小鼠部分胆管结扎术: 局部梗阻性胆汁淤积症的对照模型
March 28th, 2018
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。
这种手术干预的总体目标是观察局部阻塞性胆汁淤积的效果。这种方法可以帮助回答肝病学领域关于如何识别肝脏再生机制的关键问题。该技术的主要优点是可以在同一动物体内连接的实验叶和未连接的对照叶之间进行有意义的比较。
今天演示该程序的是移植手术实验室的博士后 Toshimasa Nakao 博士。在确认麻醉的 8 至 12 周龄 C57 black 6 小鼠对脚趾捏无反应后，用理发剪剃掉腹部区域，并将动物置于手术显微镜下。将肢体固定在桌子上，并使用无菌拭子依次用 0.3 至 0.5 毫升聚维酮碘和 70% 乙醇对手术区域进行消毒。
接下来，从剑突到耻骨做一个中线切口，并使用微型牵开器打开腹腔。使用蚊子镊子将剑突缩向小鼠头部，并使用软粘土将镊子和剑突保持在该位置。用 1 到 2 毫升消毒的 37 摄氏度 PBS 滋润腹腔，并使用湿棉签轻轻地将小肠移向小鼠头部。
剪断小肠和腹膜后之间的韧带，以免损伤组织，并用湿的无纺布纱布海绵包裹小肠和大肠。将肠组织放在腹腔尾侧，用小的湿无纺布纱布海绵轻轻地将整个肝脏向剑突缩回，露出肝门和肝十二指肠韧带。找到肝门、门静脉腹侧的胆总管以及排入肝门处主要胆道汇合处的左右肝胆管。
使用 10-0 尼龙缝合线和缝合针的钝侧，结扎左肝胆管叶。缝合后，将小肠和大肠放回其解剖位置，并确认没有肠扭转或腹腔内出血。然后取下蚊子镊子，分别用连续 4-0 薇乔线缝合肌肉和皮肤层，并将鼠标置于监测的加热灯下，直至完全恢复。
部分胆管结扎小鼠和完全胆管结扎小鼠转基因不溶性肝蛋白聚集的血清肝脏生化比较显示，部分胆管结扎小鼠的肝损伤或胆汁淤积最小，而完全胆管结扎的转基因动物则严重损伤和酶积聚。随着接受部分胆管结扎的小鼠随着时间的推移发展为叶特异性阻塞性胆汁淤积，与保持正常深红棕色的未结扎的叶相比，结扎的叶在手术后 14 天的颜色略浅。与未结扎的肺叶相比，结扎的肺叶还表现出胆管增殖和纤维化的增加，以响应结扎诱导的局部胆汁淤积。
一旦掌握，如果执行得当，这项技术可以在 10 到 12 分钟内完成。在此程序之后，可以使用谱系追踪等其他方法来回答有关干细胞归巢到肝损伤和再生部位的其他问题。观看此视频后，您应该对如何使用部分胆管结扎术进行有意义的比较以识别局部影响与全身影响有很好的了解。
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概述
本文介绍了部分胆管结扎作为研究啮齿动物肝脏损伤和再生的一种外科模型。该方法允许观察局部阻塞性胆汁淤积及其对肝脏机制的影响。
关键研究组成部分
科学领域
- 肝病学
- 肝脏再生
- 外科模型
背景
- 了解肝脏损伤和再生对肝病学至关重要。
- 局部阻塞性胆汁淤积可以提供对肝脏机制的洞察。
- 在同一动物内进行比较研究可以提高结果的可靠性。
- 这个模型特别适用于研究肝脏恢复的动力学。
研究目的
- 观察部分胆管结扎对肝脏损伤的影响。
- 识别肝脏再生涉及的机制。
- 建立一个可靠的外科模型以用于未来的肝病学研究。
使用的方法
- 使用麻醉的C57黑色6号小鼠进行手术。
- 在手术前剃毛并对腹部进行消毒。
- 将四肢固定在手术台上以保持稳定。
- 在手术显微镜下进行手术以确保精确。
主要结果
- 证明了该外科模型的可行性。
- 建立了比较结扎和未结扎肝叶的方法。
- 提供了对肝脏再生过程的洞察。
- 促进了对肝脏损伤机制的进一步研究。
结论
- 部分胆管结扎是研究肝脏损伤的有价值模型。
- 该技术允许在同一生物体内进行受控的比较。
- 未来的研究可以在此模型的基础上进一步探索肝脏再生。
常见问题
Executive Industry Relevance
Partial bile duct ligation (pBDL) provides a controlled model for studying localized obstructive cholestasis, enabling direct comparison of ligated and unligated liver lobes within the same animal. This internal control design reduces variability, lowers animal use, and supports mechanistic de-risking in hepatology target validation. The approach enhances predictive confidence by isolating lobe-specific responses to bile accumulation and fibrosis pathways.
Strategic Applications in Biopharma R&D
Early Discovery & Target Validation
- Scientific Value: Enables interrogation of therapeutic hypotheses related to localized liver injury and regeneration pathways.
- Operational Value: Supports biological de-risking by comparing lobe-specific responses to cholestasis within identical genetic and environmental backgrounds.
- Predictive Value: Facilitates portfolio triage by distinguishing systemic versus localized drug effects on bile duct proliferation and fibrosis.
Screening & Assay Development
- Scientific Value: Prepares validated disease-relevant systems for downstream assay standardization using lobe-specific biochemical readouts.
- Operational Value: Enhances reproducibility through internal controls, reducing need for separate sham groups and minimizing inter-animal variability.
- Scalability: Supports platform reuse in preclinical models investigating medication mechanisms and cell migration in cholestatic contexts.
Translational & Preclinical Research
- Scientific Value: Aligns with disease-relevant systems by modeling human obstructive cholestasis with lobe-specific bile duct proliferation and fibrosis.
- Translational Continuity: Bridges discovery to preclinical validation by enabling lineage tracing and stem cell homing studies post-ligation.
- Risk-Adjusted Advancement: Supports go/no-go decisions by quantifying localized versus systemic biomarker responses to retained bile components.
Pipeline & Workflow Integration
pBDL fits within the discovery continuum from target validation through preclinical research, particularly for hepatology-focused programs investigating liver injury mechanisms and regenerative responses.
- Discovery Biology: Supports hypothesis testing by enabling comparison of ligated and unligated lobes to isolate local effects of cholestasis.
- Screening: Delivers assay-ready biological systems with quantifiable outputs such as serum liver biochemistries and histological changes in bile duct proliferation.
- Analytics: Provides quantitative dependent variable measurements including enzyme levels, lobe coloration, and fibrosis scores to compare experimental conditions.
- Translational Research: Connects to preclinical continuity by supporting post-surgical analyses like lineage tracing to study stem cell recruitment to injury sites.
- Enterprise Reuse: Functions as a reusable surgical platform for studying medication effects and cellular mechanisms in localized liver disease models.
Operational & Enterprise Impact
- Scientific Value: Increases predictive confidence by reducing mechanistic ambiguity through internal lobe-to-lobe comparisons.
- Operational Value: Improves standardization and reproducibility by eliminating inter-animal variability via within-animal controls.
- Strategic Value: Enhances capital efficiency by reducing animal losses and expenses compared to complete bile duct ligation models.
- Portfolio Impact: Enables risk-adjusted prioritization by distinguishing localized drug toxicity from systemic liver effects.
Implementation Considerations
- Requires microsurgical expertise and access to surgical microscopes for precise ligation of the left hepatic bile duct.
- Depends on instrumentation including 10-0 nylon sutures, mosquito forceps, and sterile field maintenance tools.
- Necessitates cross-team standardization of postoperative monitoring and serum biochemistry sampling procedures.
- Involves adaptation considerations when extending the model to other mouse strains or investigating different lobes.
- Limited to studies where lobe-specific obstruction is sufficient to model the human condition of interest.
