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Precision-cut human liver slice cultures as an immunological platform.

ABSTRACT: The liver is the central metabolic organ in the human body, and also plays an essential role in innate and adaptive immunity. While mouse models offer significant insights into immune-inflammatory liver disease, human immunology differs in important respects. It is not easy to address those differences experimentally. Therefore, to improve the understanding of human liver immunobiology and pathology, we have established precision-cut human liver slices to study innate immunity in human tissue. Human liver slices collected from resected livers could be maintained in ex vivo culture over a two-week period. Although an acute inflammatory response accompanied by signs of tissue repair was observed in liver tissue following slicing, the expression of many immune genes stabilized after day 4 and remained stable until day 15. Remarkably, histological evidence of pre-existing liver diseases was preserved in the slices for up to 7?days. Following 7?days of culture, exposure of liver slices to the toll-like receptor (TLR) ligands, TLR3 ligand Poly-I:C and TLR4 ligand LPS, resulted in a robust activation of acute inflammation and cytokine genes. Moreover, Poly-I:C treatment induced a marked antiviral response including increases of interferons IFNB, IL-28B and a group of interferon-stimulated genes. Therefore, precision-cut liver slices emerge as a valuable tool to study human innate immunity.

SUBMITTER: Wu X

PROVIDER: S-EPMC6689534 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

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Precision-cut human liver slice cultures as an immunological platform.

Wu Xia X Roberto Jessica B JB Knupp Allison A Kenerson Heidi L HL Truong Camtu D CD Yuen Sebastian Y SY Brempelis Katherine J KJ Tuefferd Marianne M Chen Antony A Horton Helen H Yeung Raymond S RS Crispe Ian N IN

Journal of immunological methods 20180201

The liver is the central metabolic organ in the human body, and also plays an essential role in innate and adaptive immunity. While mouse models offer significant insights into immune-inflammatory liver disease, human immunology differs in important respects. It is not easy to address those differences experimentally. Therefore, to improve the understanding of human liver immunobiology and pathology, we have established precision-cut human liver slices to study innate immunity in human tissue. H ...[more]

PMID: 29408707

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Project description:Precision-cut liver tissue slice (PCLS) contains all major cell types of the liver parenchyma and preserves the original cell-cell and cell-matrix contacts. It represents a promising ex vivo model to study liver fibrosis and test the antifibrotic effect of experimental compounds in a physiological environment. In this study using RNA sequencing, we demonstrated that various pathways functionally related to fibrotic mechanisms were dysregulated in PCLSs derived from rats subjected to bile duct ligation. The activin receptor-like kinase-5 (Alk5) inhibitor SB525334, nintedanib, and sorafenib each reversed a subset of genes dysregulated in fibrotic PCLSs, and of those genes we identified 608 genes whose expression was reversed by all three compounds. These genes define a molecular signature characterizing many aspects of liver fibrosis pathology and its attenuation in the model. A panel of 12 genes and 4 secreted biomarkers including procollagen I, hyaluronic acid (HA), insulin-like growth factor binding protein 5 (IGFBP5), and WNT1-inducible signaling pathway protein 1 (WISP1) were further validated as efficacy end points for the evaluation of antifibrotic activity of experimental compounds. Finally, we showed that blockade of ?V-integrins with a small molecule inhibitor attenuated the fibrotic phenotype in the model. Overall, our results suggest that the rat fibrotic PCLS model may represent a valuable system for target validation and determining the efficacy of experimental compounds. NEW & NOTEWORTHY We investigated the antifibrotic activity of three compounds, the activin receptor-like kinase-5 (Alk5) inhibitor SB525334, nintedanib, and sorafenib, in a rat fibrotic precision-cut liver tissue slice model using RNA sequencing analysis. A panel of 12 genes and 4 secreted biomarkers including procollagen I, hyaluronic acid (HA), insulin-like growth factor binding protein 5 (IGFBP5), and WNT1-inducible signaling pathway protein 1 (WISP1) were then established as efficacy end points to validate the antifibrotic activity of the ?V-integrin inhibitor CWHM12. This study demonstrated the value of the rat fibrotic PCLS model for the evaluation of antifibrotic drugs.

Project description:Background & aimsIncreased sensitivity towards tumor necrosis factor (TNF)-induced cell death in virus-infected hepatocytes has revealed a so far unrecognized hepatocyte-intrinsic antiviral immune surveillance mechanism, for which no in vitro or ex vivo model is available. We aimed to establish precision-cut liver slices (PCLS) as a model system to study hepatocyte-intrinsic regulation of apoptosis.MethodsPreparation of PCLS from mouse and human liver tissue was optimized for minimal procedure-associated apoptosis. Functionality of liver cells in PCLS was characterized using extracellular flux analysis to determine mitochondrial respiration, and viral infection with recombinant adenovirus and lymphocytic choriomeningitis virus (LCMV) was used to probe for hepatocyte-intrinsic sensitivity towards apoptosis in PCLS. Apoptosis was detected by immunohistochemical staining for cleaved-caspase 3 and quantified by detection of effector caspase activity in PCLS.ResultsWe established an optimized protocol for preparation of PCLS from human and mouse models using agarose-embedding of liver tissue to improve precision cutting and using organ-protective buffer solutions to minimize procedure-associated cell death. PCLS prepared from virus-infected livers showed preserved functional metabolic properties. Importantly, in PCLS from adenovirus- and LCMV-infected livers we detected increased induction of apoptosis after TNF challenge ex vivo.ConclusionWe conclude that PCLS can be used as model system to ex vivo characterize hepatocyte-intrinsic sensitivity to cell death. This may also enable researchers to characterize human hepatocyte sensitivity to apoptosis in PCLS prepared from patients with acute or chronic liver diseases.Lay summaryVirus-infected hepatocytes in vivo show an increased sensitivity towards induction of cell death signaling through the TNF receptor. Studying this hepatocyte-intrinsic antiviral immune surveillance mechanism has been hampered by the absence of model systems that reciprocate the in vivo finding of increased apoptosis of virus-infected hepatocytes challenged with TNF. Herein, we report that an optimized protocol for generation of precision-cut liver slices can be used to study this hepatocyte-intrinsic surveillance mechanism ex vivo.

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Precision-cut human liver slice cultures as an immunological platform.

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Precision-cut human liver slice cultures as an immunological platform.

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