Khan 博士承认来自 NIH/发育研究院 K12HD052892、Alpha-1 基金会、基金会和匹兹堡肝脏研究中心的赠款支持。Michalopoulos 博士承认来自 NIH/NIDDK P01DK096990 的赠款支持。我们感谢安妮的出色和慷慨的技术援助。我们还感谢大卫. 盖勒博士提供了进入移植/啮齿动物显微外科实验室的机会。

所有程序都是按照匹兹堡大学机构动物保育和使用委员会的道德准则进行的。
1. 基本技术和一般程序
<ol><li>所有的显微外科设备使用前。</li>
	<li>为了避免麻醉杀死老鼠, 仔细检查它的呼吸模式。</li>
	<li>尽可能少接触器官和肠道, 以避免损害器官和造成出血。用湿棉签或纱布海绵按压出血点。</li>
	<li>用湿棉签对器官进行温和的操作。湿的, 无纺布纱布海绵用于两个大小, 3.5 x 3.5 厘米和 6.0 x 6.0 厘米. 用小的来收回整个肝脏。用大一点的包包裹小肠和大肠。</li>
</ol>2. 术前准备
<ol><li>使用雄性 c57/b6 小鼠, 8-12 周的年龄 (当体重约20-35 克) 的部分胆管结扎。</li>
	<li>允许老鼠免费进入水和食物, 直到麻醉的诱导。</li>
</ol>3. 部分胆总管结扎术
<ol><li>麻醉把动物放在一个感应腔内, 混合 1.5-2.0% 异氟醚和1升/分氧。观察呼吸速率, 仔细监测动物。当动物的呼吸速率为每秒一次呼吸时, 老鼠被麻醉的很好。因为前面肢体撤退反射更加迅速地消失, 捏后肢脚并且证实完全缺乏撤退反射。眼睛应与眼科润滑剂润滑, 以防止角膜溃疡。注意, 第一剂量的镇痛可以预先手术, 以防止刺激疼痛受体。</li>
	<li>用发剪刀刮腹壁, 将鼠标放在手术显微镜下的手术台上。用胶带把四肢固定在桌子上。使用异氟醚吸入在全麻下维持小鼠。<ol>
<li>调整异氟醚蒸发器, 并使用 2.0-3.0% 异氟醚诱导。</li>
	</ol>
</li>
	<li>使用无菌拭子, 先用 0.3-0.5 毫升聚维酮碘对腹壁进行消毒, 其次采用70% 乙醇。动物应该用不育的褶皱覆盖。</li>
	<li>用剪刀和 Adson 钳从剑突到耻骨进行长中线腹部切口。做腹部切口后, 将异氟醚减少到 0.8-1.0% 进行维护。</li>
	<li>使用微型牵引器暴露腹腔。用蚊子钳握住剑突, 然后向鼠标头缩回。用软粘土固定好蚊虫钳。</li>
	<li>将1-2 毫升温热 (37 °c) 消毒的磷酸盐缓冲盐水 (PBS) 注入腹腔, 以避免损伤腹部器官。用湿棉签握住小肠, 轻轻地将它们缩回鼠标的头部。减少小肠和腹腔之间的韧带, 以避免伤害小肠 (图 1A)。</li>
	<li>用湿的无纺布海绵包裹小肠和大肠。把它们放在腹腔的尾侧。肠道包裹在无菌纱布可以放置在无菌覆盖的领域, 以避免污染。</li>
	<li>用一个小的湿无纺布海绵将肝脐和肝-十二指肠韧带 (图 1B) 轻轻地收回整个肝脏, 剑突。</li>
	<li>检查以确认肝门的解剖。胆总管位于门静脉的腹侧。确认右侧和左肝胆管的位置, 将其排入肝门的主胆汁汇合处。</li>
	<li>用缝合针的钝面 (图 2A) 确定左侧肝胆管引流左叶, 并用10-0 尼龙缝线将其包围。避免损伤门静脉、肝动脉和肝表面。</li>
	<li>用10-0 尼龙结扎左胆管。(图 2B)。</li>
	<li>用湿棉拭子将小肠和大肠放在手术前的相同位置。</li>
	<li>确认没有肠道扭转和腹腔内出血, 然后关闭肌肉和皮肤两层 4-0 vicryl。编织缝合 (Vicryl) 用于皮肤闭合, 但单丝更倾向于减少感染的可能性。 注: 关闭筋膜由剑突到耻骨的持续缝合, 并通过从耻骨到剑突的持续缝合来关闭皮肤。</li>
	<li>关闭切口后停止麻醉。</li>
</ol>4. 术后治疗及随访
<ol><li>手术后, 捏住老鼠的背皮, 注射 Buprenex (0.1 毫克/千克) 和头孢唑啉 (100 毫克/千克), 其次是 Buprenex 注射液每12小时3天, 每24小时头孢唑啉分别为3天11,12 ,13。注意, 第一剂量的镇痛可以预先手术, 以防止刺激疼痛受体。头孢唑啉和其他围手术期抗生素只需要免疫抑制动物, 如用于移植研究。 注: 如果手术成功, 老鼠通常会在30分钟内痊愈, 而老鼠的活动与术前情况差不多。</li>
	<li>每天检查老鼠是否有任何遇险迹象。<ol>
<li>在常规麻醉下, 通过颈椎脱位牺牲小鼠, 在 pBDL 后, 在规定的时间点 (即, 7, 14, 21 天) 获得肝脏和其他组织样本。</li>
	</ol>
</li>
</ol>

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作者没有披露。

虽然 cBDL 是阻塞性胆汁淤积症最常见的实验技术, 但它可以在啮齿动物中提出多种挑战。根据完成终点所需的术后天数, 随着时间的推移, 动物的发病率和死亡率会显著下降。在一些转基因小鼠肝病模型的作用下, 组织损伤和胆道坏死的程度进一步恶化, 表现为肝实质异常和基线功能。与 cBDL 相比, pBDL 的报道是有限的, 但这种技术可以作为选择实验的替代品, 以减少负面结果和动物损失。
还报告了 pBDL 方法的变化。在 Heinrich et. 的方法中, 外部弯曲针放置在普通胆管旁边, 然后在普通胆管和针17周围结扎。然后在腹壁闭合前取出针头, 导致胆汁流通过狭窄的腔内 (即, 小计梗阻) 胆道17。在此过程中, 胆囊也被移除, 以防止胆囊炎17。作者认为, 这种方法更有利于研究急性胆汁淤积症, 而不是慢性变化17。相比之下, 我们在这里描述的 pBDL 方法是严密控制的, 在同一个动物中, 结扎和 unligated ("假") 裂片, 所以每个实验需要较少的动物。即使是具有严重潜在肝病的转基因小鼠, 也可以维持较长的终点 (2-3 周)。必须注意充分暴露和识别胆管树使用显微解剖技术, 有限的操作的血管结构和胆管, 以防止伤害。
在理想的实验中, 从结扎 (左) 和 unligated (右) 裂片的组织将同时收获, 因此直接比较胆汁分泌功能和转运体可以用组织学和分子技术。虽然血清肝生化不会反映严重的胆汁淤积, 结扎叶将显示组织学变化与梗阻性胆汁淤积症10。该模型有助于研究保留胆汁成分 (例如、胆汁酸、血脂、胆固醇) 在肝脏中的直接微环境效应。作为未来的研究, pBDL 可能被认为是调查 cholehepatic 分流, 其中某些胆汁酸和药物吸收直接从胆管到肝细胞18,19。肝细胞结合的内源胆汁酸被动地被胆道上皮吸收, 接受 cholehepatic 分流, 从而导致 hypercholeresis 和 alkanlinization 胆汁由增加的碳酸氢钠分泌物20。pBDL 技术的应用可以通过鉴别肝病中新的胆汁酸疗法的药理机制来促进药物的发现。
pBDL 的其他应用可以延伸到胆道树之外。cBDL 的可逆性及其相关影响已被描述, 这也可以应用到 pBDL 与内部控制21直接比较。同样, 在 cBDL22后, 研究了荧光记者在小鼠肝脏中的纵向体内bioimaging, 对 pBDL 实验有很强的影响。此外, 使用 pBDL 的啮齿类动物实验有可能将诸如骨髓间充质细胞等循环细胞的谱系追踪或 "归巢" 纳入肝损伤部位, 从而使它们能在体内被跟踪内更精度23,24,25。
显然, pBDL 作为一种内部控制和重现性的技术来研究肝脏损伤和修复, 以及胆道生理学和药物代谢, 都有广泛的应用。为了评估慢性胆汁淤积症的进展和系统性影响, cBDL 仍然是最可取的方法;然而, pBDL 仍将增加补充数据, 以支持这些发现。

外科模型的急性肝损伤和修复, 如部分肝切除或胆总管结扎 (cBDL), 已被用于研究肝再生和病理1,2。在 cBDL, 胆总管 (排泄所有肝脏产生的胆汁) 结扎, 导致梗阻性胆汁淤积, 炎症和纤维化在手术后的前2-3 周, 最终进展为肝硬化3。胆管增生, 转分化和诱导肝祖细胞也是 cBDL4,5的特征。虽然 cBDL 阐明了阻塞性 cholangiopathy 的具体机制, 其特点是在野生型小鼠中具有可重现的结果, 在这些菌株与正常肝脏6,7,8,在某些转基因小鼠肝脏疾病模型中, 由于胆道梗阻的发病率和死亡率随着时间的推移而增加, 需要的动物比预期的要多。这项技术也有利于研究具有潜在肝病的转基因小鼠, 否则可能无法容忍 cBDL 后长时间课程实验的压力。
pBDL 可以提供一个合理的替代方案来帮助克服其中的一些挑战。野生型小鼠的初步 pBDL 研究旨在区分炎症的局部作用和系统性介质9。在 pBDL 中, 胆总管上方右、左肝胆管的主胆汁汇合处仔细地可视于脐, 只结扎左肝胆管, 产生局部梗阻性胆汁淤积。pBDL 为 cBDL 提供了一些优势。在 pBDL 中, unligated 右叶作为结扎左叶的一个相同的内部控制, 受到相同的系统性影响, 如营养状况或循环因素。虽然全身性的炎症介质存在, 大泽et等) 观察到较少的纤维化和坏死的 unligated 裂片, 但增加炎症细胞和转化生长因子β表达在结扎叶 9.
最近, pBDL 被重新定义为研究保留胆汁的肝脏实质作用10。通过使用同样的动物进行内部控制 (unligated "假") 和实验组, pBDL 有效地将每项实验所需的动物数量减半, 这反过来又更具成本效益。小鼠对 pBDL 的耐受效果好得多, 在结扎叶中可实现理想的时间过程实验 (≥2周)。最重要的是, pBDL 可以区分梗阻性胆汁淤积症和保留胆汁成分的直接肝内效应, 与 unligated 控制叶相比。总的来说, pBDL 是研究肝局部胆汁淤积症叶特异效应的一种很有吸引力的方法。

<table><tbody><tr><td>Microscope-Leica Wild M650, 6&amp;ndash;40&amp;times; magnification</td><td>Leica Microsystems</td><td>n/a</td><td></td></tr><tr><td>Tec 3 isoflurane funnel fill vaporizer</td><td>General Anesthetic Services</td><td>n/a</td><td></td></tr><tr><td>Stevens Tenotomy Scissors</td><td>Accurate Surgical &amp;amp; Scientific Instruments Corporation</td><td>ASSI.9193</td><td></td></tr><tr><td>Adson Forceps</td><td>Fine science tools</td><td>11006-12</td><td></td></tr><tr><td>Mosquito classic delicate hemostatic forceps</td><td>Codman</td><td>30-4472</td><td></td></tr><tr><td>Micro-retractor</td><td>Murdock; Roboz Surgical Instrument</td><td>RS-6550</td><td></td></tr><tr><td>Nonwoven gauze sponges</td><td>Fisherband</td><td>870-PC-DBL</td><td></td></tr><tr><td>Puritan 6&amp;rdquo; Small Cotton Swab w/Wooden Handle</td><td>Puritan Medical Products Company</td><td>868-WCS</td><td></td></tr><tr><td>Dumont SS Forceps - Standard Tips/Straight/Inox/13.5cm</td><td>Fine science tools</td><td>11203-23</td><td></td></tr><tr><td>Dumont SS Forceps - Standard Tips/Angled 45&amp;deg;/Inox/13.5cm</td><td>Fine science tools</td><td>11203-25</td><td></td></tr><tr><td>Vannas Spring Scissors - 3mm Cutting Edge</td><td>Fine science tools</td><td>15000-00</td><td></td></tr><tr><td>Microneedle holder</td><td>Aesculap Surgical Instruments</td><td>FD231R</td><td></td></tr><tr><td>Micro AROSuture, sterile 10-0 nylon suture, 70 &amp;micro;m, TAP points</td><td>AROSurgical Instruments Corporation</td><td>T4A10N07</td><td></td></tr><tr><td>4-0 Vicryl</td><td>Ethicon</td><td>J662H</td><td></td></tr><tr><td>5-ml Syringe</td><td>BD</td><td>309646</td><td></td></tr><tr><td>Falcon tissue culture dish, 60 &amp;times; 15 mm</td><td>Corning</td><td>353002</td><td></td></tr><tr><td>Isoflurane, United States Pharmacopeia (USP) liquid for inhalation, 250 ml</td><td>Piramal Healthcare</td><td>NDC 66794-017-25</td><td></td></tr><tr><td>Buprenex injectable (buprenorphine hydrochloride)</td><td>Reckitt Benckiser Pharmaceuticals</td><td>NDC 12496-0757-1</td><td></td></tr><tr><td>Cefazolin for injection , USP</td><td>APP Pharmaceuticals</td><td>NDC 63323-238-61</td><td></td></tr><tr><td>PVP scrub solution (povidone&amp;ndash;iodine 7.5%)</td><td>Medline</td><td>NDC 12496-0757-1</td><td></td></tr><tr><td>70% Ethanol</td><td>Decon Labs</td><td>2716</td><td></td></tr><tr><td>Phosphate Buffered Saline Without Calcium or Magnesium</td><td>LONZA</td><td>17-516F</td><td></td></tr><tr><td>Sirius Red</td><td>Sigma</td><td>365548</td><td></td></tr><tr><td>Hematolxylin</td><td>Fisher (Richard-Allan Scientific)</td><td>22-050-111 (7211)</td><td></td></tr><tr><td>Eosin</td><td>Anatech (Fisher)</td><td>832 (NC9686037)</td><td></td></tr><tr><td>Formalin</td><td>Fisher</td><td>SF100-20</td><td></td></tr></tbody></table>

partial bile duct ligation in the mouse: a controlled model of localized obstructive cholestasis

在啮齿动物中, 胆总管完全胆管结扎术 (cBDL) 是研究梗阻性胆汁淤积症和胆管增生的一种成熟的外科技术。然而, 长期实验可能导致发病率和死亡率的增加。在选择有潜在肝病的小鼠菌株中, 甚至可以通过结扎肝单叶来进行有意义的比较, 这样可以减少动物的损失和开支。在这里, 我们描述的部分胆管结扎 (pBDL) 在小鼠, 其中只有左肝胆管结扎, 导致胆道梗阻的左叶, 而不是其余的裂片。通过仔细的显微外科技术, pBDL 实验可以是成本效益, 因为 unligated 叶作为一个内部控制的结扎裂片, 当受到相同的条件在同一动物。不像 cBDL, 一个单独的假操作控制组是不必要的。pBDL 是非常有用的直接比较局部和系统性的影响胆汁淤积症和其他保留胆汁成分。pBDL 也可以作为一种新的方法来研究与药物和细胞迁移有关的机制。

对于这一典型的实验, pBDL 在三 PiZ 转基因小鼠中进行, 概括了 alpha-1 胰岛素 (A1AT) 缺乏中发现的人肝病。PiZ 小鼠 overexpresses 一种转基因, 由 DNA 的多个基因组片段组成, 其中包含突变体的人类 A1AT 基因和启动子的编码区域, 以及上游和下游侧翼区域的 2 kilobases14, 15。正常 A1AT 是一种分泌的血清蛋白在肝脏中产生。A1AT 的突变体产生不溶性的蛋白质聚集物, 自发地积聚在肝脏中引起毒性压力。实验利用3月大的雄性 PiZ 小鼠, 表达了这种疾病最严重的表型, 当蛋白质聚集峰值16。
血清肝生化比较 pBDL 和 cBDL PiZ 小鼠 (表 1)10。肝酶丙氨酸转氨酶 (ALT) 和天门冬氨酸转氨酶 (AST) 是肝细胞损伤的生物标志物, 而总血清胆红素 () 测定胆汁淤积程度。总的来说, 受 pBDL 的小鼠没有出现严重的肝损伤或胆汁淤积的迹象, 因为他们的血清值与基线10相似。重要的是要注意的是, pBDL 失败的老鼠 (由于技术问题, 如胆管或血管损伤) 将显示与 cBDL 小鼠相似的血清价值, 显著升高的肝酶和血药。临床上, pBDL 失败的小鼠会出现更多的黄疸和苦恼, 发现胆汁泄漏, 腹水, 腹膜炎和/或血肿, 最终将需要牺牲。
由于受 pBDL 的小鼠发育出叶特定的梗阻性胆汁淤积症, 他们没有获得在 cBDL 中看到的扩张, 黑暗, 胆汁填充胆囊。只有结扎的裂片的颜色显得苍白, 而 unligated 裂片将保持其正常的, 深色的, 红褐色的着色 (图 3)。肝组织学也存在着重要的差异, 在结扎的肺叶中胆管增生和纤维化增加 (图 4)。10
<img alt="Table 1" src="/files/ftp_upload/56930/56930table1.jpg"> 表 1:代表血清肝生化从部分胆管结扎术 (pBDL) 与完全胆管结扎术 (cBDL) 比较)10.每个时间点代表3-4 只老鼠。注意 PiZ 转基因小鼠的 c57/bl6 背景。实验室参考范围也在括号中给出。(野生型;总血清胆红素;ALT, 丙氨酸转氨酶;碱性磷酸酶)。
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/56930/56930fig1.jpg"> 图 1:腹腔和肝胆系统暴露.(A) 小肠被轻轻地缩回以避免肠道损伤, 连接韧带从腹腔中被切断。(B) 肝脏被轻轻地收回以允许肝脏脐 (箭头) 上的胆管树的最大可视化。<a href="//cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/56930/56930fig1large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/56930/56930fig2.jpg"> 图 2: 部分胆总管结扎.(A) 手术针的钝端用于仔细地将尼龙缝合线环绕在左肝胆管周围, 与胆总管的汇合处 (原放大 40x)。(B) 成功地结扎左肝胆管, 通常排泄左肝叶。其余的胆导管是 unligated (原放大 40X)。<a href="//cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/56930/56930fig2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/56930/56930fig3.jpg"> 图 3: pBDL 后14天 (c57/b6) 小鼠肝脏的总外观.请注意, 结扎左叶的颜色显得苍白 (箭头), 而 unligated 裂片保留其正常, 暗, 红褐色的着色。<a href="//cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/56930/56930fig3large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/56930/56930fig4.jpg"> 图 4: 从14天 pBDL 比较 unligated 右 (R) 叶与结扎左 (L) 叶 (c57/b6) 小鼠的代表性肝组织切片中的叶特异变化.(A) 苏木精和伊红 (H & E) 染色显示结扎叶中胆管增生增加 (箭头)。(B) 天狼星红染色显示在结扎叶中伴有胆管增生的桥状纤维化。PiZ 鼠标的组织学以前已报告为10。<a href="//cloudflare-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/56930/56930fig4large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>

Watch this Scientific Journal Video about Partial Bile Duct Ligation in the Mouse: A Controlled Model of Localized Obstructive Cholestasis at JoVE.com

Partial Bile Duct Ligation in the Mouse: A Controlled Model of Localized Obstructive Cholestasis

在这里, 我们提出了部分胆管结扎作为一个外科模型的肝损伤和再生的啮齿动物。

小鼠部分胆管结扎术: 局部梗阻性胆汁淤积症的对照模型

In rodents, complete bile duct ligation (cBDL) of the common bile duct is an established surgical technique for studying obstructive cholestasis and bile ...

partial-bile-duct-ligation-mouse-controlled-model-localized

Nanyang Technological University

Research

JoVE Journal

Biology

15.4K 次观看.  Jichi Medical University. 在这里, 我们提出了部分胆管结扎作为一个外科模型的肝损伤和再生的啮齿动物。

视频: Partial Bile Duct Ligation in the Mouse: A Controlled Model of Localized Obstructive Cholestasis

Creation of Reversible Cholestatic Rat Model

Cholestasis is a clinical condition commonly encountered by both surgeons and gastroenterologists. Cholestasis can cause various physiological changes and affect the nutritional status and surgical outcomes. Study of the pathophysiological changes occurring in the liver and other organs is of importance. Various studies have been done in cholestatic rat models.
We used a reversible cholestatic rat model in our recent study looking at the role of methylprednisolone in the ischemia reperfusion injury. Various techniques for creation of a reversible cholestatic model have been described. Creation of a reversible cholestatic rat model can be challenging in view of the smaller size and unique hepatopancreatobiliary anatomy in rats. This video article demonstrates the creation of a reversible cholestatic model.
This model can be used in various studies, such as looking at the changes in nutritional, physiological, pathological, histological and immunological changes in the gastrointestinal tract. This model can also be used to see the effects of cholestasis and various therapeutic interventions on major hepatic surgeries.

Cholestasis is a clinical condition commonly encountered by both surgeons and gastroenterologists. Creation of a reversible cholestatic rat model can be challenging in view of the smaller size and unique hepatopancreatobiliary anatomy in rats. This video article demonstrates the creation of a reversible cholestatic model.

可逆的胆汁淤积大鼠模型的建立

Cholestasis is a clinical condition commonly encountered by both surgeons and gastroenterologists. Cholestasis can cause various physiological changes and ...

科研

医学

Determining Bile Duct Density in the Mouse Liver

Mouse is broadly used as a model organism to study biliary diseases. To evaluate the development and function of the biliary system, various techniques are used, including serum chemistry, histological analysis, and immunostaining for specific markers. Although these techniques can provide important information about the biliary system, they often do not present a full picture of bile duct (BD) developmental defects across the whole liver. This is in part due to the robust ability of the mouse liver to drain the bile even in animals with significant impairment in biliary development. Here we present a simple method to calculate the average number of BDs associated with each portal vein (PV) in sections covering all lobes of mutant/transgenic mice. In this method, livers are mounted and sectioned in a stereotypic manner to facilitate comparison among various genotypes and experimental conditions. BDs are identified via light microscopy of cytokeratin-stained cholangiocytes, and then counted and divided by the total number of PVs present in liver section. As an example, we show how this method can clearly distinguish between wild-type mice and a mouse model of Alagille syndrome. The method presented here cannot substitute for techniques that visualize the three-dimensional structure of the biliary tree. However, it offers an easy and direct way to quantitatively assess BD development and the degree of ductular reaction formation in mice.

We present a rather simple and sensitive method for accurate quantification of bile duct density in the mouse liver. This method can aid in determining the effects of genetic and environmental modifiers and the effectiveness of potential therapies in mouse models of biliary diseases.

确定小鼠肝脏中的胆汁密度

Mouse is broadly used as a model organism to study biliary diseases. To evaluate the development and function of the biliary system, various techniques ...

发育生物学

Bile Duct Ligation in Mice: Induction of Inflammatory Liver Injury and Fibrosis by Obstructive Cholestasis

In most vertebrates, the liver produces bile that is necessary to emulsify absorbed fats and enable the digestion of lipids in the small intestine as well as to excrete bilirubin and other metabolic products. In the liver, the experimental obstruction of the extrahepatic biliary system initiates a complex cascade of pathological events that leads to cholestasis and inflammation resulting in a strong fibrotic reaction originating from the periportal fields. Therefore, surgical ligation of the common bile duct has become the most commonly used model to induce obstructive cholestatic injury in rodents and to study the molecular and cellular events that underlie these pathophysiological mechanisms induced by inappropriate bile flow. In recent years, different surgical techniques have been described that either allow reconnection or reanastomosis after bile duct ligation (BDL), e.g., partial BDL, or other microsurgical methods for specific research questions. However, the most frequently used model is the complete obstruction of the common bile duct that induces a strong fibrotic response after 21 to 28 days. The mortality rate can be high due to infectious complications or technical inaccuracies. Here we provide a detailed surgical procedure for the BDL model in mice that induce a highly reproducible fibrotic response in accordance to the 3R rule for animal welfare postulated by Russel and Burch in 1959.

Disruption of bile flow results in severe inflammatory cholestatic liver injury with a characteristic time-dependent sequence of morphological alterations. Here we present a protocol for the surgical ligation of the common bile duct in mice that allows to induce a strong fibrotic response after 21 to 28 days.

胆管结扎小鼠:感应炎性肝损伤和纤维化的阻塞性胆汁淤积

In most vertebrates, the liver produces bile that is necessary to emulsify absorbed fats and enable the digestion of lipids in the small intestine as well ...

A Silver Nanoparticle Method for Ameliorating Biliary Atresia Syndrome in Mice

Biliary atresia (BA) is a severe type of cholangitis with high mortality in children of which the etiology is still not fully understood. Viral infections may be one possible cause. The typical animal model used for studying BA is established by inoculating a neonatal mouse with a rhesus rotavirus. Silver nanoparticles have been shown to exert antibacterial and antiviral effects; their function in the BA mouse model is evaluated in this study. Currently, in BA animal experiments, the methods used to improve the symptoms of BA mice are generally symptomatic treatments given via food or other drugs. The aim of this study is to demonstrate a new method for ameliorating BA syndrome in mice by the intraperitoneal injection of silver nanoparticles and to provide detailed methods for preparing the silver nanoparticle gel formulation. This method is simple and widely applicable and can be used to research the mechanism of BA, as well as in clinical treatments. Based on the BA mouse model, when the mice exhibit jaundice, the prepared silver nanoparticle gel is injected intraperitoneally to the surface of the lower liver. The survival status is observed, and biochemical indicators and liver histopathology are examined. This method allows a more intuitive understanding of both the establishment of the BA model and novel BA treatments.

This article describes in detail a method based on silver nanoparticles for ameliorating biliary atresia syndrome in an experimental biliary atresia mouse model. A solid understanding of the reagent preparation process and the neonatal mouse injection technique will help familiarize researchers with the method used in neonatal mouse model studies.

银纳米粒子法改善小鼠胆道闭锁综合征

Biliary atresia (BA) is a severe type of cholangitis with high mortality in children of which the etiology is still not fully understood. Viral infections ...

免疫与感染

Ileectomy-induced Bile Overaccumulation in Mouse Intestine

Intestinal resection is a common therapeutic approach for human diseases such as obesity, inflammatory bowel disease, Crohn's disease, and colon cancer that often results in severe short bowel syndrome-like adverse effects including bile acid diarrhea, dehydration, electrolyte disturbances, and nutrient malabsorption. Here we introduce a murine ileal resection model, termed ileectomy, to evaluate tissue communication and the maintenance of systemic homeostasis. After ileal resection, circulating blood is permanently devoid of the ileum-specific endocrine hormone fibroblast growth factor 15 (FGF15), which releases its endocrinal inhibition of bile acid synthesis in the liver. In combination with the increased production and abolished reabsorption of bile acids after removing the ileum, mice that underwent surgery suffer from bile salt overaccumulation in the intestine and associated diarrhea, morbidity, and mortality. Novel usage of the surgery model introduced in this study may provide mechanistic and functional insights into ileal control of systemic metabolic regulation in physiology and disease.

Small intestine-dependent bile acid reabsorption and feedback inhibition of hepatic bile acid synthesis is important for systemic homeostasis and health. In this study, we describe a mouse model for ileal resection to evaluate ileectomy-induced bile malabsorption, overaccumulation, and toxicity in mouse intestine.

回肠致胆汁积滞在小鼠肠道中

Intestinal resection is a common therapeutic approach for human diseases such as obesity, inflammatory bowel disease, Crohn's disease, and colon cancer ...

A Mouse Model of Chronic Liver Fibrosis for the Study of Biliary Atresia

Biliary atresia (BA) is a fatal disease involving obstructive jaundice, and it is the most common indication for liver transplantation in children. Due to the complex etiology and unknown pathogenesis, there are still no effective drug treatments. At present, the classic BA mouse model induced by rhesus rotavirus (RRV) is the most commonly used model for studying the pathogenesis of BA. This model is characterized by growth retardation, jaundice of the skin and mucosa, clay stools, and dark yellow urine. The histopathology shows severe liver inflammation and obstruction of the intrahepatic and extrahepatic bile ducts, which are similar to the symptoms of human BA. However, the livers of end-stage mice in this model lack fibrosis and cannot fully simulate the characteristics of liver fibrosis in clinical BA. The presented study developed a novel BA mouse model of chronic liver fibrosis by injecting 5-10 &#181;g of anti-Ly6G antibody four times, with gaps of 2 days after each injection. The results showed that some of the mice successfully formed chronic BA with typical fibrosis after the time lapse, meaning these mice represent a suitable animal model for the virus-induced liver fibrosis mechanistic study of BA and a platform for developing future BA treatments.

We established a mouse model of chronic liver fibrosis, which provides a suitable animal model for the virus-induced liver fibrosis mechanistic study of biliary atresia (BA) and a platform for future BA treatments.

慢性肝纤维化小鼠模型用于研究胆道闭锁

Biliary atresia (BA) is a fatal disease involving obstructive jaundice, and it is the most common indication for liver transplantation in children. Due to ...

A Mouse Model for Chronic Pancreatitis via Bile Duct TNBS Infusion

Chronic pancreatitis (CP) is a complex disease involving pancreatic inflammation and fibrosis, glandular atrophy, abdominal pain and other symptoms. Several rodent models have been developed to study CP, of which the bile duct 2,4,6 -trinitrobenzene sulfonic acid (TNBS) infusion model replicates the features of neuropathic pain seen in CP. However, bile duct drug infusion in mice is technically challenging. This protocol demonstrates the procedure of bile duct TNBS infusion for generation of a CP mouse model. TNBS was infused into the pancreas through the ampulla of Vater in the duodenum. This protocol optimized drug volume, surgical techniques, and drug handling during the procedure. TNBS-treated mice showed features of CP as reflected by bodyweight and pancreas weight reductions, changes in pain-associated behaviors, and abnormal pancreatic morphology. With these improvements, mortality associated with TNBS injection was minimal. This procedure is not only critical in generating pancreatic disease models but is also useful in local pancreatic drug delivery.

Chronic pancreatitis (CP) is a disease characterized by inflammation and fibrosis of the pancreas, often associated with intractable abdominal pain. This article focuses on refining the technique to generate a mouse model of CP via bile duct infusion with 2,4,6 -trinitrobenzene sulfonic acid (TNBS).

通过胆管 TNBS 输液治疗慢性胰腺炎的鼠标模型

Chronic pancreatitis (CP) is a complex disease involving pancreatic inflammation and fibrosis, glandular atrophy, abdominal pain and other symptoms. ...

Extrahepatic Bile Duct and Gall Bladder Dissection in Nine-Day-Old Mouse Neonates

The dissection of murine neonatal bile ducts has been described as difficult. The main aim of the described standard operating procedure is the isolation of the extrahepatic bile duct (EBD) in mouse neonates without damaging the bile duct during preparation. Because of its exceptionally close preparation compared to the cholangiocytes cell line and harvesting of the entire extrahepatic bile duct system (EBDS), the described approach is extremely useful in researching animal models of newborn bile duct disorders, such as biliary atresia. After euthanasia, the peritoneal cavity was accessed, and the bile duct system, duodenum, and liver were extracted with the unique En-bloc-Resection (EbR). The extracted sample is placed on a foam mat, and the EBD is dissected from contaminating cells atraumatically without necessary touch. The dissection of the entire EBDS is a significant advantage of this method. Caution must be taken due to the small size and amount of bile duct tissue. Using the described technique, there is no damage to the cholangiocytes. Further, the purity of the technique is reproducible (n = 10). Therefore, optimally comparable samples can be harvested. Furthermore, no bile duct tissue is harmed, because any contact with the bile duct system can be avoided during preparation, leaving the bile fluid inside the gall bladder. Most importantly, while performing the final gall bladder and bile duct dissection, atraumatic microinstruments were used only slightly lateral of the bile duct without squeezing it. This is the key to a clean and intact sample, and essential for further histological investigation or the isolation of cholangiocytes. To summarize, the described innovative dissection technique enables especially inexperienced operators with the necessary equipment to isolate the EBDS as cleanly as possible.

For the observation of murine neonatal bile duct disorders, an intact bile duct and efficient preparation are required. Therefore, a new approach for isolating the entire extrahepatic bile duct system in murine neonates was successfully developed while maintaining the integrity of the bile duct.

9日龄小鼠新生儿肝外胆管和胆囊清扫术

The dissection of murine neonatal bile ducts has been described as difficult. The main aim of the described standard operating procedure is the isolation ...

<meta charset="utf8"></head><body>在分期肝切除术动物模型中研究 ALLPS 的肝脏再生

A Mouse Model of the Associating Liver Partition and Portal Vein Ligation for Staged Hepatectomy Procedure Aided by Microscopy

Hepatectomy is widely regarded as the primary treatment for hepatic malignancies; yet, postoperative liver failure remains a major cause of perioperative mortality, severely impacting patient outcomes. In a robust hepatic environment, the future liver remnant (FLR) must exceed 25%, and in cases of cirrhosis, this requirement increases to over 40%. The inadequacy of FLR is currently a major obstacle in the progression of hepatic surgery.
Traditional methods to enhance FLR hypertrophy mainly focus on portal vein embolization (PVE), but its effectiveness is considerably limited. In recent years, there have been numerous reports on a novel biphasic hepatectomy method involving hepatic partitioning and portal vein ligation, known as associating liver partition and portal vein ligation for staged hepatectomy (ALPPS). ALPPS surpasses PVE in efficiently and considerably inducing FLR hypertrophy. However, the detailed mechanisms driving ALPPS-facilitated hepatic regeneration are not fully understood. Thus, replicating ALPPS in animal models is crucial to thoroughly investigate the molecular mechanisms of hepatic regeneration, offering valuable theoretical and practical insights.

<meta charset="utf8"></head><body>作者聚焦：通过 ALPPS 小鼠模型推进肝脏再生研究

Associating Liver Partition and Portal Vein Ligation for Staged Hepatectomy (ALPPS) has been shown to result in remarkable liver regeneration after several weeks post first-stage surgery. Animal models that have undergone ALPPS have been developed to explore potential liver regeneration ability and achieve favorable clinical outcomes.

显微镜辅助分期肝切除术的关联肝分区和门静脉结扎的小鼠模型

Hepatectomy is widely regarded as the primary treatment for hepatic malignancies; yet, postoperative liver failure remains a major cause of perioperative ...

小鼠部分胆管结扎术: 局部梗阻性胆汁淤积症的对照模型

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9日龄小鼠新生儿肝外胆管和胆囊清扫术

显微镜辅助分期肝切除术的关联肝分区和门静脉结扎的小鼠模型

显微镜辅助分期肝切除术的关联肝分区和门静脉结扎的小鼠模型