The CYP2D6 animal model: how to induce autoimmune hepatitis in mice

Autoimmune hepatitis is a rare but life threatening autoimmune disease of the liver of unknown etiology1,2. In the past many attempts have been made to generate an animal model that reflects the characteristics of the hu
Autoimmune hepatitis is a rare but life threatening autoimmune disease of the liver of unknown etiology1,2. In the past many attempts have been made to generate an animal model that reflects the characteristics of the human disease 3-5. However, in various models the induction of disease was rather complex and often hepatitis was only transient3-5. Therefore, we have developed a straightforward mouse model that uses the major human autoantigen in type 2 autoimmune hepatitis (AIH-2), namely hCYP2D6, as a trigger6. Type 1 liver-kidney microsomal antibodies (LKM-1) antibodies recognizing hCYP2D6 are the hallmark of AIH-27,8. Delivery of hCYP2D6 into wildtype FVB or C57BL/6 mice was by an Adenovirus construct (Ad-2D6) that ensures a direct delivery of the triggering antigen to the liver. Thus, the ensuing local inflammation generates a fertile field9 for the subsequent development of autoimmunity. A combination of intravenous and intraperitoneal injection of Ad-2D6 is the most effective route to induce a long-lasting autoimmune damage to the liver (section 1). Here we provide a detailed protocol on how autoimmune liver disease is induced in the CYP2D6 model and how the different aspects of liver damage can be assessed. First, the serum levels of markers indicating hepatocyte destruction, such as aminotransferases, as well as the titers of hCYP2D6 antibodies are determined by sampling blood retroorbitaly (section 2). Second, the hCYP2D6-specific T cell response is characterized by collecting lymphocytes from the spleen and the liver. In order to obtain pure liver lymphocytes, the livers are perfused by PBS via the portal vein (section 3), digested in collagen and purified over a Percoll gradient (section 4). The frequency of hCYP2D6-specific T cells is analyzed by stimulation with hCYP2D6 peptides and identification of IFNγ-producing cells by flow cytometry (section 5). Third, cellular infiltration and fibrosis is determined by immunohistochemistry of liver sections (section 6). Such analysis regimen has to be conducted at several times after initiation of the disease in order to prove the chronic nature of the model. The magnitude of the immune response characterized by the frequency and activity of hCYP2D6-specific T and/or B cells and the degree of the liver damage and fibrosis have to be assessed for a subsequent evaluation of possible treatments to prevent, delay or abrogate the autodestructive process of the liver.
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Metadaten
Author:Edith Hintermann, Janine Ehser, Urs Christen
URN:urn:nbn:de:hebis:30:3-288743
DOI:http://dx.doi.org/10.3791/3644
ISSN:1940-087X
Pubmed Id:http://www.ncbi.nlm.nih.gov/pubmed?term=22331063
Parent Title (English):Journal of visualized experiments : JoVE
Publisher:[s.n.]
Place of publication:[S.l.]
Document Type:Article
Language:English
Date of Publication (online):2012/02/03
Date of first Publication:2012/02/03
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2013/02/14
Tag:autoantigen; autoimmunity; fibrosis; liver; perfusion
Volume:60
Issue:e3644
Pagenumber:7
Note:
Copyright © 2012, Journal of Visualized Experiments. This is an open-access article distributed under the terms of the Creative Commons Attribution Non-Commercial License, which permits non-commercial use, distribution, and reproduction, provided the original work is properly cited.
Note:
The video component of this article can be found at http://www.jove.com/video/3644/
HeBIS PPN:331501716
Institutes:Medizin
Zentrum für Arzneimittelforschung, Entwicklung und Sicherheit
Dewey Decimal Classification:610 Medizin und Gesundheit
Sammlungen:Universitätspublikationen
Licence (German):License LogoCreative Commons - Namensnennung 3.0

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