Name: the murine choline-deficient, ethionine-supplemented (cde) diet model of chronic liver injury
Uploaded: 2017-10-21T14:00:00
Description: Watch this Scientific Journal Video about The Murine Choline-Deficient, Ethionine-Supplemented CDE Diet Model of Chronic Liver Injury at JoVE.com

这项工作得到了澳大利亚国家卫生和医学研究理事会 (APP1042370、APP1061332、APP1087125) 澳洲的资助。作者希望感谢科科卫生创新研究所的工作人员提供技术援助。

1. 动物实验 在本调查中描述的所有动物研究都是由科大学动物伦理委员会 (批准编号: AEC_2014_28) 在开始之前批准的根据《澳大利亚动物保育和使用守则》进行实验和执行. 
<ol> <li> 动物 <ol> <li> 使用六周的老雄性 C57BL/6J 小鼠进行实验. </li>
	</ol> </li> <li> 实验设计 <ol> 到达动物设施后, 允许小鼠在任何实验开始前适应四天. <li>
		<li> 在小麦谷壳床上?...

<ol>
	<li>Taub, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Liver+regeneration:+from+myth+to+mechanism.">Liver regeneration: from myth to mechanism.</a> Nat Rev Mol Cell Biol. 5, 836-847 (2004).</li><li>Kohn-Gaone, J., Gogoi-Tiwari, J., Ramm, G. A., Olynyk, J. K., Tirnitz-Parker, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+liver+progenitor+cells+during+liver+regeneration,+fibrogenesis,+and+carcinogenesis.">The role of liver progenitor cells during liver regeneration, fibrogenesis, and carcinogenesis.</a> Am J Physiol Gastrointest Liver Physiol. 310, G143-G154 (2016).</li><li>Lu, W. Y., 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=Hepatic+progenitor+cells+of+biliary+origin+with+liver+repopulation+capacity.">Hepatic progenitor cells of biliary origin with liver repopulation capacity.</a> Nat Cell Biol. 17, 971-983 (2015).</li><li>Prakoso, E., 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=Analysis+of+the+intrahepatic+ductular+reaction+and+progenitor+cell+responses+in+hepatitis+C+virus+recurrence+after+liver+transplantation.">Analysis of the intrahepatic ductular reaction and progenitor cell responses in hepatitis C virus recurrence after liver transplantation.</a> Liver Transpl. 20, 1508-1519 (2014).</li><li>Tirnitz-Parker, J. E., Tonkin, J. N., Knight, B., Olynyk, J. K., Yeoh, G. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation,+culture+and+immortalisation+of+hepatic+oval+cells+from+adult+mice+fed+a+choline-deficient,+ethionine-supplemented+diet.">Isolation, culture and immortalisation of hepatic oval cells from adult mice fed a choline-deficient, ethionine-supplemented diet.</a> Int J Biochem Cell Biol. 39, 2226-2239 (2007).</li><li>Van Hul, N. K., Abarca-Quinones, J., Sempoux, C., Horsmans, Y., Leclercq, I. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Relation+between+liver+progenitor+cell+expansion+and+extracellular+matrix+deposition+in+a+CDE-induced+murine+model+of+chronic+liver+injury.">Relation between liver progenitor cell expansion and extracellular matrix deposition in a CDE-induced murine model of chronic liver injury.</a> Hepatology. 49, 1625-1635 (2009).</li><li>Clouston, A. 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=Fibrosis+correlates+with+a+ductular+reaction+in+hepatitis+C:+roles+of+impaired+replication,+progenitor+cells+and+steatosis.">Fibrosis correlates with a ductular reaction in hepatitis C: roles of impaired replication, progenitor cells and steatosis.</a> Hepatology. 41, 809-818 (2005).</li><li>Forbes, S. J., Newsome, P. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Liver+regeneration+-+mechanisms+and+models+to+clinical+application.">Liver regeneration - mechanisms and models to clinical application.</a> Nat Rev Gastroenterol Hepatol. 13, 473-485 (2016).</li><li>Akhurst, 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+modified+choline-deficient,+ethionine-supplemented+diet+protocol+effectively+induces+oval+cells+in+mouse+liver.">A modified choline-deficient, ethionine-supplemented diet protocol effectively induces oval cells in mouse liver.</a> Hepatology. 34, 519-522 (2001).</li><li>Shinozuka, H., Lombardi, B., Sell, S., Iammarino, R. 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J., Olynyk, J. K., Ramm, G. A., Tirnitz-Parker, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=TWEAK+and+LTbeta+Signaling+during+Chronic+Liver+Disease.">TWEAK and LTbeta Signaling during Chronic Liver Disease.</a> Front Immunol. 5, 39 (2014).</li><li>Karin, M., Clevers, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reparative+inflammation+takes+charge+of+tissue+regeneration.">Reparative inflammation takes charge of tissue regeneration.</a> Nature. 529, 307-315 (2016).</li><li>Ruddell, R. 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慢性肝病通常是一种无声的疾病, 大多数患者无症状, 它是一个主要的贡献, 发病率和死亡率在世界范围内。慢性酒精中毒和丙肝病毒感染是主要原因。慢性肝损伤的特点是肝脏炎症、纤维化和严重的肝硬化、肝癌和最终肝功能衰竭。目前尚无治疗方法, 虽然已取得重大进展, 了解肝脏疾病的机制, 但仍然迫切需要新的治疗途径。
时间饮食是一个简单的, 容易管理的模型, 诱导和?...

肝脏是身体的最大的腺新陈代谢的器官并且有许多复杂作用。肝脏的关键作用包括消化、新陈代谢、解毒、储存必要的营养素、生产血浆蛋白组分、以及通过驻留巨噬细胞或枯否体细胞介导的免疫。肝脏有很大的能力再生, 即使高达70-90% 的总质量损失。在急性肝损伤的情况下, 如看到部分肝切除或扑热息痛中毒, 其余健康肝细胞增殖, 以修复在高度协调的过程中的损害1。然而, 当肝细胞因长期病毒感染、酒精或非酒精性脂肪肝而长期受伤时, 炎症微环境引发肝纤维化的活化, 并肝祖细胞增殖 (LPCs), 有可能分化为胆管或肝细胞2,3,4,5。确切的起源, 分化命运的 LPCs, 他们对肝脏再生的贡献, 和肝癌一直是激烈辩论的话题, 很可能取决于伤害的严重性和上下文2。早期再生相关事件的顺序也有争议的讨论, 一些研究人员指出, 肝星状细胞活化和基质重塑对于产生 LPC 偏爱的小生境6是必不可少的, 而其他报告, 要触发纤维7, 需要 LPC 扩展和 so-called 胆管反应。有许多动物模型研究损伤和再生的具体方面, 试图了解所有的潜在因素, 调节疾病进展, 并最终制定新的治疗策略的病人8。
本发明的胆碱缺乏和氨基补充的饮食模型最初是为大鼠使用的, 后来改性用于小鼠的慢性肝损伤诱导,9,10。膳食缺乏的胆碱导致受损的组装和分泌非常低密度脂蛋白。结合 hepatocarcinogen dl-氨基, 这种方案导致过量的肝脏脂肪负荷, 连续炎症, periportal 纤维化, LPC 反应和长期肝癌的发展11,12.然而, 重要的是, 不同的小鼠菌株表现出鲜明的炎症, 化和 LPC 反应动力学13。本协议描述了慢性肝损伤诱导的 C57BL/6J 小鼠, 最常用的近交系小鼠品系。
在慢性肝病的研究中, 典型的分析包括苏木精和红素的组织学评估以及天狼星红色染色, 以可视化胶原沉积、免疫组织化学或荧光检测肝细胞的数量,组通过复杂生长因子和细胞因子网络协调诱导细胞变化的肝脏微环境分析14,15,16,17,18。

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	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:332px;">Six-week-old male C57BL/6J mice&#160;</td>
			<td style="width:367px;">Animal Resource Centre of Western Australia, Murdoch, WA,&#160; Australia</td>
			<td style="width:121px;">N/A</td>
			<td style="width:163px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">10 Kg Steam Cut Wheaten Chaff&#160;</td>
			<td>Specialty Feeds, Glen Forrest, WA, Australia&#160;</td>
			<td>N/A</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Water for irrigation 1000ml bottle (Baxter)&#160;</td>
			<td>Surgical House, Perth, WA, Australia&#160;</td>
			<td>AHF7114A</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Choline- deficient diet, modified (pellets)&#160;</td>
			<td colspan="2">MP Biomedicals Australasia Pty Limited, WA, Australia 02960210</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">DL-Ethionine&#160;</td>
			<td>Sigma-Aldrich, Castle Hill, NSW,&#160; Australia&#160;</td>
			<td>E5139-25G</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Ketamil injection&#160;</td>
			<td>Troy Laboratories Pty Limited, Glendenning, NSW, Australia&#160;</td>
			<td>N/A</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Ilum Xylazil-20 injection&#160;</td>
			<td>Troy Laboratories Pty Limited, Glendenning, NSW, Australia&#160;</td>
			<td>N/A</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">27G x 1/2&#34;, Regular Wall Needle&#160;</td>
			<td>Terumo Australia Pty Limited, NSW, Australia&#160;</td>
			<td>NN-2713R</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Syringes Terumo 1ml and 10ml&#160;</td>
			<td>Terumo Australia Pty Limited, Macquarie Park, NSW, Australia&#160;</td>
			<td>1018242, 1018037</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Tissue-Tek OCT compound&#160;</td>
			<td>VWR International Pty Limited, Tingalpa, QLD, Australia&#160;</td>
			<td>25608-930</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Neutral buffered formalin&#160;</td>
			<td>Amber Scientific, Midvale, WA, Australia&#160;</td>
			<td>NBF-2.5L</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Ethanol absolute anaLaR normalpur&#160;</td>
			<td>VWR International Pty Limited, Tingalpa, QLD, Australia&#160;</td>
			<td>20821.33</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Superfrost Plus slides&#160;</td>
			<td>Grale Scientific Pty Limited, Ringwood, VIC, Australia&#160;</td>
			<td>SF41296SP</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Dako Protein Block, serum-free&#160;</td>
			<td>Dako Australia Pty Limited, North Sydney, NSW, Australia&#160;</td>
			<td>&#160;X090930-2</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Dako antibody diluent&#160;</td>
			<td>Dako Australia Pty Limited, North Sydney, NSW, Australia&#160;&#160;</td>
			<td>s0809</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">rat anti-CD45&#160;</td>
			<td>eBioscience, San Diego, California, USA&#160;</td>
			<td>m0701&#160;</td>
			<td>1/200 dilution</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">rabbit anti-panCK&#160;</td>
			<td>Dako Australia Pty Limited, North Sydney, NSW, Australia&#160;&#160;</td>
			<td>Z0622</td>
			<td>1/300 dilution</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Goat anti-rabbit (Alexa Fluor 488)</td>
			<td>Life Technologies Australia Pty Limited, Mulgrave, VIC, Australia</td>
			<td>A-11008</td>
			<td>1/500 dilution</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Goat anti-rat IgG (Alexa Fluor 594)&#160;</td>
			<td>Life Technologies Australia Pty Limited, Mulgrave, VIC, Australia</td>
			<td>&#160;A-11007</td>
			<td>1/500 dilution</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">ProLong Gold Antifade Reagent with DAPI&#160;</td>
			<td>Life Technologies Australia Pty Limited, Mulgrave, VIC, Australia&#160;</td>
			<td>P36935&#160;&#160;</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Picrosirius Red Stain Kit</td>
			<td>Polysciences Inc., Warrington, PA, USA&#160;</td>
			<td>ab150681&#8195;</td>
			<td></td>
		</tr>
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the murine choline-deficient, ethionine-supplemented (cde) diet model of chronic liver injury

慢性肝病, 如病毒性肝炎、酒精性肝病或非酒精脂肪肝, 其特征是持续性炎症、肝实质的渐进破坏和再生, 肝祖细胞增殖和纤维化。每个慢性肝病的终末期是肝硬化, 是肝癌发展的主要危险因素。为了研究控制疾病的起始、建立和进展的过程, 实验室中使用了几种动物模型。在这里, 我们描述了六周的时间过程中的胆碱缺乏和氨基补充 (多) 小鼠模型, 其中包括喂养六周的老雄性 C57BL/6J 小鼠与胆碱缺乏的 chow 和 0.15% DL 氨基补充饮用水。对动物健康的监测和典型的体重损失曲线进行了解释。该协议显示了经心脏穿刺治疗的肝脏和血液采集的总检查结果, 随后进行血清分析。其次, 肝脏灌注技术和收集不同的肝叶的标准评价显示, 包括肝组织学评估的苏木精和曙红或天狼星红色染色, 荧光检测肝细胞数量以及对肝脏微环境的转录分析。这种小鼠模型适于研究慢性肝病诱发的炎症、化和肝祖细胞动力学, 并可用于测试可能调节这些过程的潜在治疗剂。

在六周的时间过程中, 多因素诱发慢性肝损伤, 参数评估的天数 7 (归纳阶段), 14 和 21 (建立阶段) 和 42 (维护阶段)。与对照组小鼠相比, 经治疗的小鼠在初始适应阶段损失了20% 的初始体重, 并倾向于在建立和维护阶段重拾重量(图 1)。体重与血清丙氨酸转氨酶水平呈反比关系, 这是肝细胞损伤的指标(图 2)。...

Watch this Scientific Journal Video about The Murine Choline-Deficient, Ethionine-Supplemented CDE Diet Model of Chronic Liver Injury at JoVE.com

The Murine Choline-Deficient, Ethionine-Supplemented CDE Diet Model of Chronic Liver Injury

在这里, 我们描述了一个常见的方法来诱导慢性肝损伤小鼠通过喂养的胆碱缺乏和氨基补充 (多) 饮食。我们展示健康监测, 肝脏灌注, 隔离和保存。六周的时间过程可以告知肝损伤、病理、纤维化、炎症和肝祖细胞反应。

慢性肝损伤小鼠胆碱缺乏、氨基补充的饮食模型

Chronic liver diseases, such as viral hepatitis, alcoholic liver disease, or non-alcoholic fatty liver disease, are characterized by continual ...

Research

JoVE Journal

Biology

The Murine Choline-Deficient, Ethionine-Supplemented (CDE) Diet Model of Chronic Liver Injury

Laser-Induced Chronic Ocular Hypertension Model on SD Rats

激光诱导SD大鼠慢性高眼压高压模型

Glaucoma is one of the major causes of blindness in the world. Elevated intraocular pressure is a major risk factor. Laser photocoagulation induced ocular ...

科研

生物学

Murine Model of Hindlimb Ischemia 

肢体缺血小鼠模型

In the United States, peripheral arterial disease (PAD) affects about 10 million individuals, and is also prevalent worldwide.  Medical therapies for ...

Murine Renal Transplantation Procedure

小鼠肾移植程序

Renal orthotopic transplantation in mice is a technically challenging procedure. Although the first kidney transplants in mice were performed by Russell ...

Chronic Salmonella Infected Mouse Model

Chronic Salmonella Infected Mouse Model

慢性沙门氏菌感染小鼠模型

The bacterial infected mouse model is a powerful model system for studying areas such as infection, inflammation, immunology, signal transduction, and ...

Do-It-Yourself Device for Recovery of Cryopreserved Samples Accidentally Dropped into Cryogenic Storage Tanks

做自己动手恢复冻存样品的设备不慎掉落到低温储罐

Liquid nitrogen is colorless, odorless, extremely cold (-196 &deg;C) liquid kept under pressure. It is commonly used as a cryogenic fluid for long term ...

Isolation of Murine Valve Endothelial Cells

鼠阀门内皮细胞的分离

Normal valve structures consist of stratified layers of specialized extracellular matrix (ECM) interspersed with valve interstitial cells (VICs) and ...

Murine Aortic Crush Injury: An Efficient In Vivo Model of Smooth Muscle Cell Proliferation and Endothelial Function

Murine Aortic Crush Injury: An Efficient In Vivo Model of Smooth Muscle Cell Proliferation and Endothelial Function

小鼠主动脉夹层损伤：高效<em&gt;体内</em&gt;平滑肌细胞增殖和内皮功能模型

Arterial reconstruction, whether angioplasty or bypass surgery, involves iatrogenic trauma causing endothelial disruption and vascular smooth muscle cell ...

A Murine Model of Vertical Sleeve Gastrectomy

小鼠垂直套胃切除术的动物模型

Bariatric surgery, such as vertical sleeve gastrectomy (VSG), is a surgery of the gastrointestinal tract that is performed for the purpose of weight loss. ...

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

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 ...

Mass Spectrometry Analysis to Identify Ubiquitylation of EYFP-tagged CENP-A (EYFP-CENP-A)

Mass Spectrometry Analysis to Identify Ubiquitylation of EYFP-tagged CENP-A EYFP-CENP-A

质谱分析，用于识别 EYFP 标记 CENP-A （EYFP-CENP-A） 的泛化

Studying the structure and the dynamics of kinetochores and centromeres is important in understanding chromosomal instability (CIN) and cancer ...