, R. L., Wilt, W. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diagnosis+and+Management+of+Ductal+Carcinoma+In+Situ+(DCIS).">Diagnosis and Management of Ductal Carcinoma In Situ (DCIS).</a> Evidence Report/Technology Assessment. , 185 (2009).</li><li>Mills, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Examination+of+Duct+Physiology+in+the+Human+Mammary+Gland.">Examination of Duct Physiology in the Human Mammary Gland.</a> PLoS One. 11 (4), e0150653 (2016).</li><li>King, B. L., Love, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Intraductal+Approach+to+the+Breast:+Raison+d'Etre.">The Intraductal Approach to the Breast: Raison d'Etre.</a> Breast Cancer Res. 8 (2), 206 (2006).</li><li>Bu, W., Xin, L., Toneff, M., Li, L., Li, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lentivirus+vectors+for+stably+introducing+genes+into+mammary+epithelial+cells+in+vivo+.">Lentivirus vectors for stably introducing genes into mammary epithelial cells in vivo .</a> J Mammary Gland Biol Neoplasia. 14, 401-404 (2009).</li><li>Lyons, W. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Direct+Mammotrophic+Action+of+Lactogenic+Hormone.">The Direct Mammotrophic Action of Lactogenic Hormone.</a> Proc. Soc. Exp. Bio. Med. 51 (2), 308-311 (1942).</li><li>Fiddler, T. J., Birkinshaw, M., Falconer, I. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+Of+Intraductal+Prolactin+On+Some+Aspects+of+the+Ultrastructure+and+Biochemistry+of+Mammary+Tissue+in+the+Pseudopregnant+Rabbit.">Effects Of Intraductal Prolactin On Some Aspects of the Ultrastructure and Biochemistry of Mammary Tissue in the Pseudopregnant Rabbit.</a> J Endocrinol. 49 (3), 459-469 (1971).</li><li>Falconer, I. R., Fiddler, T. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+Intraductal+Administration+Of+Prolactin,+Actinomycin+D+and+Cycloheximide+on+Lipoprotein+Lipase+Activity+in+the+Mammary+Glands+of+Pseudopregnant+Rabbits.">Effects of Intraductal Administration Of Prolactin, Actinomycin D and Cycloheximide on Lipoprotein Lipase Activity in the Mammary Glands of Pseudopregnant Rabbits.</a> Biochim Biophys Acta. 218 (3), 508-514 (1970).</li><li>Singh, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+Molecular+Size+on+the+Retention+of+Polymeric+Nanocarrier+Diagnostic+Agents+in+Breast+Ducts.">Influence of Molecular Size on the Retention of Polymeric Nanocarrier Diagnostic Agents in Breast Ducts.</a> Pharm Res. 29 (9), 2377-2388 (2012).</li><li>Jain, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Atypical+Ductal+Hyperplasia:+Interobserver+and+Intraobserver+Variability.">Atypical Ductal Hyperplasia: Interobserver and Intraobserver Variability.</a> Mod Pathol. 24 (7), 917-923 (2011).</li><li>Betsill, W. L., Rosen, P. P., Lieberman, P. H., Robbins, G. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intraductal+Carcinoma.+Long-term+Follow-up+After+Treatment+by+Biopsy+Alone.">Intraductal Carcinoma. Long-term Follow-up After Treatment by Biopsy Alone.</a> JAMA. 239 (18), 1863-1867 (1978).</li><li>Eusebi, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+Follow-up+of+In+Situ+Carcinoma+of+the+Breast.">Long-term Follow-up of In Situ Carcinoma of the Breast.</a> Semin Diagn Pathol. 11 (3), 223-235 (1994).</li><li>Sanders, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Natural+History+Of+Low-Grade+Ductal+Carcinoma+In+Situ+Of+The+Breast+In+Women+Treated+By+Biopsy+Only+Revealed+Over+30+Years+Of+Long-Term+Follow-Up.">The Natural History Of Low-Grade Ductal Carcinoma In Situ Of The Breast In Women Treated By Biopsy Only Revealed Over 30 Years Of Long-Term Follow-Up.</a> Cancer. 103 (12), 2481-2484 (2005).</li><li>Esserman, L. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Addressing+Overdiagnosis+and+Overtreatment+in+Cancer:+A+Prescription+For+Change.">Addressing Overdiagnosis and Overtreatment in Cancer: A Prescription For Change.</a> Lancet Oncol. 15 (6), e234-e242 (2014).</li></ol>

The authors have nothing to disclose.

可用于超声对比试剂和许多其他的水溶液，包括重要的染料和治疗剂导管内输送到兔的乳腺的此方法。以前的研究已经表明激素17，18，19的导管内交付。在啮齿动物模型中，核酸8的导管内输送，化学治疗6,7和纳米颗粒的载体8，20<...

<html>治疗剂的导管内输送已经研究了在啮齿动物模型中和在早期的人体试验3，4，5，6，11，12。最近的I期临床试验证明导管内卡铂或女性乳房等待为浸润癌2治疗导管内聚乙二醇化脂质体阿霉素的安全性和可行性。 对导管内输送以前的协议已被开发用于小鼠和大鼠乳腺6，7，8，9。出于研究目的，导管内的肿瘤细胞注射和癌基因的慢病毒载体递送也已在啮齿动物模型中进行的裁判"&gt; 13，14，15，16，但是， 在导管内输送过程的体内模型的理想应允许新的类治疗化合物的发展和促进临床前评估。啮齿动物和人类之间的解剖学差异有复杂的这些翻译学习。 不像小鼠，其中每个管道在一个单独的乳头结束时，人乳腺癌由5至9个独立的导管系统中，每个与奶嘴结束一个单独的开口。兔乳腺怀有四个独立导管系统中，每个分别通过在一个单一的奶头4孔可接触。兔模型更接近人体解剖，并允许导管内给药的研究更相关的情境。 在这里，我们用两种方法来评估导管内交付。的一个共同给予活体染料允许可视化通过皮肤，并提供该方法的一个简单的和快速的确认。对于一些应用，导管的更高分辨率映射可以是优选的。我们在座的管道超声成像协议通过非靶向造影剂的导管内交付。

<table><tbody><tr><td>MicroMarker non-targeted contrast reagent</td><td>VisualSonics</td><td>VS-11694</td><td></td></tr><tr><td>Luer Lock 1 mL Syringes</td><td>BD</td><td>309628</td><td></td></tr><tr><td>Glycopyrrolate 0.2 mg/mL</td><td>Wedgewood Compounding Pharmacy</td><td>GLYCOP-INJ013VC</td><td>6 month shelf life, supply may be limited.&amp;nbsp;</td></tr><tr><td>Atropine Sulfate 0.5 mg/mL</td><td>Animal Health International</td><td>15320764</td><td>If glycopyrrolate is unavailable. Not to be combined with glycopyrrolate.</td></tr><tr><td>Ketamine HCL 100 mg/mL</td><td>Animal Health International</td><td>21250699</td><td>&lt;a href=&quot;http://www.animalhealthinternational.com/&quot;&gt;http://www.animalhealthinternational.com/&lt;/a&gt;</td></tr><tr><td>Acepromazine 10 mg/mL</td><td>Animal Health International</td><td>17640541</td><td></td></tr><tr><td>Xylazine 20 mg/mL</td><td>Animal Health International</td><td>20101547</td><td></td></tr><tr><td>Yohimbine 0.2 mg/mL</td><td>Animal Health International</td><td>14588965</td><td></td></tr><tr><td>Hair Removing Cream</td><td>Veet</td><td></td><td>Sensitive skin solution. Available through local retailers.</td></tr><tr><td>Blunt tip infusion needles</td><td>Sai Infusion Technology</td><td>B14-50</td><td>&lt;a href=&quot;http://www.sai-infusion.com/collections/blunt-needles&quot;&gt;http://www.sai-infusion.com/collections/blunt-needles&lt;/a&gt;</td></tr><tr><td>Veterinary Pulse Oximeter</td><td>EdanUSA</td><td>VE-H100B</td><td>&lt;a href=&quot;http://www.edanusa.com/Product/VE-H100B-Veterinary-Pulse-Oximeter.html&quot;&gt;http://www.edanusa.com/Product/VE-H100B-Veterinary-Pulse-Oximeter.html&lt;/a&gt;</td></tr><tr><td>Warm Water Pump</td><td>Gaymar</td><td>TP700</td><td></td></tr><tr><td>Warm Water Blanket</td><td>Animal Health International</td><td>21232696</td><td>Maxi-Therm Lite Warming Pads</td></tr><tr><td>Ultrasound system</td><td>VisualSonics</td><td>Vevo 2100</td><td></td></tr></tbody></table>

intraductal delivery to the rabbit mammary gland

对于乳腺癌的局部导管的治疗提供了潜在的优势，包括有效递送至肿瘤，并降低全身毒性和副作用1，2，3，4，5，6，7。然而，一些挑战依然存在之前这些处理可以更广泛地应用。在合适的动物模型的发展和导管治疗的有效性促进导管内治疗策略对患者的发展。而小鼠乳腺已被广泛地用作乳腺发育和肿瘤的模型系统中，解剖来自人腺不同。较大的动物模型，如兔，可作为乳腺结构和管内治疗开发一个更好的模型。在Contrast小鼠，在十个由导管树木在空间沿体轴分布，在一个单独的乳头每个终止，兔子乳腺更类似于人类腺，具有多个重叠导管系统该出口通过在一个乳头分开的开口。这里，我们提出了试剂的递送微创方法直接进入兔乳腺导管和用于递送本身具有高分辨率超声波成像的可视化。

这里，我们表明，对比试剂兔的乳腺导管的导管内输送而不创伤可以实现对组织（ 图2）。家兔，四个单独的导管系统会聚在同一乳头，并且因此可以被访问和使用这种方法单独地成像。个别导管开口容易显现;注意箭头标记相邻于图2B中的空心管的第二导管开口。 与线性成像模式不相关的对比试剂的?...

Watch this Scientific Journal Video about Intraductal Delivery to the Rabbit Mammary Gland at JoVE.com

Intraductal Delivery to the Rabbit Mammary Gland

Here, we describe a technique for the localized delivery of reagents to the rabbit mammary gland via an intraductal injection. In addition, we describe a protocol for visualization and the confirmation of delivery by high-resolution ultrasound imaging of contrast agents.

导管内送货到兔乳腺

Localized intraductal treatments for breast cancer offer potential advantages, including efficient delivery to the tumor and reduced systemic toxicity and ...

intraductal-delivery-to-the-rabbit-mammary-gland

Research

JoVE Journal

Bioengineering

9.3K 次观看.  The University of Texas at Austin. Here, we describe a technique for the localized delivery of reagents to the rabbit mammary gland via an intraductal injection. In addition, we describe a protocol for visualization and the confirmation of delivery by high-resolution ultrasound imaging of contrast agents. 

视频: Intraductal Delivery to the Rabbit Mammary Gland

Minimally Invasive Embryo Transfer and Embryo Vitrification at the Optimal Embryo Stage in Rabbit Model

Assisted reproductive techniques (ARTs), such as in vitro embryo culture or embryo cryopreservation, affect natural development patterns with perinatal and postnatal consequences. To ensure the innocuousness of ART applications, studies in animal models are necessary. In addition, as a last step, embryo development studies require evaluation of their capacity to develop full-term healthy offspring. Here, embryo transfer to the uterus is indispensable to perform any ARTs-related experiment.
The rabbit has been used as a model organism to study mammalian reproduction for over a century. In addition to its phylogenetic proximity to the human species and its small size and low maintenance cost, it has important reproductive characteristics such as induced ovulation, a chronology of early embryonic development similar to humans and a short gestation that allow us to study the consequences of ART application easily. Moreover, ARTs (such as intracytoplasmic sperm injection, embryo culture, or cryopreservation) are applied with suitable efficiency in this species.
Using the laparoscopic embryo transfer technique and the cryopreservation protocol presented in this article, we describe 1) how to transfer embryos through an easy, minimally invasive technique and 2) an effective protocol for long-term storage of rabbit embryos to provide time-flexible logistical capacities and the ability to transport the sample. The outcomes obtained after transferring rabbit embryos at different developmental stages indicate that morula is the ideal stage for rabbit embryo recovery and transfer. Thus, an oviductal embryo transfer is required, justifying the surgical procedure. Furthermore, rabbit morulae are successfully vitrified and laparoscopically transferred, proving the effectiveness of the described techniques.

Assisted reproductive techniques (ARTs) are in continuous evaluation to improve outcomes and reduce the associated risks. This manuscript describes a minimally invasive embryo transfer procedure with an efficient cryopreservation protocol that allows the use of rabbits as an ideal animal model of human reproduction.

家兔模型最佳胚胎阶段微创胚胎移植与胚胎玻璃化

Assisted reproductive techniques (ARTs), such as in vitro embryo culture or embryo cryopreservation, affect natural development patterns with perinatal ...

科研

发育生物学

Intraductal Delivery and X-ray Visualization of Ethanol-Based Ablative Solution for Prevention and Local Treatment of Breast Cancer in Mouse Models

Breast cancer is the most prevalent cancer and the second-leading cause of cancer-related death for women in the USA. For high-risk women, prophylactic mastectomy is the most effective primary prevention strategy. Prophylactic mastectomy is an aggressive surgical procedure that completely removes the mammary epithelial cells from which breast cancer arises along with the surrounding tissue. We seek to develop a minimally invasive intraductal procedure as an alternative to prophylactic mastectomy to locally ablate the mammary epithelial cells before they can become malignant. We and others have developed an intraductal delivery procedure to reach and treat these epithelial cells in rodent models of breast cancer. While the mouse mammary gland with a single non-anastomosed ductal tree opening at the nipple has a much less complex and tortuous architecture than the human breast, chemically induced and genetically engineered mouse models of breast cancer are valuable to produce proof-of-concept studies of new preventative strategies. Here, we describe a procedure for intraductal delivery of an ethanol-based ablative solution containing micro-CT/X-ray tantalum-based contrast agent within the mouse mammary ductal tree for the therapeutic purpose of primary prevention of breast cancer. Intraductal delivery of aqueous reagents (e.g., cytotoxic compounds, siRNAs, AdCre) has been previously described in mouse models. Thus, we focus our protocol description on methodological modifications and unique experimental considerations for optimizing delivery of ethanol, for minimizing local and systemic side effects of ethanol administration, and for in vivo visualization of ductal tree filling via micro-CT/fluoroscopy imaging. Visualization of the ductal tree immediately after injection of a contrast-containing solution allows for confirmation of complete filling or unsuccessful outcomes such as underfilling or overfilling. This procedure can be applied for delivery and imaging of other ablative compounds aimed at either preventing tumor formation or locally treating early-stage tumors accessible via the ductal tree.

A method of intraductal injection of reagents for an ethanol-based ablative solution to the mouse mammary ductal tree for in vivo imaging and breast cancer prevention is described. Injection directly into the nipple opening allows for targeting mammary epithelial cells with minimal collateral tissue damage.

导管内递送和X射线可视化乙醇烧蚀溶液在小鼠模型中预防和局部治疗乳腺癌

Breast cancer is the most prevalent cancer and the second-leading cause of cancer-related death for women in the USA. For high-risk women, prophylactic ...

癌症研究

X-Ray Visualization of Intraductal Ethanol-Based Ablative Treatment for Prevention of Breast Cancer in Rat Models

There are still a limited number of primary interventions for prevention of breast cancer. For women at a high risk of developing breast cancer, the most effective intervention is prophylactic mastectomy. This is a drastic surgical procedure in which the mammary epithelial cells that can give rise to breast cancer are completely removed along with the surrounding tissue. The goal of this protocol is to demonstrate the feasibility of a minimally invasive intraductal procedure that could become a new primary intervention for breast cancer prevention. This local procedure would preferentially ablate mammary epithelial cells before they can become malignant. Intraductal methods to deliver solutions directly to these epithelial cells in rodent models of breast cancer have been developed at Michigan State University and elsewhere. The rat mammary gland consists of a single ductal tree that has a simpler and more linear architecture compared to the human breast. However, chemically induced rat models of breast cancer offer valuable tools for proof-of-concept studies of new preventive interventions and scalability from mouse models to humans. Here, a procedure for intraductal delivery of an ethanol-based ablative solution containing tantalum oxide nanoparticles as X-ray contrast agent and ethyl cellulose as gelling agent into the rat mammary ductal tree is described. Delivery of aqueous reagents (e.g., cytotoxic compounds, siRNAs, AdCre) by intraductal injection has been described previously in mouse and rat models. This protocol description emphasizes methodological changes and steps that pertain uniquely to delivering an ablative solution, formulation consideration to minimize local and systemic side effects of the ablative solution, and X-ray imaging for in vivo assessment of ductal tree filling. Fluoroscopy and micro-CT techniques enable to determine the success of ablative solution delivery and the extent of ductal tree filling thanks to compatibility with the tantalum-containing contrast agent.

A procedure for the delivery of a chemical ablative solution to the rat mammary ductal tree for image-guided preventive treatment of breast cancer is described. Mammary epithelial cells can be targeted with minimal collateral tissue damage through cannulation directly into the nipple opening and intraductal infusion of a 70% ethanol-based ablative solution.

导管内乙醇消融治疗在大鼠模型中预防乳腺癌的X射线可视化

There are still a limited number of primary interventions for prevention of breast cancer. For women at a high risk of developing breast cancer, the most ...

In Vivo Gene Delivery into Mouse Mammary Epithelial Cells Through Mammary Intraductal Injection

Mouse mammary glands comprise ductal trees, which are lined by epithelial cells and have one opening at the tip of each nipple. The epithelial cells play a major role in mammary gland function and are the origin of most mammary tumors. Introducing genes of interest into mouse mammary epithelial cells is a critical step in evaluating gene function in epithelial cells and generating mouse mammary tumor models. This goal can be accomplished through the intraductal injection of a viral vector carrying the genes of interest into the mouse mammary ductal tree. The injected virus subsequently infects mammary epithelial cells, bringing in the genes of interest. The viral vector can be lentiviral, retroviral, adenoviral, or adenovirus-associated viral (AAV). This study demonstrates how a gene of interest is delivered into mammary epithelial cells through mouse mammary intraductal injection of a viral vector. A lentivirus carrying GFP is used to show stable expression of a delivered gene, and a retrovirus carrying Erbb2 (HER2/Neu) is used to demonstrate oncogene-induced atypical hyperplastic lesions and mammary tumors.

In Vivo Gene Delivery into Mouse Mammary Epithelial Cells Through Mammary Intraductal Injection

The present protocol describes intraductal injection of viral vectors via the teat to deliver genes of interest into the mammary epithelial cells.

体内 通过乳腺导管内注射将基因传递到小鼠乳腺上皮细胞

Mouse mammary glands comprise ductal trees, which are lined by epithelial cells and have one opening at the tip of each nipple. The epithelial cells play ...

Intranasal Immunization and Milk Collection in Studies of Maternal Immunization in New Zealand White Rabbits (Oryctolagus cuniculus)

Due to similarities in placentation and antibody transfer with humans, rabbits are an excellent model of maternal immunization. Additional advantages of this research model are the ease of breeding and sample collection, relatively short gestation period, and large litter sizes. Commonly assessed routes of immunization include subcutaneous, intramuscular, intranasal, and intradermal. Nonterminal sample collection for the chronological detection of the immunologic responses to these immunizations include the collection of blood, from both dams and kits, and milk from the lactating does. In this article, we will demonstrate techniques our lab has utilized in studies of maternal immunization in New Zealand White rabbits (Oryctolagus cuniculus), including intranasal immunization and milk collection.

Intranasal Immunization and Milk Collection in Studies of Maternal Immunization in New Zealand White Rabbits Oryctolagus cuniculus

This article describes and demonstrates the administration of intranasal vaccines and the collection of milk from lactating rabbits (Oryctolagus cuniculus) as a means to assess mucosal immunity in a translationally appropriate model of maternal immunization.

新西兰白兔产妇免疫研究中的内在免疫和牛奶收集

Due to similarities in placentation and antibody transfer with humans, rabbits are an excellent model of maternal immunization. Additional advantages of ...

免疫与感染

Intraductal Injection for Localized Drug Delivery to the Mouse Mammary Gland

Herein we describe a protocol to deliver various reagents to the mouse mammary gland via intraductal injections. Localized drug delivery and knock-down of genes within the mammary epithelium has been difficult to achieve due to the lack of appropriate targeting molecules that are independent of developmental stages such as pregnancy and lactation. Herein, we describe a technique for localized delivery of reagents to the mammary gland at any stage in adulthood via intraductal injection into the nipples of mice. The injections can be performed on live mice, under anesthesia, and allow for a non-invasive and localized drug delivery to the mammary gland. Furthermore, the injections can be repeated over several months without damaging the nipple. Vital dyes such as Evans Blue are very helpful to learn the technique. Upon intraductal injection of the blue dye, the entire ductal tree becomes visible to the eye. Furthermore, fluorescently labeled reagents also allow for visualization and distribution within the mammary gland. This technique is adaptable for a variety of compounds including siRNA, chemotherapeutic agents, and small molecules.

A protocol for the non-invasive intraductal delivery of aqueous reagents to the mouse mammary gland is described. The method takes advantage of localized injection into the nipples of mammary glands targeting mammary ducts specifically. This technique is adaptable for a variety of compounds including siRNA, chemotherapeutic agents and small molecules.

导管内注射局部给药的小鼠乳腺

Herein we describe a protocol to deliver various reagents to the mouse mammary gland via intraductal injections. Localized drug delivery and knock-down of ...

生物学

Simplifying Transcutaneous Intratracheal Drug Delivery in the Newborn Preterm Rabbit

Intratracheal (IT) drug delivery allows the direct delivery of pharmaceutical substances to the lung, maximizing potential pulmonary benefit and minimizing systemic drug exposure. The transcutaneous technique is simple and allows for the IT delivery of substances to the lung of prematurely born rabbits shortly after birth. Newborn pups are anesthetized with inhaled Isoflurane before being placed in a supine position with the neck extended. The larynx is identified and stabilized before transcutaneous placement of a 26-gauge (G) catheter into the trachea. Following catheterization of the trachea, a 30 G blunt needle attached to a Hamilton syringe is introduced into the IT catheter and is used for delivering a precise volume into the trachea during spontaneous respiration. After the IT injection is completed, the needle and catheter are withdrawn, and the pup is allowed to recover from anesthesia. Transcutaneous IT injection delivers a large proportion of the injected substance to the lung, with the majority remaining in the lung 3 hours after the intervention. The injections are well tolerated from the day of birth and can be repeated for multiple consecutive days without influencing survival. This technique can be used to investigate the effect of pharmaceutical agents on lung development and in the prevention of neonatal lung injury in preterm rabbits.

Transcutaneous intratracheal injection allows for effective intrapulmonary drug delivery during spontaneous respiration. Single and multiple injections are well tolerated with no effect on survival. The technique is simple to perform and can examine the effect of substances on lung development and the prevention of lung injury in newborn rabbits.

Intratracheal (IT) drug delivery allows the direct delivery of pharmaceutical substances to the lung, maximizing potential pulmonary benefit and ...

医学

Pregnancy and Nursing Management for Embryo-Transferred and Genetically Modified Rabbits

With the advancement of scientific research, the demand for gene-edited rabbit models is increasing. However, there are limited pregnancy and feeding management systems for gene-edited rabbits, leading to low survival rates among gene-edited rabbits prepared by many inexperienced researchers. Therefore, proper guidance is essential. This article summarizes the pregnancy and feeding practices for genetically modified rabbits developed in the author's laboratory and outlines a set of fundamental processes. These include pregnancy diagnosis, antenatal care, midwifery, assisted breastfeeding, weaning, and other procedures, along with the rescue and care of weak newborn rabbits. Compared to the traditional natural childbirth and nurturing methods used in rabbit farms, this approach involves more refined management, requiring additional time and effort but significantly increasing the survival rate of suckling rabbits. The methods described in this article are suitable for most laboratory breeding scenarios involving gene-edited or embryo-transferred rabbits and provide a straightforward and effective reference for other researchers.

This protocol describes a pregnancy and feeding management technique for embryo-transferred and genetically modified rabbits, aimed at reducing newborn rabbit mortality and enhancing the preparation efficiency of gene-edited rabbits.

胚胎转移和转基因兔的妊娠和哺乳管理

With the advancement of scientific research, the demand for gene-edited rabbit models is increasing. However, there are limited pregnancy and feeding ...

Intraductal Injection: Delivering Injection Mix into the Ducts of the Mouse Mammary Gland

Source: Xiang, D., et al. <a href="https://www.jove.com/v/59502/modeling-breast-cancer-via-an-intraductal-injection-cre-expressing">Modeling Breast Cancer via an Intraductal Injection of Cre-expressing Adenovirus into the Mouse Mammary Gland.</a> J. Vis. Exp (2019).
This video describes an intraductal injection technique where we deliver injection mix into the ducts of mouse mammary glands.

导管内注射:将注射混合物输送到小鼠乳腺的导管中

Source: Xiang, D., et al. Modeling Breast Cancer via an Intraductal Injection of Cre-expressing Adenovirus into the Mouse Mammary Gland. J. Vis. Exp ...

JoVE Encyclopedia of Experiments

乳腺癌

Procedures and techniques

Rabbit Intraductal Injection: Localized Delivery of Solution of Interest into the Rabbit Mammary Gland

Source: Clark et. al., <a href="https://www.jove.com/v/55209/intraductal-delivery-to-the-rabbit-mammary-gland">Intraductal Delivery to the Rabbit Mammary Gland.</a> J. Vis. Exp. (2017).
In this video we describe a technique of administering a solution of interest into the mammary duct of a rabbit and confirm the delivery by high-resolution ultrasound imaging of contrast agents.

兔导管内注射:将目标溶液局部输送到兔乳腺中

Source: Clark et. al., Intraductal Delivery to the Rabbit Mammary Gland. J. Vis. Exp. (2017).
In this video we describe a technique of administering a ...

导管内送货到兔乳腺

摘要

探索更多视频

此视频中的章节

家兔模型最佳胚胎阶段微创胚胎移植与胚胎玻璃化

导管内递送和X射线可视化乙醇烧蚀溶液在小鼠模型中预防和局部治疗乳腺癌

导管内乙醇消融治疗在大鼠模型中预防乳腺癌的X射线可视化

体内通过乳腺导管内注射将基因传递到小鼠乳腺上皮细胞

新西兰白兔产妇免疫研究中的内在免疫和牛奶收集

导管内注射局部给药的小鼠乳腺

Simplifying Transcutaneous Intratracheal Drug Delivery in the Newborn Preterm Rabbit

胚胎转移和转基因兔的妊娠和哺乳管理

导管内注射:将注射混合物输送到小鼠乳腺的导管中

兔导管内注射:将目标溶液局部输送到兔乳腺中

导管内送货到兔乳腺

摘要

探索更多视频

此视频中的章节

家兔模型最佳胚胎阶段微创胚胎移植与胚胎玻璃化

导管内递送和X射线可视化乙醇烧蚀溶液在小鼠模型中预防和局部治疗乳腺癌

导管内乙醇消融治疗在大鼠模型中预防乳腺癌的X射线可视化

体内 通过乳腺导管内注射将基因传递到小鼠乳腺上皮细胞

新西兰白兔产妇免疫研究中的内在免疫和牛奶收集

导管内注射局部给药的小鼠乳腺

Simplifying Transcutaneous Intratracheal Drug Delivery in the Newborn Preterm Rabbit

胚胎转移和转基因兔的妊娠和哺乳管理

导管内注射:将注射混合物输送到小鼠乳腺的导管中

兔导管内注射:将目标溶液局部输送到兔乳腺中

体内通过乳腺导管内注射将基因传递到小鼠乳腺上皮细胞