Ex-vivo perfusion of human liver tissue as model for pharmacological liver inury and hepatic steatosis

我们能够证明，我们的灌注系统可以模拟中毒离体对乙酰氨基酚在人体肝脏组织标本在目前资助的项目。我们所采用的系统提供了在天然结构和合作细胞结构中使用完整的人肝组织的优点。灌注系统中的对乙酰氨基酚毒性反映了患者临床观察结果的个体间差异，这在体外或动物模型中不会发生。MicroRNA-122释放到灌注液和线粒体损伤被确定为对乙酰氨基酚诱导的肝损伤的早期事件。这再次反映了对乙酰氨基酚引起的急性肝衰竭的临床情况。在拟定的随访项目中，对乙酰氨基酚剂量将进行调整，以达到100%的中毒率。这将允许系统表征，以在未来分析未知物质的肝毒性。因此，我们希望确定对乙酰氨基酚诱导的急性肝衰竭的个体、患者特异性危险因素。此外，它将能够测试对乙酰氨基酚诱导的急性肝衰竭的既定和实验性治疗方法，并确定所涉及的细胞类型和分子机制。为此，将收集来自灌注液以及来自肝组织的样品，并且在对乙酰氨基酚剂量和N-乙酰半胱氨酸剂量中的任一种之后，将评估用于损伤和用于特定细胞类型的标志物的分泌以及表达。（已建立的疗法）或血栓素合酶抑制剂的应用灌注模型也将被扩展以产生肝脏脂肪变性，独立于我们系统中对乙酰氨基酚毒性的表征。与世界范围内的肥胖流行并行，非酒精性脂肪性肝病（NAFLD）正在上升，并且是目前工业化国家最常见的慢性肝病。NAFLD的基础是肝脏中过量的脂肪积累（脂肪变性）。脂肪积累和随后的肝损伤的早期过程仍然不完全了解或主要基于体外模型。在人体离体灌注模型中，我们可以产生脂肪变性，并将在拟议的项目中阐明这一过程的细胞和调控机制。特别是，我们将能够观察脂肪变性发展的早期机制和随后的代谢变化，以及在完整的合作细胞结构中人肝组织中非实质肝细胞的行为。我们期待对NAFLD可能的预防或治疗方法的新见解。

We were able to demonstrate that our perfusion system can mimic poisoning by acetaminophen ex vivo in human liver tissue specimen in the currently funded project. The system employed by us offers the advantage of using intact human liver tissue in a native architecture and cooperative cellular structure. Acetaminophen toxicity in the perfusion system reflected inter-individual variance from clinical observations in patients, which do not occur in vitro or in animal models. MicroRNA-122 release into the perfusate and mitochondrial damage were identified as early events in acetaminophen-induced liver injury. This again reflects the clinical situation in acetaminophen-induced acute liver failure.In the proposed follow up project the acetaminophen dose will be adapted to achieve 100 % poisoning rate. This will allow characterization of the system to analyze liver toxicity of unknown substances in the future. Thus we hope to identify individual, patient specific risk factors for acetaminophen-induced acute liver failure. Furthermore it will enable testing of established as well as experimental therapeutic approaches for acetaminophen-induced acute liver failure and to identify the involved cell types and molecular mechanisms. To this end, samples from the perfusate as well as from liver tissue will be collected and secretion as well as expression of markers for injury and for specific cell types will be assessed after acetaminophen dosage and either N-acetylcysteine dosage (established therapy) or application of thromboxane-synthase inhibitors (experimental therapy).The perfusion model will also be expanded for generation of liver steatosis, independent from characterization of acetaminophen toxicity in our system. In parallel to the world-wide obesity pandemic non-alcoholic fatty liver disease (NAFLD) is on the rise and is the most common chronic liver disease in industrialized countries by now. The basis for NAFLD is excess fat accumulation (steatosis) in the liver. Early processes of fat accumulation and subsequent liver injury are still incompletely understood or are largely based on in vitro models. In the human ex vivo perfusion model we can generate steatosis and will elucidate cellular and regulatory mechanisms of this process in the proposed project. In particular, we will be able to observe the early mechanisms of steatosis development and subsequent metabolic changes as well as behavior of non-parenchymal liver cells in human liver tissue in an intact cooperative cell structure. We expect novel insights into possible preventive or therapeutic approaches against NAFLD.