We thank Dr. John Chang (Radiation Oncology, LSU Health Science Center) for assistance with radiation dose measurements. The study was supported by the American Cancer Society (Grant number: 116945-RSG-09-038-01-CCE), National Institute of Health (Grant number: R21CA173162) and the Feist-Weiller Cancer Center.

所有的程序是由路易斯安那州立大学健康，什里夫波特，动物护理和使用委员会批准，并分别按照照顾和使用实验动物的指导方针NIH。 1.鼠颌下腺涎腺插管<ol><li> 注射器管组件的研制 <ol><li>切PE10聚乙烯管的一个10cm的长度用手术刀。容纳食指和拇指之间的管道的两端。加热上述平缓火焰管的中间部分，并轻轻拉伸软化的管道通过拉双方加倍其长度。 </li><li>在切割用手术刀中间管以45°角拿到2套管-每一个锥形端。通过在0.5毫升的胰岛素注射器的29克针嵌合它扩大插管的非锥形端。 </li><li>取出套管，并绘制220微升盐溶液进入注射器。点击赶走气泡。适合cannul一个在针，推动注射器的柱塞以排出空气，并确保通过套管的溶液自由流动。音量调整到200微升。 </li></ol></li><li> 颌下腺导管的口腔内插管 注:高压灭菌仪器之前的程序，并在两者之间进行消毒在热珠灭菌程序。 <ol><li>权衡只SD大鼠，并分配氯胺酮的计算量（42毫克/千克）/赛拉嗪（8毫克/千克）/乙酰丙嗪（1.4毫克/千克）的混合在一个皮下注射器。 </li><li>通过抓住食指和一只手的拇指之间的尾巴的基部和滑另一方面在主体把握它抑制动物。其余部分的食指和中指沿头部两侧，同时容纳用拇指和其余手指躯干。 </li><li>注入在后肢肌肉组织的麻醉剂。确认通过脚趾捏和眼睑反射麻醉深度。敷眼润滑剂防止干燥而动物处于麻醉状态。 </li><li>放置一个专门设计的平台（ 图1）上的动物，和横杆上接合的上门牙。通过循环围绕下门牙的橡胶带和它锚定到平台拉下颌向下。 </li><li>通过无菌缝合通过舌头将其提起，以提高口底。夹住缝合用止血，并且将它传递横杆。 </li><li>扩大与定制的脸颊吊具（ 图1）的脸颊，并在解剖显微镜下找到在口底舌下乳头。 </li><li>掌握使用细腻镊子预制PE10管的锥形端。轻轻操纵套管的尖端插入舌下乳头乳腺导管口。通过线程它3确认导管的放置 - 5 mm产品到管道;确保它传递而没有任何阻碍。 </li></ol></li><li> 颌下腺Instillation <ol><li>注射阿托品（0.5毫克/千克）皮下的颈背，并等待10分钟在唾液分泌减少。 </li><li>固定在插管到导管孔用氰基丙烯酸酯（胶）的下降，并使其干燥。灌输通过在〜50微升/分钟的速度缓慢地按下针筒柱塞在压盖（200微升/腺）的溶液中。 </li><li>粉碎用止血钳管道，并小心地取出注射器。保留在管道中的管道30 - 60分钟，直到动物恢复意识。卸下将舌头缝合。 </li><li>动物转移到一个单独的笼子里，并用热灯恢复过程中保持温暖。不要离开无人看管的动物，直到它已恢复了意识，以保持胸骨靠着。 </li><li>动物是完全卧床后，在不受限制地访问食物和水动物饲养安置它。 </li></ol></li></ol> 2.局部分割照射颌下腺<ol><li>抑制动物如之前描述的，并用氯胺酮肌内给药（33毫克/千克）/赛拉嗪（6毫克/公斤）/乙酰丙嗪（1毫克/千克）混合在后肢麻痹。确认麻醉深度，并使用润滑剂涂抹到眼睛。 </li><li>广场上的直线加速器桌面动物仰卧，并通过倾斜头部延长它的脖子。准直辐射场3厘米（狭缝宽度），以从下颌骨到胸骨的顶部的下边界包围的区域。 </li><li>放置1厘米组织等效丸剂以上的区域中，并调整辐射源和丸剂至100cm的顶部之间的距离。 </li><li>照射使用线性加速器的6 MV光子束的动物（2.5 Gy）进行。从辐射源剂量率，字段大小和距离弹丸表面将决定的曝光时间。在设置当前，动物在〜1戈瑞/分钟的剂量率照射。 </li><li>重复曝光;总共8 2.5格雷/天天; 4天/周，其间有2天的间隔。保持动物的体温恢复过程中。动物转移到动物饲养后，它完全卧床。 </li><li>收集刺激下颌下腺唾液8周照射后，测量腺体功能14。由冷的4％多聚甲醛/磷酸盐缓冲盐水pH 7.2的的心脏灌注安乐死麻醉下的动物。消灭颌下腺组织学和免疫组化分析14。 </li></ol>

<ol>
	<li>Delporte, C., 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=Increased+fluid+secretion+after+adenoviral-mediated+transfer+of+the+aquaporin-1+cDNA+to+irradiated+rat+salivary+glands.">Increased fluid secretion after adenoviral-mediated transfer of the aquaporin-1 cDNA to irradiated rat salivary glands.</a> Proc Natl Acad Sci U S A. 94 (7), 3268-3273 (1997).</li><li>Chambers, M. S., Rosenthal, D. I., Weber, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Radiation-induced+xerostomia.">Radiation-induced xerostomia.</a> Head Neck. 29 (1), 58-63 (2007).</li><li>Sciubba, J. J., Goldenberg, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oral+complications+of+radiotherapy.">Oral complications of radiotherapy.</a> Lancet Oncol. 7 (2), 175-183 (2006).</li><li>Rodrigues, N. 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=A+prospective+study+of+salivary+gland+function+in+lymphoma+patients+receiving+head+and+neck+irradiation.">A prospective study of salivary gland function in lymphoma patients receiving head and neck irradiation.</a> Int J Radiat Oncol Biol Phys. 75 (4), 1079-1083 (2009).</li><li>Coracin, F. L., 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=Major+salivary+gland+damage+in+allogeneic+hematopoietic+progenitor+cell+transplantation+assessed+by+scintigraphic+methods.">Major salivary gland damage in allogeneic hematopoietic progenitor cell transplantation assessed by scintigraphic methods.</a> Bone Marrow Transplant. 37 (10), 955-959 (2006).</li><li>Jensen, S. 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=A+systematic+review+of+salivary+gland+hypofunction+and+xerostomia+induced+by+cancer+therapies:+management+strategies+and+economic+impact.">A systematic review of salivary gland hypofunction and xerostomia induced by cancer therapies: management strategies and economic impact.</a> Support Care Cancer. 18 (8), 1061-1079 (2010).</li><li>de Castro, G., Federico, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation,+prevention+and+management+of+radiotherapy-induced+xerostomia+in+head+and+neck+cancer+patients.">Evaluation, prevention and management of radiotherapy-induced xerostomia in head and neck cancer patients.</a> Curr Opin Oncol. 18 (3), 266-270 (2006).</li><li>Epperly, M. W., Carpenter, M., Agarwal, A., Mitra, P., Nie, S., Greenberger, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intraoral+manganese+superoxide+dismutase-plasmid/liposome+(MnSOD-PL)+radioprotective+gene+therapy+decreases+ionizing+irradiation-induced+murine+mucosal+cell+cycling+and+apoptosis.">Intraoral manganese superoxide dismutase-plasmid/liposome (MnSOD-PL) radioprotective gene therapy decreases ionizing irradiation-induced murine mucosal cell cycling and apoptosis.</a> In Vivo. 18 (4), 401-410 (2004).</li><li>Cotrim, A. P., Sowers, A., Mitchell, J. B., Baum, B. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prevention+of+irradiation-induced+salivary+hypofunction+by+microvessel+protection+in+mouse+salivary+glands.">Prevention of irradiation-induced salivary hypofunction by microvessel protection in mouse salivary glands.</a> Mol Ther. 15 (12), 2101-2106 (2007).</li><li>Zheng, C., 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=Prevention+of+radiation-induced+salivary+hypofunction+following+hKGF+gene+delivery+to+murine+submandibular+glands.">Prevention of radiation-induced salivary hypofunction following hKGF gene delivery to murine submandibular glands.</a> Clin Cancer Res. 17 (9), 2842-2851 (2011).</li><li>Palaniyandi, 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=Adenoviral+delivery+of+Tousled+kinase+for+the+protection+salivary+glands+against+ionizing+radiation+damage.">Adenoviral delivery of Tousled kinase for the protection salivary glands against ionizing radiation damage.</a> Gene Ther. 18 (3), 275-282 (2011).</li><li>Baum, B. J., Voutetakis, A., Wang, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Salivary+glands:+novel+target+sites+for+gene+therapeutics.">Salivary glands: novel target sites for gene therapeutics.</a> Trends Mol Med. 10 (12), 585-590 (2004).</li><li>Limesand, K. H., 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=Insulin-like+growth+factor-1+preserves+salivary+gland+function+after+fractionated+radiation.">Insulin-like growth factor-1 preserves salivary gland function after fractionated radiation.</a> Int J Radiat Oncol Biol Phys. 78 (2), 579-586 (2010).</li><li>Timiri Shanmugam, P. 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=Recombinant+AAV9-TLK1B+administration+ameliorates+fractionated+radiation-induced+xerostomia.">Recombinant AAV9-TLK1B administration ameliorates fractionated radiation-induced xerostomia.</a> Hum Gene Ther. 24 (6), 604-612 (2013).</li><li>Coppes, R. P., Vissink, A., Konings, A. W. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+radiosensitivity+of+rat+parotid+and+submandibular+glands+after+different+radiation+schedules.">Comparison of radiosensitivity of rat parotid and submandibular glands after different radiation schedules.</a> Radiother Oncol. 63 (3), 321-328 (2002).</li><li>Sunavala-Dossabhoy, G., Palaniyandi, S., Richardson, C., De Benedetti, A., Schrott, L., Caldito, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=TAT-mediated+delivery+of+Tousled+protein+to+salivary+glands+protects+against+radiation-induced+hypofunction.">TAT-mediated delivery of Tousled protein to salivary glands protects against radiation-induced hypofunction.</a> Int J Radiat Oncol Biol Phys. 84 (1), 257-265 (2012).</li><li>Baum, B. 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=Transfer+of+the+AQP1+cDNA+for+the+correction+of+radiation-induced+salivary+hypofunction.">Transfer of the AQP1 cDNA for the correction of radiation-induced salivary hypofunction.</a> Biochim Biophys Acta. 1758 (8), 1071-1077 (2006).</li><li>Tran, S. 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=Paracrine+effects+of+bone+marrow+soup+restore+organ+function,+regeneration,+and+repair+in+salivary+glands+damaged+by+irradiation.">Paracrine effects of bone marrow soup restore organ function, regeneration, and repair in salivary glands damaged by irradiation.</a> PLoS One. 8 (4), e61632 (2013).</li><li>Nanduri, L. 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=Salisphere+derived+c-Kit++cell+transplantation+restores+tissue+homeostasis+in+irradiated+salivary+gland.">Salisphere derived c-Kit+ cell transplantation restores tissue homeostasis in irradiated salivary gland.</a> Radiother Oncol. 108 (3), 458-463 (2013).</li><li>Arany, S., Benoit, D. S., Dewhurst, S., Ovitt, C. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanoparticle-mediated+gene+silencing+confers+radioprotection+to+salivary+glands+in+vivo.">Nanoparticle-mediated gene silencing confers radioprotection to salivary glands in vivo.</a> Mol Ther. 21 (6), 1182-1194 (2013).</li><li>Voutetakis, 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=Reengineered+salivary+glands+are+stable+endogenous+bioreactors+for+systemic+gene+therapeutics.">Reengineered salivary glands are stable endogenous bioreactors for systemic gene therapeutics.</a> Proc Natl Acad Sci U S A. 101 (9), 3053-3058 (2004).</li></ol>

The authors have nothing to disclose.

唾液腺经常接受的辐射剂量超出经历了头颈部肿瘤，颈淋巴结选修消融或地区血液系统恶性肿瘤放疗的病人组织恢复的门槛。尽管腺的流体分泌腺泡细胞终末分化，它们是矛盾的对辐射敏感。分泌功能辐射和不可逆转的破坏腺体结果的第一周内滴在慢性低口水输出。为了打击穷人腺体功能和随之而来的口腔干燥，保存或分泌功能的恢复是至关重要的。新的治疗方法，以防止或修复辐射的伤害正在?...

健康组织的侧支破坏占了许多癌症治疗的有害副作用。与该辐射场位于大唾液腺的一部分或全部都不可避免地破坏。因此，经历骨髓移植前头部和颈部癌，子宫颈癌淋巴瘤，或全身辐射放疗大多数患者遭受的辐射，唾液腺机能减退2-6的最常见和最 ​​持久的不利效果之一。 唾液腺的流体产生腺泡细胞对辐射十分敏感。损坏唾液腺导致唾液流，称为唾液腺机能减退的条件的急剧减少。在唾液流量慢性减少损害的关键口腔活动，如咀嚼，吞咽，语言和口味，但剧烈的疼痛，黏膜撕裂，吞咽困难，机会性感染病态的后遗症，和龋齿恶化一个病人的福祉和功能2,3。 由于放疗相关的唾液腺细胞的损失是不可逆的，有口干没有矫正治疗。专注于人工唾液替代品和药品prosecretory平息目前的症状治疗无效的长期救援6。虽然改进的辐射递送技术已经帮助减少的病症的严重程度，正常组织的毒性及其并发症留在癌症治疗中6,7-一个限制因素。先发制人的措施，以防止放射治疗相关的并发症，因此，成为常态。正在探索的清除自由基的氧，促进细胞再增殖，或增强DNA修复放射保护剂，以避免唾液功能减退8-11。 外分泌腺唾液腺分泌物流入通过主排泄管道口。个的口腔内插管E对于造影剂注射排泄管道作为门诊手术常规做。利用类似的方法，唾液腺可以直接针对局部治疗12。除了降低的全身性副作用的风险，retroductal腺滴注增加的好处。围绕导管树唾液腺细胞的单层布置允许指定所有唾液腺上皮细胞，以及腺充当屏障纤维封装，以减少不必要的治疗性传播。从本质上讲，唾液腺是最适宜于腺体的苦难，如放射性唾液腺功能减退的有针对性的治疗。 （ - 2.5格雷/分/天，每周五天1.8），为期两周的用于癌症治疗常规辐射中的小剂量输送。因此，无线电防护治疗，显示在实验模型对旷日持久的辐射方案的疗效有较大的临床影响。康博分次辐射后mised唾液腺功能已被记录在小动物，但辐射源，剂量分数，和使用的协议是多种多样9,10,13。 该报告建立了retroductal交付使用病人相关的辐射源和剂量率大鼠颌下腺局部放射方法。

<table border="0" cellpadding="0" cellspacing="0" width="573">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Intramedic Polyethylene tubing (PE10)</td>
			<td style="width:71px;">Becton Dickson</td>
			<td align="right" style="width:119px;">427401</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">1/2 cc Insulin Syringe U-100</td>
			<td style="width:71px;">Becton Dickson</td>
			<td align="right" style="width:119px;">309306</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Artificial Tears</td>
			<td style="width:71px;">Miller Vet Supply&#160;</td>
			<td style="width:119px;">5098-9840-64</td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Hot Bead Sterilizer</td>
			<td style="width:71px;">Fine Science Tools</td>
			<td style="width:119px;">18000-45</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Perma-Hand silk suture</td>
			<td style="width:71px;">Ethicon</td>
			<td style="width:119px;">K833H</td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Graefe forcep</td>
			<td style="width:71px;">Fine Science Tools</td>
			<td style="width:119px;">11051-10</td>
			<td></td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Olympus SZX16 Stereo Microscope</td>
			<td style="width:71px;">Hunt Optics and Imaging</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">6MV Linear Accelerator</td>
			<td style="width:71px;">Elekta</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Bolus - Skinless&#160;</td>
			<td style="width:71px;">Civco</td>
			<td style="width:119px;">MTCB410</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Heat Lamp</td>
			<td style="width:71px;">Braintree Scientific</td>
			<td style="width:119px;">HL-1&#160;110V</td>
			<td></td>
		</tr>
	</tbody>
</table>

retroductal submandibular gland instillation and localized fractionated irradiation in a rat model of salivary hypofunction

Normal tissues that lie within the portals of radiation are inadvertently damaged. Salivary glands are often injured during head and neck radiotherapy. Irreparable cell damage results in a chronic loss of salivary function that impairs basic oral activities, and increases the risk of oral infections and dental caries. Salivary hypofunction and its complications gravely impact a patient's comfort. Current symptomatic management of the condition is ineffective, and newer therapies to assuage the condition are needed. 
Salivary glands are exocrine glands, which expel their secretions into the mouth via excretory ducts. Cannulation of these ducts provides direct access to the glands. Retroductal delivery of a contrast agent to major salivary glands is a routine out-patient procedure for diagnostic imaging. Using a similar procedure, localized treatment of the glands is feasible. However, performing this technique in preclinical studies with small animals poses unique challenges. In this study we describe the technique of retroductal administration in rat submandibular glands, a procedure that was refined in Dr. Bruce Baum's laboratory (NIH)1, and lay out a procedure for local gland irradiation.

适应微创技术造影，大唾液腺的局部治疗是可行的。在大鼠颌下腺唾液腺Retroductal给药试图通过沃顿管道口腔内插管（ 图2）。沃顿的唾液腺导管上位于口底的舌下乳头打开，但这些孔不容易看到。插管的插入物，因此，通过温和探测完成。为了避免不良出血或穿孔管，同时将套管没有使用武力。在导管插管的光滑无电阻的通道是由管子的平缓来回运动证实。 ...

Watch this Scientific Journal Video about Retroductal Submandibular Gland Instillation and Localized Fractionated Irradiation in a Rat Model of Salivary Hypofunction at JoVE.com

Retroductal Submandibular Gland Instillation and Localized Fractionated Irradiation in a Rat Model of Salivary Hypofunction

Salivary gland hypofunction, a major adverse effect of head and neck radiotherapy diminishes a patient's quality of life. The demonstration of efficacy of new therapies in animal models is a prerequisite before clinical transition. This protocol describes retroductal administration and local irradiation of rat submandibular glands.

Retroductal颌下腺灌输和本地化分割照射在涎腺功能减退症大鼠模型

Normal tissues that lie within the portals of radiation are inadvertently damaged. Salivary glands are often injured during head and neck radiotherapy. ...

retroductal-submandibular-gland-instillation-localized-fractionated

Research

JoVE Journal

Medicine

11.3K 次观看.  Louisiana State University Health Sciences Center. Salivary gland hypofunction, a major adverse effect of head and neck radiotherapy diminishes a patient's quality of life. The demonstration of efficacy of new therapies in animal models is a prerequisite before clinical transition. This protocol describes retroductal administration and local irradiation of rat submandibular glands.

视频: Retroductal Submandibular Gland Instillation and Localized Fractionated Irradiation in a Rat Model of Salivary Hypofunction

Cannulation of the Mouse Submandibular Salivary Gland via the Wharton's Duct

Severe salivary gland hypofunction is frequently found in patients with Sj&ouml;gren's syndrome and those who receiving therapeutic 
irradiation in their head and neck regions for cancer treatment. Both groups of patients experience symptoms such as xerostomia (dry mouth), dysphagia 
(impaired chewing and swallowing), severe dental caries, altered taste, oro-pharyngeal infections (candidiasis), mucositis, pain and discomfort.
One innovative approach of regenerative medicine for the treatment of salivary gland hypo-function is speculated in RS Redman, E Mezey 
et al. 2009: stem cells can be directly deposited by cannulation into the gland as a potent method in reviving the functions of the impaired organ. Presumably, 
the migrated foreign stem cells will differentiate into glandular cells to function as part of the host salivary gland. Also, this cannulation technique is an 
expedient and effective delivery method for clinical gene transfer application.
Here we illustrate the steps involved in performing the cannulation procedure on the mouse submandibular salivary gland via the Wharton's duct 
(Fig 1). C3H mice (Charles River, Montreal, QC, Canada) are used for this experiment, which have been kept under clean conventional conditions at 
the McGill University animal resource center. All experiments have been approved by the University Animal Care Committee and were in accordance with the 
guidelines of the Canadian Council on Animal Care.
For this experiment, a trypan blue solution is infused into the gland through the opening of the Wharton's duct using a 
insulin syringe with a 29-gauge needle encased inside a polyethylene tube. Subsequently, the mouse is dissected to show that the infusions migrated 
into the gland successfully.

A protocol for the cannulation of the mouse submandibular salivary gland via the Wharton's duct is described. For this experiment, the trypan blue solution is used as a dyer to demonstrate how this technique effectively delivers infusions into the targeted gland, and to suggest the reliability of this new approach as a potential clinical drug/cell therapy for the regeneration of salivary glands.

沃顿商学院的导管插管通过小鼠颌下腺唾液腺

Severe salivary gland hypofunction is frequently found in patients with Sj&ouml;gren's syndrome and those who receiving therapeutic 
irradiation in their ...

科研

医学

A Model of Chronic Nutrient Infusion in the Rat

Chronic exposure to excessive levels of nutrients is postulated to affect the function of several organs and tissues and to contribute to the development of the many complications associated with obesity and the metabolic syndrome, including type 2 diabetes. To study the mechanisms by which excessive levels of glucose and fatty acids affect the pancreatic beta-cell and the secretion of insulin, we have established a chronic nutrient infusion model in the rat. The procedure consists of catheterizing the right jugular vein and left carotid artery under general anesthesia; allowing a 7-day recuperation period; connecting the catheters to the pumps using a swivel and counterweight system that enables the animal to move freely in the cage; and infusing glucose and/or Intralipid (a soybean oil emulsion which generates a mixture of approximately 80% unsaturated/20% saturated fatty acids when infused with heparin) for 72 hr. This model offers several advantages, including the possibility to finely modulate the target levels of circulating glucose and fatty acids; the option to co-infuse pharmacological compounds; and the relatively short time frame as opposed to dietary models. It can be used to examine the mechanisms of nutrient-induced dysfunction in a variety of organs and to test the effectiveness of drugs in this context.

A protocol for chronic infusions of glucose and Intralipid in rats is described. This model can be used to study the impact of nutrient excess on organ function and physiological parameters.

在大鼠模型慢性营养输液

Chronic exposure to excessive levels of nutrients is postulated to  affect the function of several organs and tissues and to contribute to the  ...

Collecting Saliva and Measuring Salivary Cortisol and Alpha-amylase in Frail Community Residing Older Adults via Family Caregivers

Salivary measures have emerged in bio-behavioral research that are easy-to-collect, minimally invasive, and relatively inexpensive biologic markers of stress. This article we present the steps for collection and analysis of two salivary assays in research with frail, community residing older adults-salivary cortisol and salivary alpha amylase. The field of salivary bioscience is rapidly advancing and the purpose of this presentation is to provide an update on the developments for investigators interested in integrating these measures into research on aging. Strategies are presented for instructing family caregivers in collecting saliva in the home, and for conducting laboratory analyses of salivary analytes that have demonstrated feasibility, high compliance, and yield quality specimens. The protocol for sample collection includes: (1) consistent use of collection materials; (2) standardized methods that promote adherence and minimize subject burden; and (3) procedures for controlling certain confounding agents. We also provide strategies for laboratory analyses include: (1) saliva handling and processing; (2) salivary cortisol and salivary alpha amylase assay procedures; and (3) analytic considerations.

We demonstrate: (1) procedures for collection of salivary samples in cognitive impaired older adults by family caregivers in the home setting, (2) procedures for measuring stress activity via salivary cortisol and alpha amylase, and (3) representative profiles. Protocols that allow researchers to study stress-linked processes advance our understanding of biological sensitivity and susceptibility.

Salivary measures have emerged in bio-behavioral research  that are easy-to-collect, minimally invasive, and relatively inexpensive  biologic markers of ...

Glaucoma-inducing Procedure in an In Vivo Rat Model and Whole-mount Retina Preparation

Glaucoma is a disease of the central nervous system affecting retinal ganglion cells (RGCs). RGC axons making up the optic nerve carry visual input to the brain for visual perception. Damage to RGCs and their axons leads to vision loss and/or blindness. Although the specific cause of glaucoma is unknown, the primary risk factor for the disease is an elevated intraocular pressure. Glaucoma-inducing procedures in animal models are a valuable tool to researchers studying the mechanism of RGC death. Such information can lead to the development of effective neuroprotective treatments that could aid in the prevention of vision loss. The protocol in this paper describes a method of inducing glaucoma - like conditions in an in vivo rat model where 50 &#181;l of 2 M hypertonic saline is injected into the episcleral venous plexus. Blanching of the vessels indicates successful injection. This procedure causes loss of RGCs to simulate glaucoma. One month following injection, animals are sacrificed and eyes are removed. Next, the cornea, lens, and vitreous are removed to make an eyecup. The retina is then peeled from the back of the eye and pinned onto sylgard dishes using cactus needles. At this point, neurons in the retina can be stained for analysis. Results from this lab show that approximately 25% of RGCs are lost within one month of the procedure when compared to internal controls. This procedure allows for quantitative analysis of retinal ganglion cell death in an in vivo rat glaucoma model.

Glaucoma-inducing Procedure in an In Vivo Rat Model and Whole-mount Retina Preparation

Glaucoma is characterized by damage to retinal ganglion cells. Inducing glaucoma in animal models can provide insight into the study of this disease. Here, we outline a procedure that induces loss of RGCs in an in vivo rat model and demonstrates the preparation of whole-mount retinas for analysis.

青光眼诱导在程序<em&gt;在体内</em&gt;大鼠模型和全安装视网膜准备

Glaucoma is a disease of the central nervous system affecting retinal ganglion cells (RGCs). RGC axons making up the optic nerve carry visual input to the ...

Murine Salivary Functional Assessment via Pilocarpine Stimulation Following Fractionated Radiation

Hyposalivation is commonly observed in the autoimmune reaction of Sj&#246;gren's syndrome or following radiation injury to the major salivary glands. In these cases, questions remain regarding disease pathogenesis and effective interventions. An optimized technique that allows functional assessment of the salivary glands is invaluable for investigating exocrine gland biology, dysfunction, and therapeutics. Here, we present a step by step approach to performing pilocarpine stimulated saliva secretion, including tracheostomy and the dissection of the three major murine salivary glands. We also detail the appropriate murine head and neck anatomy accessed during these techniques. This approach is scalable, allowing for multiple mice to be processed simultaneously, thus improving the efficiency of the work flow. We aim to improve the reproducibility of these methods, each of which has further applications within the field. In addition to saliva collection, we discuss metrics for quantifying and normalizing functional capacity of these tissues. Representative data are included from submandibular glands with depressed salivary gland function 2 weeks following fractionated radiation (4 doses of 6.85 Gy).

We present a detailed approach to performing saliva collection, including murine tracheostomy and the isolation of three major salivary glands.

小鼠涎腺照射后通过匹属刺激进行唾液功能评估

Hyposalivation is commonly observed in the autoimmune reaction of Sj&#246;gren's syndrome or following radiation injury to the major salivary glands. In ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

用线 Myography 测定小鼠肠系膜动脉的等轴收缩力

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

A Pleural Effusion Model in Rats by Intratracheal Instillation of Polyacrylate/Nanosilica

Pleural effusion is a prevalent clinical finding of many pulmonary diseases. Having a useful animal pleural effusion model is very important to study these pulmonary diseases. Previous pleural effusion models paid more attention to biological factors rather than nanoparticles in the environment. Here, we introduce a model to make pleural effusion in rats by intratracheal instillation of polyacrylate/nanosilica, and a method of nanoparticle isolation in the pleural effusion. By intratracheal instillation of polyacrylate/nanosilica with concentrations of 3.125, 6.25 and 12.5 mg/kg&#8729;mL, the pleural effusion in rats presented on day 3, peaked at days 7-10 in 6.25 and 12.5 mg/kg&#8729;mL groups, then slowly decreased and disappeared on day 14. When the concentration of polyacrylate/nanosilica increased, the pleural effusion is produced more and faster. This pleural fluid was detected by ultrasound examination or CT chest scanning and confirmed by dissection of rats. Silica nanoparticles were observed in the rats' pleural effusion by transmission electron microscope. These results showed that the exposure to polyacrylate/nanosilica leads to the induction of pleural effusion, which was consistent with our previous report in humans. Additionally, this model is beneficial for the further study of nanotoxicology and the pleural effusion diseases.

Here, we present a protocol to construct a pleural effusion model in rats by intratracheal instillation of polyacrylate/nanosilica.

聚丙烯酸/纳米水的气管内灌注积液模型

Pleural effusion is a prevalent clinical finding of many pulmonary diseases. Having a useful animal pleural effusion model is very important to study ...

Radiation Treatment of Organotypic Cultures from Submandibular and Parotid Salivary Glands Models Key In Vivo Characteristics

Hyposalivation and xerostomia create chronic oral complications that decrease the quality of life in head and neck cancer patients who are treated with radiotherapy. Experimental approaches to understanding mechanisms of salivary gland dysfunction and restoration have focused on in vivo models, which are handicapped by an inability to systematically screen therapeutic candidates and efficiencies in transfection capability to manipulate specific genes. The purpose of this salivary gland organotypic culture protocol is to evaluate maximal time of culture viability and characterize cellular changes following ex vivo radiation treatment. We utilized immunofluorescent staining and confocal microscopy to determine when specific cell populations and markers are present during a 30-day culture period. In addition, cellular markers previously reported in in vivo radiation models are evaluated in cultures that are irradiated ex vivo. Moving forward, this method is an attractive platform for rapid ex vivo assessment of murine and human salivary gland tissue responses to therapeutic agents that improve salivary function.

Using three-dimensional organotypic cultures to visualize morphology and functional markers of salivary glands may provide novel insights into the mechanisms of tissue damage following radiation. Described here is a protocol to section, culture, irradiate, stain, and image 50&#8211;90 &#956;m thick salivary gland sections prior to and following exposure to ionizing radiation.

下颌骨和腮腺表型模型对组织型培养的放射治疗是机体活力特征的关键

Hyposalivation and xerostomia create chronic oral complications that decrease the quality of life in head and neck cancer patients who are treated with ...

Acupuncture in a Rat Model of Asthma

A high platform can fix rats without restriction and completely expose the acupoints on the back during acupuncture manipulation. This article describes methods for the fabrication of the high platform, establishes a rat model of asthma and measures changes in respiratory function using a noninvasive and real-time whole-body plethysmography (WBP) system.

哮喘鼠模型中的针灸

A high platform can fix rats without restriction and completely expose the acupoints on the back during acupuncture manipulation. This article describes ...

Retinal Pathophysiological Evaluation in a Rat Model

A posterior segment eye disease like diabetic retinopathy alters the physiology of the retina. Diabetic retinopathy is characterized by a retinal detachment, breakdown of the blood-retinal barrier (BRB), and retinal angiogenesis. An in vivo rat model is a valuable experimental tool to examine the changes in the structure and function of the retina. We propose three different experimental techniques in the rat model to identify morphological changes of retinal cells, retinal vasculature, and compromised BRB. Retinal histology is used to study the morphology of various retinal cells. Also, quantitative measurement is performed by retinal cell count and thickness measurement of different retinal layers. A BRB breakdown assay is used to determine the leakage of extraocular proteins from the plasma to vitreous tissue due to the breakdown of BRB. Fluorescence angiography is used to study angiogenesis and leakage of blood vessels by visualizing retinal vasculature using FITC-dextran dye.

Diabetic retinopathy is one of the leading causes of blindness. Histology, blood-retinal barrier breakdown assay, and fluorescence angiography are valuable techniques to understand the pathophysiology of the retina, which could further enhance the efficient drug screening against diabetic retinopathy.