Pro-fibrotic compounds induce stellate cell activation, ECM-remodelling and Nrf2 activation in a human 3D-multicellular model of liver fibrosis

Autor: Franziska Lampart, Vincenzo Prestigiacomo, Anna Weston, Laura Suter-Dick, Simon Messner
Rok vydání: 2017
Předmět:
0301 basic medicine
Lipopolysaccharides
Liver Cirrhosis
Physiology
Cellular differentiation
lcsh:Medicine
Thioacetamide
Biochemistry
Extracellular matrix
White Blood Cells
0302 clinical medicine
Fibrosis
Animal Cells
Immune Physiology
Medicine and Health Sciences
lcsh:Science
Innate Immune System
Multidisciplinary
Chemistry
Liver Diseases
Kupffer cell
Cell biology
Extracellular Matrix
medicine.anatomical_structure
Liver
030220 oncology & carcinogenesis
Hepatocyte
Liver Fibrosis
Cytokines
Tumor necrosis factor alpha
Cellular Types
Anatomy
Research Article
NF-E2-Related Factor 2
Immune Cells
Immunology
Gastroenterology and Hepatology
Cell Line
Transforming Growth Factor beta1
03 medical and health sciences
Albumins
medicine
Hepatic Stellate Cells
Humans
Vimentin
Blood Cells
Tumor Necrosis Factor-alpha
Macrophages
lcsh:R
Biology and Life Sciences
Proteins
Cell Biology
Molecular Development
medicine.disease
Cytoskeletal Proteins
030104 developmental biology
Methotrexate
Cell culture
Immune System
Hepatic stellate cell
Hepatocytes
lcsh:Q
Collagens
Developmental Biology
Zdroj: PLoS ONE
PLoS ONE, Vol 12, Iss 6, p e0179995 (2017)
ISSN: 1932-6203
Popis: Background & aims Currently most liver fibrosis research is performed in vivo, since suitable alternative in vitro systems which are able to recapitulate the cellular events leading to liver fibrosis are lacking. Here we aimed at generating a system containing cells representing the three key players of liver fibrosis (hepatocyte, Kupffer cells and stellate cells) and assess their response to pro-fibrotic compounds such as TGF-β1, methotrexate (MTX) and thioacetamide (TAA). Methods Human cell lines representing hepatocytes (HepaRG), Kupffer cell (THP-1 macrophages) and stellate cells (hTERT-HSC) were co-cultured using the InSphero hanging drop technology to generate scaffold-free 3D microtissues, that were treated with pro-fibrotic compounds (TGF-β1, MTX, TAA) for up to 14 days. The response of the microtissues was evaluated by determining the expression of cytokines (TNF-α, TGF-β1 and IL6), the deposition and secretion of ECM proteins and induction of gene expression of fibrosis biomarkers (e.g. αSMA). Induction of Nrf2 and Keap1, as key player of defence mechanism, was also evaluated. Results We could demonstrate that the multicellular 3D microtissue cultures could be maintained in a non-activated status, based on the low expression levels of activation markers. Macrophages were activated by stimulation with LPS and hTERT-HSC showed activation by TGF-β1. In addition, MTX and TAA elicited a fibrotic phenotype, as assessed by gene-expression and protein-deposition of ECM proteins such as collagens and fibronectin. An involvement of the antioxidant pathway upon stimulation with pro-fibrotic compounds was also observed. Conclusion Here, for the first time, we demonstrate the in vitro recapitulation of key molecular and cellular events leading to liver fibrosis: hepatocellular injury, antioxidant defence response, activation of Kupffer cells and activation of HSC leading to deposition of ECM.
Databáze: OpenAIRE
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