This study was supported and funded by the Avenir Foundation (Fondation de l'Avenir, 255 rue de Vaugirard, 75719 Paris cedex 15, Paris, France). This study would never have been conducted without the precious help of the veterinary team of the Laboratory of Biosurgical Research from the Alain Carpentier Foundation.

所有的动物都根据动物研究伦理委员会（法国农业部的指导方针）处理。该协议获得在巴黎第五大学授权的动物experimentations当地伦理委员会的批准（注册号MESR 2035.02;医学学部巴黎笛卡尔，法国巴黎）。 1. ADSC文化和标签<ol><li>获得一个私人机构证实脂肪干细胞。在37℃和5％的CO 2与含10％胎儿小牛血清和1％抗生素（青霉素和链霉素）的α极限必需培养基培养同种异体脂肪干细胞。 </li><li>使用标准的细胞扩增程序（37℃和5％CO 2）得到每只动物约24×10 6个脂肪干细胞。收获使用标准的胰蛋白酶溶液（10毫升的T150皿）的脂肪干细胞。用血球进行细胞计数。 </li><li>执行小区使用PKH67染色PR标记移植前一天ocedure如下所述。 注:在染色过程的目标是允许移植后细胞跟踪。 <ol><li>为了标注2×10 6的脂肪干细胞，准备溶液A（2×10 6细胞用100微升稀释的C）和溶液B（PKH67 4μL用1毫升稀释液的C）。 </li><li>孵育溶液A和B为2分钟。然后，添加100μL胎儿小牛血清1分钟。 </li><li>冲洗以1500 xg离心与柠檬酸的RPMI 300微升的溶液（RPMI的298.5微升和柠檬酸1.5μL），离心5分钟。重复此操作2次。保持标记细胞，并保护他们免受光，直到板结构。 </li></ol></li></ol> 2.双击ADSC表构建<ol><li>一天移植前，准备与每细胞培养基的阱4毫升，如上所述，PKH标记之前12孔温度响应细胞培养皿。种子菜1.5×10 6 PKH-labele每孔D细胞孵育12小时（37℃和5％CO 2）。 </li><li>上移植的当天，通过温育在室温下将培养皿30分钟分离汇合细胞片。 </li><li>轻轻吸了吸取1 ADSC表和图层它的疏水纸（1.5厘米直径）。使用这种纸张作为支撑膜。 </li><li>掌握另一ADSC片，轻轻地沉积其上的另一个的顶部，以获得一个双层构建体。 </li><li>重复这些程序根据需要（4每只动物），得到尽可能多的ADSC表结构。 </li></ol> 3.食管内镜黏膜下剥离术（ESD） <ol><li>上移植的当天，用10毫克/千克肌内氯胺酮预先用药的动物，并以8毫克/公斤静脉内异丙酚诱导它们。然后，执行气管插管并保持与异氟烷2.5％吸入麻醉。 </li><li>一旦动物是在全身麻醉下，进行ESD与gastroscOPE，一个视频内窥镜，以及电外科单元。 <ol><li>使用内窥镜刀和软凝固，以获得半周背侧标记范围从40厘米到从牙弓45厘米。 </li><li>注入含有靛蓝胭脂红染料成用于粘膜层从肌肉层分离的粘膜下层甘油溶液。 </li><li>使用内窥镜刀endocut I模式来获得圆周切口。 </li><li>使用的内窥镜刀强迫凝固模式来执行粘膜解剖，从近端切口移动到远端切口。 </li></ol></li><li>在静电放电过程结束时，观察半周，5-厘米长的食管瘢痕露出肌层。 </li></ol> 4.移植内镜<ol><li>静电放电后，立即内镜移植动物4双ADSC表结构。 <ol><li>掌握内镜钳和ADSC表结构通过使用大量的，透明的内窥镜盖保护它运输到创伤部位中。 </li><li>的ADSC表构建体的整个表面施加到溃疡床。 </li><li>轻轻地应用ADSC表结构为10分钟获得一个完整的，稳定的植入。为应用程序，使用内窥镜钳子或内窥镜帽并应用ADSC的整个表面构造到食道粘膜下层。 </li></ol></li><li>重复此步骤，对每个动物四次，以获得4移栽双贴壁细胞表结构。 注:我们的目标是覆盖近似一半的受伤面积。 </li></ol> 5.术后评估及随访<ol><li>对于每个动物，确保后的ESD镇痛为0.2毫克/千克吗啡为第一天并用埃索美拉唑的28天的抗酸处理（40毫克/天），每天三次肌内注射。执行与amoxicill抗生素预防在（1克/天）7天。授权于翌日第1天的液体和固体。 </li><li>跟进动物28天。获得使用圆润平卡斯吞咽困难得分16，体重变化的测量日常临床评估。 </li><li>多式联运狭窄的评价:全身麻醉，在预定的3天，14和28下，担负狭窄发生多式联运评价。 <ol><li>执行使用瘢痕描述，狭窄测量和纸张支撑膜检测内窥镜评价。使用胃镜通过狭窄跨越测量狭窄（胃镜的直径为8毫米）。观察疤痕，关注炎性方面（粘膜红斑，粘膜水肿，和粘膜自发性出血）和瘢痕内不存在或纸张支撑膜的存在。 </li><li>通过胃镜操作者运河直接申请pCLE绿色探头插入疤痕床。搜索一个自发并组织与PKH67标记ADSC表结构植入兼容绿色信号。 </li><li>执行重晶石食管造影的2正交发生率与放射性箍（正面和左路的发生率）。保持为狭窄评价最紧密的发生率（的狭窄（％度）= [1-（狭窄下短轴长度/狭窄下正常轴）的长度×100）]）17，18。 </li></ol></li><li>动物牺牲。 <ol><li>在第28天，在随访期结束时，执行动物牺牲。虽然动物仍然是在全身麻醉下，注入100毫克/公斤静脉注射苯巴比妥。确保动物死亡利用临床检查为缺乏心跳和呼吸运动。 </li><li>胸骨切开术和器官曝光后，取出食道从每个动物处死（用10％福尔马林固定，石蜡包埋，4微米厚的部分）。染色幻灯片用苏木和藏红花15。对于数字化计算机分析幻灯片和比较动物组15。 </li></ol></li></ol>

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作者什么都没有透露。

在这种猪模型中，ADSC表移植在技术上是成功的，并允许细胞植入监测体内 pCLE评价。临床，内窥镜，放射性，和组织学评价证明增强的ESD后在食管狭窄预防的内窥镜ADSC片的有效性。 含有靛蓝胭脂红染料片的ADSC甘油溶液的内窥镜移植是在再生医学的创新方法。大木等。第一次描述了内镜移植过程。在他们的研究中，细胞表构建体组成层叠在聚偏二氟乙烯的薄片口腔粘膜上皮细胞片，用作载体膜10，11，19的。支持膜与内镜钳抓住并小心地放在床上溃疡。切尔温和应用的10分钟后，获得的L-片的粘附性。这个过程还没有被再现。在这项研究中，双ADSC表构建物移植多种原因。 ADSC的选择是基于几个研究，证明了其抗炎能力，特别是在皮肤愈合过程。脂肪干细胞是公知的调节角质形成细胞和成纤维细胞的相互作用7。纤维化发展的机制是复杂的，仅部分的理解。的旁分泌效应似乎发挥ADSC动作的主要作用，包括抗炎因子的生产，如TGFβ和促血管生成因子20。 内镜ADSC表移植有几个限制。首先，被应用细胞片10分钟以食管瘢痕获得稳定的粘合性。在这部分程序是具有挑战性的，技术上的困难，不得不由有经验的内镜医师进行。然而，所有的动物都成功地移植，确认了内窥镜检查过程的可行性。最近，以改善细胞片移植的成功使用3D印刷装置的报告21。第二，这种技术具有未知存活率有关。食管瘢痕的细胞一个积极的环境。温度响应盘允许生产的细胞片的构造通过细胞外基质和细胞的连接的形成。因此，为了提高生存率和ADSC相互作用，一个双ADSC表构建体被选择22,23。这项研究并不是用来评估两种细胞存活率或移植对食管狭窄预防剂量效应，这两者都是重要的问题。第三，如在大木等。 ，使用支撑膜的必要携带ADSC表构建体。前前后后呃移植，纸支撑膜不能没有构造破坏拆除。此帧内食管异物负责增加炎症。一个优化的技术将允许移除纸张支撑膜。 与pCLE移植的监测也具有挑战性。这个新的内窥镜成像技术可以在体内的组织学分析，包括腺图案的可视化，微血管和细胞的改变。 pCLE在许多消化系统疾病，如炎性肠疾病，巴雷特食管，和粘液性胰腺囊肿24,25进行评价。所述pCLE兼容PKH67荧光团被用于细胞标记。与ADSC片的存在相容的荧光信号在4只动物中观察到第3天这肯定结果反映的ADSC表移植的成功。然而，贝科使用PKH67具有降低的信号随着时间的推移和细胞分裂后，pCLE评价移植26后仅几天是可能的。大木等。和金井等人。通过免疫染色监测细胞片移植，和他们证实细胞的存在，直到移植后8一天。成熟细胞在这项研究中，具有特定的免疫染色的优点，使用。 食管狭窄是以下的ESD的最严重的不良事件，从而限制了技术发展中的一个。最佳的技术来防止狭窄形成仍然缺乏。在临床实践中，患者具有扩展食管癌的ESD被规定不同的预防性治疗方法，如皮质类固醇，内窥镜球囊扩张术，或食道支架置入27。生物材料的局部应用已经进行，结果各不相同28,29 </SUP&gt;。再生医学的目的是要重建使用各种方法，如组织替代或局部免疫调节的组织或器官。对组织替代，使用具有或不具有上皮细胞的生物支架在文献30,31进行评价。在食管瘢痕观察到早熟，提高再上皮化。对于局部免疫调节，小数据是在大约ADSC注射或羊膜应用32,33的文献报道。一个延迟的，狭窄的形成和降低狭窄率进行观察。 在这种猪模型，成功地进行了内镜细胞表移植过程。在双细胞片移植组织的脂肪干细胞表现出了对防止超期ESD后食管狭窄形成能力。参与ADSC愈合机制过程知之甚少，必须在今后的研究进一步调查。因此，鉴于这些有前景的结果的，临床试验应进行评估扩展后的ESD患者ADSC表构建体的效力。

浅表性食管肿瘤的管理层已与新的内镜技术的发展变化。如今，内窥镜切除是一线治疗。事实上，它是具有较低的发病率和死亡率比等于肿瘤结果1，2，3一手术相关。内窥镜粘膜切除术（EMR）和内镜黏膜下切除术（ESD）是最广泛使用的技术。在扩展浅表肿瘤的情况下，静电放电是优选的。相比EMR，ESD允许整块切除，无论病灶大小和形状4，5，6。 ESD的主要延迟并发症是食道狭窄的形成，这通常发生一周和两周之间切除后。最近公布的研究表明，狭窄形成是相关的切除的大小。日本内镜学会建议避免ESD尺寸大于¾食管周长，因为它们是在病例的90％以上与狭窄发展相关联，并且负责严重馈送麻烦和生活质量严重恶化的要大。 食管狭窄的预防仍然是具有挑战性的。参与狭窄形成机制只知道部分。狭窄的地层仿佛来自两个不同的机制，该协会导致:（1）促炎症细胞招聘和（2）过度纤维化发展7。几个预防性治疗方法已被提出。然而，结果并不令人满意，有什么好处和严重的副作用8,9。最近，日本队，大木等人。 ，提出了自体口腔粘膜细胞的单层细胞片移植到esophageal疤痕。在ESD 10,11后立即进行移植。他们证明这种创新方法的有效性，首先在犬模型，然后在病人。 脂肪组织来源的基质细胞（脂肪干细胞）是有希望在再生医学。其在几个领域中的应用已表明有趣的结果，特别是在伤口愈合过程。 ADSC疗法提供了几个优点，因为细胞是容易分离的，并具有抗炎特性，局部免疫调节作用，血管新生诱导，分化能力为间充质和非间充质系12，13，14相关联。 在先前的研究中，我们的团队表现出双ADSC表镜下移植治疗食管狭窄上一页成效在猪模型15增强的ESD后ention。在这篇文章中，ADSC板结构和内窥镜移植技术的报告介绍。

<table><tbody><tr><td>Transparent endoscopic cap Q180 compatible</td><td>Olympus Optical Co</td><td></td><td></td></tr><tr><td>GIF-Q180 gastroscope&amp;nbsp;</td><td>Olympus Optical Co</td><td></td><td></td></tr><tr><td>Videoscope System Exera II&amp;nbsp;</td><td>Olympus Optical Co</td><td></td><td></td></tr><tr><td>Injection needle 18 G</td><td>Olympus Optical Co</td><td></td><td></td></tr><tr><td>Electrosurgery unit ERBE ICC 350&amp;nbsp;</td><td>ERBE Technology</td><td></td><td></td></tr><tr><td>Indigo carmin 1%</td><td>Life</td><td></td><td></td></tr><tr><td>Endoscopic hybrid knife</td><td>Life</td><td></td><td></td></tr><tr><td>Minisonde Z pCLE green probe</td><td>Mauna Kea Technology</td><td></td><td>You must learn how to use the probe. The manipulation could be difficult.</td></tr><tr><td>Fetal bovine serum</td><td>Sigma Aldrich</td><td>12105C</td><td></td></tr><tr><td>Trypsin</td><td>Sigma Aldrich</td><td>T146</td><td></td></tr><tr><td>Alpha minimum essential medium</td><td>Thermo Fisher</td><td>22561-021</td><td></td></tr><tr><td>Phosphate-Buffered Salines</td><td>Thermo Fisher</td><td>10010-023</td><td></td></tr><tr><td>PKH67 dye kit</td><td>Sigma Aldrich</td><td>Mini67-KT</td><td></td></tr><tr><td>12-well temperature responsive cell culture dish</td><td>Upcell Thermo Scientific</td><td>174900</td><td>Feel the weel with 4 mL standard medium culture 30 min before seeding cells</td></tr><tr><td>Esomeprazole 40 mg&amp;nbsp;</td><td>Biogaran</td><td></td><td></td></tr><tr><td>Moprhine sulfate 50 mg/mL</td><td>Lavoisier</td><td></td><td></td></tr><tr><td>Amoxicilline 1 g</td><td>Biogaran</td><td></td><td></td></tr><tr><td>Ketamine 250 mg/5 mL</td><td>Panpharma</td><td></td><td></td></tr><tr><td>Propofol 10 mg/mL</td><td>Fresenius</td><td></td><td></td></tr><tr><td>Hydrophobic paper</td><td>Carrefour</td><td></td><td></td></tr></tbody></table>

adsc-sheet transplantation to prevent stricture after extended esophageal endoscopic submucosal dissection

在过去几年中，细胞的表结构刺激了再生医学的广泛兴趣，尤其是对重建手术程序。多样化的技术脂肪组织衍生的基质细胞（脂肪干细胞）与各种生物材料相结合的发展导致了多种类型的组织工程替代品，如骨，软骨，和从啮齿动物，猪或人脂肪干细胞的脂肪组织的结构。扩展食管内镜黏膜下剥离术（ESD）负责食管狭窄的形成。狭窄预防仍然是具有挑战性的，没有可用的有效的治疗方法。先前的研究报道了犬模型和人类中的粘膜细胞片移植的有效性。脂肪干细胞归因抗炎特性，局部免疫调节作用，血管新生诱导，分化能力为间充质和非间充质细胞谱系。这个原始的研究描述了TRA内镜一个ADSC组织工程构建的nsplantation防止食管狭窄的猪模型。在ADSC结构是由一纸支撑膜在彼此分层2同种异体ADSC表。将脂肪干细胞标记的PKH67荧光允许基于探针的共聚焦激光显微内镜（pCLE）监控。上移植当天，进行5厘米和已知诱导食管狭窄半周的ESD。动物立即用胃镜ADSC 4移植结构。温和应用的10分钟后，获得的ADSC构建体的完整的粘合。动物处死28天所有动物均成功移植。移植证实第3天以积极的pCLE的评价。相比于移植的动物，对照组动物出现严重狭窄，与主要的纤维化组织的发展，越来越频繁的消化道麻烦，减少体重增加。在我们的模型中，allogeni移植Ç脂肪干细胞，双细胞片组织，增强的ESD后成功并以较低的食管狭窄率密切相关。

脂肪干细胞并获得ADSC片的过程的培养示于图1。 图2示出了移植物的结构，在其纸载体膜在彼此层叠2 ADSC片组成。脂肪干细胞以前与PKH67荧光团标记，以便与 pCLE 体内移植监控。 图3示出了延长的食管内窥镜黏膜下层剥离术的不同步骤，从而产生一个5厘米和半周食管疤痕。 图4示出的内窥镜移植过程。移植是成功的所有动物平缓应用的10分钟后。 图5示出在第3天的正pCLE评价，证实了移植过程的成功。 允许多狭窄analys随访期<html>是。不同的评估被显示在下面的附图和表格。与对照动物相比，移植的动物组显示出较不频繁的消化道麻烦和更大的增益的日28.一个动物体重从移植组相比对照组中的所有动物发展为严重的食管狭窄。相比于移植的动物，动物控制开发显著纤维化组织。 表1示出在随访期临床和内窥镜的发现。 图6示出在移植和非移植动物随访期结束时的内窥镜和放射学发现。 图7显示了移植和控制动物之间的不同组织的调查结果。这些数据证实ADSC表移植治疗食管狭窄预防的效果。 018fig1.jpg"/&gt; 图1. ADSC表建设。 A和B早通道脂肪干细胞在不同的放大倍率标准的培养皿。 ADSC张由商业温度反应培养皿培养PKH标记的脂肪干细胞获得。 C.当培养温度降低至20℃（室温）下，所有的细胞从培养皿表面作为完整的ADSC片自发地分离，而不需要酶处理。利用转移膜移植施工前的四片ADSC收获。 <a href="https://ecsource-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/55018/55018fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图2" src="/files/ftp_upload/55018/55018fig2.jpg" /> 图2.双ADSC表建设。 A，B和C. transpla前一天ntation，PKH标记脂肪干细胞收获在早期传代次数（P4 - P5），接种在12孔温度响应细胞培养，并在37℃和5％的CO 2用α-最低必需培养基中培养的，包括10％胎小牛肉血清和1％抗生素（青霉素和链霉素）。各孔涂敷有聚N-异丙基膜（在每个孔的底部）中，用1.5×10 6个细胞接种。当培养温度降低至20℃（室温）下，所有的细胞自发地脱离如无需酶处理完整ADSC片。 D. 2 ADSC片构成的双细胞表构建体的理论架构层叠在彼此并施加到纸载体膜。一个完整的双单元表结构的E.宏观视图。从参考15.修改<a href="https://ecsource-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/55018/55018fig2large.jpg" target="_blank">请点击这里Ť</a><html> Ø查看此图的放大版本。 <img alt="图3" src="/files/ftp_upload/55018/55018fig3.jpg" /> 图3.内镜黏膜下剥离过程。 答内窥镜标记创建划定切除区域，范围从40到45厘米的牙弓和涉及一半圆周。 B.含有靛蓝染料的甘油溶液的黏膜下注射。这种喷射的目的是粘膜层从肌肉层，以露出粘膜下层分离。 C.周围切口被外部执行的内窥镜标记，揭示注入粘膜下层。 D.在黏膜下层剥离术的端部的内窥镜视图，示出5厘米长和半周食管疤痕。等="_空白"&gt;点击此处查看该图的放大版本。 <img alt="图4" src="/files/ftp_upload/55018/55018fig4.jpg" /> 图4.双击ADSC表内镜移植过程。 A.四ADSC构建准备移栽。 B.在ADSC保护移植前的大帽内窥镜下施工。动物咽部的交叉，显示了内窥镜下盖保护的ADSC构造C.内镜视图。 D.应用的10分钟是足以获得片粘附。使用内窥镜帽或内镜钳进行应用。 <a href="https://ecsource-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/55018/55018fig4large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图5" srC ="/文件/ ftp_upload / 55018 / 55018fig5.jpg"/&gt; 图5. pCLE结果。在第3天pCLE示出了类似于那些在体外获得，以成功的细胞片移植兼容自发和强烈的信号。无信号于14日和28 A.悬浮脂肪干细胞的体外 pCLE信号找到。 B.在细胞片组织脂肪干细胞的体外 pCLE信号。 C. 体内 pCLE探针应用到食管疤痕。 ð。脂肪干细胞的体内 pCLE信号移植后第3天显示。从参考15.修改<a href="https://ecsource-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/55018/55018fig5large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图6" src="/files/ftp_upload/55018/55018fig6.jpg" /> 图6.结果形态分析在28天相比于对照组，内镜与第28天放射学研究结果显示短而轻狭窄而不上游扩张和完全再上皮黏膜。从参考15.修改<a href="https://ecsource-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/55018/55018fig6large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图7" src="/files/ftp_upload/55018/55018fig7.jpg" /> 图7.狭窄区域的组织学评价。后HES-标签和幻灯片数字化进行的组织学分析。 在移植的动物（A）中 ，愈合过程进行了改进，以增加再上皮和减少纤维化的发展，相比 非移植动物（B）。激烈的纤维化组织的发展是由于观察到粘膜肌层破坏，纤维变性内肌层浸润和大的粘膜缺损。从参考15.修改<a href="https://ecsource-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/55018/55018fig7large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="表格1" src="/files/ftp_upload/55018/55018table1.jpg" /> 在随访期间表1.临床评估。每天使用临床评估根据麦罗平卡斯得分和权重的变化进行临床评估。 （*）醇厚平卡斯得分16。从参考15修改。

Watch this Scientific Journal Video about ADSC-sheet Transplantation to Prevent Stricture after Extended Esophageal Endoscopic Submucosal Dissection at JoVE.com

ADSC-sheet Transplantation to Prevent Stricture after Extended Esophageal Endoscopic Submucosal Dissection

这项研究报告内窥镜脂肪组织的基质细胞（ADSC）的成功的方法折叠移植食管狭窄预防猪模型的扩展内镜黏膜下剥离术（ESD）之后。

ADSC表移植到扩展食管内镜黏膜下剥离术之后防止狭窄

In past years, the cell-sheet construct has spurred wide interest in regenerative medicine, especially for reconstructive surgery procedures. The ...

adsc-sheet-transplantation-to-prevent-stricture-after-extended

Universidad San Sebastian

Research

JoVE Journal

Medicine

8.4K 次观看.  Université Paris Descartes Sorbonne Paris Cité. 这项研究报告内窥镜脂肪组织的基质细胞（ADSC）的成功的方法折叠移植食管狭窄预防猪模型的扩展内镜黏膜下剥离术（ESD）之后。 

视频: ADSC-sheet Transplantation to Prevent Stricture after Extended Esophageal Endoscopic Submucosal Dissection

An Ivor Lewis Esophagectomy Designed to Minimize Anastomotic Complications and Optimize Conduit Function

We describe a novel esophagogastric anastomotic technique (&ldquo;side-to-side: staple line-on-staple line&rdquo;, STS) for intrathoracic anastomoses designed to create a large diameter anastomosis while simultaneously maintaining conduit blood supply. This technique aims to minimize the incidence of anastomotic leaks and strictures, which is a frequent source of morbidity and occasional mortality after esophagectomy. We analyze the results of this STS technique on 368 patients and compared outcomes to 112 patients who underwent esophagogastric anastomoses using an end-to-end stapler (EEA) over an 8-year time interval at our institution.
The STS technique involves aligning the remaining intrathoracic esophagus over the tip of the lesser curve staple line of a stomach tube, created as a replacement conduit for the esophagus. A linear stapling device cuts through and restaples the conduit staple line to the lateral wall of the esophagus in a side-to-side fashion. The open common lumen is then closed in two layers of sutures.
There was a total of 12 (3.8%) anastomotic leaks in patients who underwent STS esophagogastric anastomosis. Two of eight patients (25%) had anastomotic leaks after esophagectomy for end-stage achalasia as compared to a 2.8% leak rate (10/336) after esophagectomy for other conditions. Eighteen (5.2%) patients required a median of 2 dilatations for anastomotic stricture after STS anastomosis. Supplemental jejunostomy feedings were required in only 11.1% of patients undergoing STS anastomoses following hospital discharge. In contrast, patients undergoing EEA anastomoses demonstrated anastomotic leak and stricture rates of 16.1% and 14.3% respectively (p&lt;0.01). Time analysis of postoperative contrast studies following the STS technique typically demonstrated a straight/uniform diameter conduit with essentially complete contrast emptying into the small bowel within 3 minutes in 88.4% of patients.
The incidence of esophagogastric anastomotic leaks and strictures were extremely low using this novel anastomotic technique. Additionally we believe that based on time and qualitative analyses of postoperative contrast studies, this technique appears to optimize postoperative upper gastrointestinal tract function; however, further comparative studies are needed.

We describe a novel intrathoracic esophagogastric anastomotic technique designed to create a large diameter anastomosis while simultaneously maintaining conduit blood supply to minimize the incidence of anastomotic leaks and strictures. Construction and orientation of the stomach conduit designed to optimize upper gastrointestinal tract function is also described.

静脉造影切除术，旨在最大限度地减少麻醉并发症和优化导管功能

We describe a novel esophagogastric anastomotic technique (&ldquo;side-to-side: staple line-on-staple line&rdquo;, STS) for intrathoracic anastomoses ...

科研

癌症研究

Conditional Reprogramming of Pediatric Human Esophageal Epithelial Cells for Use in Tissue Engineering and Disease Investigation

Identifying and expanding patient-specific cells in culture for use in tissue engineering and disease investigation can be very challenging. Utilizing various types of stem cells to derive cell types of interest is often costly, time consuming and highly inefficient. Furthermore, undesired cell types must be removed prior to using this cell source, which requires another step in the process. In order to obtain enough esophageal epithelial cells to engineer the lumen of an esophageal construct or to screen therapeutic approaches for treating esophageal disease, native esophageal epithelial cells must be expanded without altering their gene expression or phenotype. Conditional reprogramming of esophageal epithelial tissue offers a promising approach to expanding patient-specific esophageal epithelial cells. Furthermore, these cells do not need to be sorted or purified and will return to a mature epithelial state after removing them from conditional reprogramming culture. This technique has been described in many cancer screening studies and allows for indefinite expansion of these cells over multiple passages. The ability to perform esophageal screening assays would help revolutionize the treatment of pediatric esophageal diseases like eosinophilic esophagitis by identifying the trigger mechanism causing the patient's symptoms. For those patients who suffer from congenital defect, disease or injury of the esophagus, this cell source could be used as a means to seed a synthetic construct for implantation to repair or replace the affected region.

Expansion of human pediatric esophageal epithelial cells utilizing conditional reprogramming provides investigators with a patient-specific population of cells that can be utilized for engineering esophageal constructs for autologous implantation to treat defects or injury and serve as a reservoir for therapeutic screening assays.

小儿食管上皮细胞重编程的条件在组织工程和疾病使用调查

Identifying and expanding patient-specific cells in culture for use in tissue engineering and disease investigation can be very challenging. Utilizing ...

发育生物学

Creation and Transplantation of an Adipose-derived Stem Cell (ASC) Sheet in a Diabetic Wound-healing Model

Artificial skin has achieved considerable therapeutic results in clinical practice. However, artificial skin treatments for wounds in diabetic patients with impeded blood flow or with large wounds might be prolonged. Cell-based therapies have appeared as a new technique for the treatment of diabetic ulcers, and cell-sheet engineering has improved the efficacy of cell transplantation. A number of reports have suggested that adipose-derived stem cells (ASCs), a type of mesenchymal stromal cell (MSC), exhibit therapeutic potential due to their relative abundance in adipose tissue and their accessibility for collection when compared to MSCs from other tissues. Therefore, ASCs appear to be a good source of stem cells for therapeutic use. In this study, ASC sheets from the epididymal adipose fat of normal Lewis rats were successfully created using temperature-responsive culture dishes and normal culture medium containing ascorbic acid. The ASC sheets were transplanted into Zucker diabetic fatty (ZDF) rats, a rat model of type 2 diabetes and obesity, that exhibit diminished wound healing. A wound was created on the posterior cranial surface, ASC sheets were transplanted into the wound, and a bilayer artificial skin was used to cover the sheets. ZDF rats that received ASC sheets had better wound healing than ZDF rats without the transplantation of ASC sheets. This approach was limited because ASC sheets are sensitive to dry conditions, requiring the maintenance of a moist wound environment. Therefore, artificial skin was used to cover the ASC sheet to prevent drying. The allogenic transplantation of ASC sheets in combination with artificial skin might also be applicable to other intractable ulcers or burns, such as those observed with peripheral arterial disease and collagen disease, and might be administered to patients who are undernourished or are using steroids. Thus, this treatment might be the first step towards improving the therapeutic options for diabetic wound healing.

Creation and Transplantation of an Adipose-derived Stem Cell ASC Sheet in a Diabetic Wound-healing Model

Adipose-derived stem cells (ASCs) are easily isolated and harvested from the fat of normal rats. ASC sheets can be created using cell-sheet engineering and can be transplanted into Zucker diabetic fatty rats exhibiting full-thickness skin defects with exposed bone and then covered with a bilayer of artificial skin.

创作和移植的糖尿病伤口愈合模型中脂肪源性干细胞 (ASC) 表

Artificial skin has achieved considerable therapeutic results in clinical practice. However, artificial skin treatments for wounds in diabetic patients ...

医学

Tissue-Engineered Graft for Circumferential Esophageal Reconstruction in Rats

The use of biocompatible materials for circumferential esophageal reconstruction is a technically challenging task in rats and requires an optimal implant technique with nutritional support. Recently, there have been many attempts at esophageal tissue engineering, but the success rate has been limited due to difficulty in early epithelization in the special environment of peristalsis. Here, we developed an artificial esophagus that can improve the regeneration of the esophageal mucosa and muscle layers through a two-layered tubular scaffold, a mesenchymal stem cell-based bioreactor system, and a bypass feeding technique with modified gastrostomy. The scaffold is made of polyurethane (PU) nanofibers in a cylindrical shape with a three-dimensional (3D) printed polycaprolactone strand wrapped around the outer wall. Prior to transplantation, human-derived mesenchymal stem cells were seeded into the lumen of the scaffold, and bioreactor cultivation was performed to enhance cellular reactivity. We improved the graft survival rate by applying surgical anastomosis and covering the implanted prosthesis with a thyroid gland flap, followed by temporary nonoral gastrostomy feeding. These grafts were able to recapitulate the findings of initial epithelialization and muscle regeneration around the implanted sites, as demonstrated by histological analysis. In addition, increased elastin fibers and neovascularization were observed in the periphery of the graft. Therefore, this model presents a potential new technique for circumferential esophageal reconstruction.

Esophageal reconstruction is a challenging procedure, and development of a tissue-engineered esophagus that enables regeneration of esophageal mucosa and muscle and that can be implanted as an artificial graft is necessary. Here, we present our protocol to generate an artificial esophagus, including scaffold manufacturing, bioreactor cultivation, and various surgical techniques.

用于大鼠圆周食管重建的组织工程移植

The use of biocompatible materials for circumferential esophageal reconstruction is a technically challenging task in rats and requires an optimal implant ...

生物工程

Transplantation of Adipose Tissue-Derived Stem Cell Sheet to Reduce Leakage After Partial Colectomy in A Rat Model

Anastomotic leakage is a disastrous complication after colorectal surgery. Although current methods for leakage prevention have different levels of clinical efficacy, they are until now imperfect solutions. Stem cell therapy using ASC sheets could provide a solution to this problem. ASCs are considered as promising candidates for promoting tissue healing because of their trophic and immunomodulatory properties. Here, we provide methods to produce high-density ASC sheets, that are transplanted onto a colorectal anastomosis in a rat model to reduce the leakage. ASCs formed cell sheets in thermo-responsive culture dishes that could be easily detached. On the day of the transplantation, a partial colectomy with a 5-suture colorectal anastomosis was performed. Animals were immediately transplanted with 1 ASC sheet per rat. ASC sheets adhered spontaneously to the anastomosis without any glue, suture, or any biomaterial. Animal groups were sacrificed 3 and 7 days postoperatively. Compared to transplanted animals, the incidence of anastomotic abscesses and leakage was higher in control animals. In our model, the transplantation of ASC sheets after colorectal anastomosis was successful and associated with a lower leakage rate.

The preparation and transplantation of adipose tissue derived stem cell (ASC) sheets onto insufficiently sutured colorectal anastomoses in a rat model is presented. This novel application shows that ASC sheets can reduce colorectal anastomosis leakage.

脂肪组织干细胞移植对大鼠部分结肠切除术后渗漏的减少

Anastomotic leakage is a disastrous complication after colorectal surgery. Although current methods for leakage prevention have different levels of ...

Vessel-sparing Excision and Primary Anastomosis

Urethroplasty is considered to be the standard treatment for urethral strictures since it provides excellent long-term success rates. For isolated short bulbar or posterior urethral strictures, urethroplasty by excision and primary anastomosis (EPA) is recommended. As EPA only requires the excision of the narrowed segment and the surrounding spongiofibrosis, a full-thickness transection of the corpus spongiosum, as performed in the traditional transecting EPA (tEPA), is usually unnecessary. Jordan et al. introduced the idea of a vessel-sparing approach in 2007, aiming to reduce surgical trauma, especially to the dual arterial blood supply of the urethra, and, thus, potentially reducing the risk of postoperative erectile dysfunction or glans ischemia. This approach could also be beneficial for subsequent urethral interventions such as redo urethroplasty using a free graft, in which a well-vascularized graft bed is imperative. Nevertheless, these potential benefits are only assumptions as prospective studies comparing the functional outcome of both techniques with validated questionnaires are currently lacking. Moreover, vessel-sparing EPA (vsEPA) should at least be able to provide similar surgical outcomes as tEPA. The aim of this paper is to give an elaborate, step-by-step overview of how to manage patients with isolated short bulbar or posterior urethral strictures with vsEPA. The main objective of this manuscript is to outline the surgical technique and to report the representative surgical outcome. A total of 117 patients were managed according to the described protocol. The analysis was performed on the entire patient cohort and on the bulbar (n = 91) and posterior (n = 26) vsEPA group separately. Success rates were 93.4% and 88.5% for the bulbar and posterior vsEPA, respectively. To conclude, vsEPA, as outlined in the protocol, provides excellent success rates with low complication rates for isolated short bulbar and posterior urethral strictures.

Here, we present an elaborate and efficient protocol to treat isolated short bulbar or posterior urethral strictures with vessel-sparing excision and primary anastomosis.

血管保留切除术与原发性吻合术

Urethroplasty is considered to be the standard treatment for urethral strictures since it provides excellent long-term success rates. For isolated short ...

Transpupillary-Guided Trans-Scleral Transplantation of Subretinal Grafts in a Retinal Degeneration Mouse Model

Transplantation of photoreceptor cells and retinal pigment&#160;epithelial (RPE) cells provide a potential therapy for retinal degeneration diseases. Subretinal transplantation of therapeutic donor cells into mouse recipients is challenging due to the limited surgical space allowed by the small volume of the mouse eye. We developed a trans-scleral surgical transplantation platform with direct transpupillary vision guidance to facilitate the subretinal delivery of exogenous cells in mouse recipients. The platform was tested using retinal cell suspensions and three-dimensional retinal sheets collected from rod-rich Rho::EGFP mice and cone-rich OPN1LW-EGFP;NRL-/- mice, respectively. Live/dead assay showed low cell mortality for both forms of donor cells. Retinal grafts were successfully delivered into the subretinal space of a mouse model of retinal degeneration, Rd1/NS, with minimum surgical complications as detected by multimodal confocal scanning laser ophthalmoscope (cSLO) imaging. Two months post-transplantation, histological staining demonstrated evidence of advanced maturation of the retinal grafts into &#39;adult&#39; rods and cones (by robust Rho::EGFP, S-opsin, and OPN1LW:EGFP expression, respectively) in the subretinal space. Here, we provide a surgical platform that can enable highly accurate subretinal delivery with a low rate of complications in mouse recipients. This technique offers precision and relative ease of skill acquisition. Furthermore, the technique could be used not only for studies of subretinal cell transplantation but also for other intraocular therapeutic studies including gene therapies.

This protocol presents a transpupillary vision-guided trans-scleral approach to safely and precisely deliver subretinal cellular grafts, with a low rate of surgical complications, in mouse recipients with or without retinal degeneration.

经瞳孔引导下视网膜下移植物的视网膜下移植物经巩膜移植视网膜变性小鼠模型

Transplantation of photoreceptor cells and retinal pigment&#160;epithelial (RPE) cells provide a potential therapy for retinal degeneration diseases. ...

神经科学

Use of the Scissor-Type Knife During the Peroral Endoscopy Myotomy Procedure for the Treatment of Achalasia

Peroral endoscopic myotomy (POEM) is one of the first-line treatment modalities along with pneumatic dilation and Heller myotomy for patients with achalasia. Endoscopists, especially trainees during the learning phase, commonly face difficulty in tissue plane dissection and selective myotomy while working near the esophagogastric junction, with increased risks of inadvertent injury, unexpected bleeding, and inadequate myotomy. To minimize the technical difficulty and improve the safety of POEM, we describe a protocol for using a scissor-type knife for the main steps of POEM, including mucosal incision, submucosal tunneling, myotomy, and hemostasis. The standard techniques used with the scissor-type knife involve grasping the target tissue, and then dissection or coagulation. The confirmation of the cutting line after grasping improves the accuracy and reliability of dissection, which is particularly useful for the selective myotomy of the internal circular muscle. Meanwhile, the scissor-type knife provides enhanced hemostatic capability and enables hemostasis and pre-coagulation without the device exchange for hemostatic forceps. Evaluation of the clinical outcomes in three patients who successfully received POEM using the scissor-type knife revealed no perioperative adverse events. At the 3-month follow-up, all patients achieved clinical success with postoperative Eckardt scores ranging from 0 to 1. In conclusion, the use of a scissor-type knife could minimize the technical difficulty and improve the safety of the POEM procedures, which may be suitable for trainees during the learning phase.

To minimize the technical difficulty and improve the safety of peroral endoscopic myotomy (POEM), we describe a protocol for using a scissor-type knife for the main steps of POEM, including mucosal incision, submucosal tunneling, myotomy, and hemostasis.

在经口内窥镜肌切开术中使用剪刀治疗贲门失弛缓症

Peroral endoscopic myotomy (POEM) is one of the first-line treatment modalities along with pneumatic dilation and Heller myotomy for patients with ...

Robotic Myotomy and Partial Fundoplication for Achalasia

Laparoscopic Heller myotomy is currently considered the standard definitive treatment of achalasia. With the advancements in technology, robotic Heller myotomy has emerged as an alternative approach to traditional laparoscopy due to three-dimensional (3D) visualization, fine motor control, and improved ergonomics provided by the robot. 
Although there is a lack of randomized controlled trials, robotic-assisted Heller myotomy seems to be associated with lower rates of intraoperative perforations compared to the laparoscopic approach. A robotic approach may also improve surgical outcomes by providing a more complete myotomy. 
Here, we describe the detailed steps of robotic myotomy and partial fundoplication for achalasia.

Surgical myotomy with a partial fundoplication may be used in selected patients as a definitive treatment for achalasia. This article provides a step-by-step description of a robotic myotomy and partial fundoplication in a 32-year-old patient with megaesophagus.

机器人肌切开术和部分胃底折叠术治疗贲门失弛缓症

Laparoscopic Heller myotomy is currently considered the standard definitive treatment of achalasia. With the advancements in technology, robotic Heller ...

Robotic D3 Partial Duodenal Resection with Primary Side-to-Side Anastomosis

Duodenal stenosis is a condition that can be related to several diseases, being either intrinsic, such as neoplasm and inflammatory stenosis, or extrinsic, such as pancreatic pseudocyst, superior mesenteric artery syndrome, and foreign bodies. Current treatments range from endoscopic approaches, such as endoscopic resection and stent placement, to surgical approaches, including duodenal resection, pancreaticoduodenectomy, and gastrointestinal bypass. Minimally invasive robot-assisted surgery is gaining importance due to its potential to decrease surgical stress, intraoperative blood loss, and postoperative pain, while its instruments and 3D-vision facilitate fine dissection and intra-abdominal suturing, all leading to a reduced time to functional recovery and shorter hospital stay. We present a case of a 75-year-old female who underwent robotic D3 partial duodenal resection with primary side-to-side duodeno-jejunal anastomosis for a 5 cm adenoma with focal high-grade dysplasia.

This protocol presents a case of a robotic partial duodenal resection with primary side-to-side duodeno-jejunal reconstruction in a patient with a 5 cm duodenal stenosis. This is done at the third duodenal segment (D3) after an endoscopic mucosal resection (EMR) for a duodenal polyp.