Modulators of hepatic response to TGF-ß mediated cellular injury in inbred mice: Identification of modifiers of fibrosis susceptibility
Final Report Abstract
We have performed a QTL mapping experiment in cultured hepatocytes following treatment with TGF-β and combined the results with QTL mapping of fibrogenesis-related phenotypes following injection of CCl4 in vivo to delineate a 6 Mb chromosomal region on mouse chromosome 11 involved in both phenotypes. Using expression datasets from acute and chronic liver damage in vivo we refined a genomic locus at 83.5 Mb that exerts a major impact on gene regulation during both processes. The nearest coding variant that shows a difference between the parental strains of the BXD lines was found in the Expi gene, which renders this gene a creedal candidate for mediating the observed differences. A knockout mouse showed decreased collagen content following chronic liver damage by CCl4. Literature data revealed that Expi is stimulated by the TLR9-ligand CpG and synergistically increased by TLR3-ligand poly I:C, suggesting a role in innate immune defence. Biochemical analyses of Expi function in normal tissue suggests a role in priming cells for apoptosis during mammary gland development and differentiation. The immediately adjacent and highly homologous (70% amino acid identity) Wfdc17 gene is a member of the same gene family of short proteinase inhibitors with antimicrobial functions and was shown to be a negative regulator of macrophage activation in microglia. Hence, we hypothesise that Expi sensitises hepatocytes to pro-apoptotic signals by TGF-β and various other, yet to be determined signalling molecules following stimulation by CpG dinucleotides via TLR9. ApcMin/+ mice show substantial induction of Expi in gastrointestinal adenoma compared to normal colonic tissue. Slightly lower levels of upregulation were observed in mammary tumours and preneoplastic mammary tissue from ErbB2/Neu transgenic mice with deregulated TGF-β signalling. Results from a study in rabbits show beneficial effects of co-stimulating tumours with TLR9- ligand CpG oligonucleotide during chemotherapy. We speculate that Expi is part of the innate immune system and as such might be involved in the interaction between the tumour and its tissue environment. It is also conceivable that Expi plays a role during tissue remodelling and metastasis due to its proteinase inhibitor function. To clarify the role of Expi in liver fibrogenesis, we propose to characterise expression and localisation of the Expi peptide in various models of liver damage using an antibody that we will generate. To investigate the role of Expi in tumour growth and metastasis, we propose generating a double knockout by crossing Expi-/- and ApcMin/+ mice and quantitatively assess tumour formation and metastasis (in collaboration with the group of Matthias Ebert at Mannheim University Hospital, who has complementary expertise and ApcMin/+ mice). There is no orthologue of the Expi/Wfdc18 gene in humans, since humans only possess 15 WFDC genes, but human WFDC2 shows similar tumour-specific expression patterns to Expi, whereas the murine Wfdc2 is not known for any involvement in tumours. This observation implicates the Expi knockout mouse as an appropriate model to study the role of the WFDC peptides, defensin-like alarm peptides, in the interaction between tumour and surrounding tissue (stroma).We propose the Expi knockout mouse as the murine model to study the role and function of human WFDC2/HE4. Studying the role of Wfdc18/Expi in the crosstalk between immunity and cellular fate might provide novel clues on how to interpret and eventually intercept signals that lead to persistent inflammation and deregulated growth.
Publications
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IL-13 induces connective tissue growth factor in rat hepatic stellate cells via TGF-β-independent Smad signaling. J Immunol 2011 Sep 1;187(5):2814-2823
Liu Y, Meyer C, Müller A, Herweck F, Li Q, Müllenbach R, Mertens PR, Dooley S, Weng HL
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A frequent variant in the human bile salt export pump gene ABCB11 is associated with hepatitis C virus infection, but not liver stiffness in a German population. BMC Gastroenterol 2012 Jun 8;12:63
Müllenbach R, Weber SN, Krawczyk M, Zimmer V,Sarrazin C, Lammert F, Grünhage F
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Common genetic variation in vitamin D metabolism is associated with liver stiffness. Hepatology 2012 Nov;56(5):1883-1891
Grünhage F, Hochrath K, Krawczyk M, Höblinger A, Obermayer-Pietsch B, Geisel J, Trauner M, Sauerbruch T, Lammert F
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Identification of RARRES1 as a core regulator in liver fibrosis. J Mol Med 2012 Dec;90(12):1439-1447
Teufel A, Becker D, Weber SN, Dooley S, Breitkopf-Heinlein K, Maass T, Hochrath K, Krupp M, Marquardt JU, Kolb M, Korn B, Niehrs C, Zimmermann T, Godoy P, Galle PR, Lammert F
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Personalised hepatology - current concepts, developments and expectations in the post-genome era. Z Gastroenterol 2012 Jan;50(1):41-46
Teufel A, Marquardt JU, Dooley S, Lammert F, Galle PR
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IFN-γ inhibits liver progenitor cell proliferation in HBV-infected patients and in 3,5- diethoxycarbonyl-1,4-dihydrocollidine diet-fed mice. J Hepatol 2013 Oct;59(4):738-745
Weng HL, Feng DC, Radaeva S, Kong XN, Wang L, Liu Y, Li Q, Shen H, Gao YP, Müllenbach R, Munker S, Huang T, Chen JL, Zimmer V, Lammert F, Mertens PR, Cai WM, Dooley S, Gao B
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Modeling hepatic osteodystrophy in Abcb4 deficient mice. Bone 2013 Aug;55(2):501-511
Hochrath K, Ehnert S, Ackert-Bicknell CL, Lau Y, Schmid A, Krawczyk M, Hengstler JG, Dunn J, Hiththetiya K, Rathkolb B, Micklich K, Hans W, Fuchs H, Gailus-Durner V, Wolf E, de Angelis MH, Dooley S, Paigen B, Wildemann B, Lammert F, Nüssler AK
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Systems genetics of hepatocellular damage in vivo and in vitro: identification of a critical network on chromosome 11 in mouse. Physiol Genomics 2013 Oct;45(20):931-939
Liebe R, Hall RA, Williams RW, Dooley S, Lammert F
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Einfluss der Expressionsvariation des Transforming Growth Factor beta Typ 2 Rezeptors (TGFbRII) auf phänotypische Aspekte der Leberfibrose im Modell von extrem exprimierenden Mauslinien. Dissertation, Medizinische Fakultät der Universität des Saarlandes, Homburg, 2014. VIII, 87 S.
Natalie Fund
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Systems genetics of liver fibrosis: identification of fibrogenic and expression quantitative trait loci in the BXD murine reference population. PLoS One 2014 Feb;9(2):e89279
Hall RA, Liebe R, Hochrath K, Kazakov A, Alberts R, Laufs U, Böhm M, Fischer HP, Williams RW, Schughart K, Weber SN, Lammert F
