In Vitro Models for the Study of Liver Biology and Diseases: Advances and Limitations
2022; Elsevier BV; Volume: 15; Issue: 3 Linguagem: Inglês
10.1016/j.jcmgh.2022.11.008
ISSN2352-345X
AutoresSavneet Kaur, Srivatsan Kidambi, Martí Ortega‐Ribera, Lê Thị Thanh Thủy, Natalia Nieto, Victoria C. Cogger, Wei‐Fen Xie, Frank Tacke, Jordi Gracia‐Sancho,
Tópico(s)Pancreatic function and diabetes
ResumoIn vitro models of liver (patho)physiology, new technologies, and experimental approaches are progressing rapidly. Based on cell lines, induced pluripotent stem cells or primary cells derived from mouse or human liver as well as whole tissue (slices), such in vitro single- and multicellular models, including complex microfluidic organ-on-a-chip systems, provide tools to functionally understand mechanisms of liver health and disease. The International Society of Hepatic Sinusoidal Research (ISHSR) commissioned this working group to review the currently available in vitro liver models and describe the advantages and disadvantages of each in the context of evaluating their use for the study of liver functionality, disease modeling, therapeutic discovery, and clinical applicability. In vitro models of liver (patho)physiology, new technologies, and experimental approaches are progressing rapidly. Based on cell lines, induced pluripotent stem cells or primary cells derived from mouse or human liver as well as whole tissue (slices), such in vitro single- and multicellular models, including complex microfluidic organ-on-a-chip systems, provide tools to functionally understand mechanisms of liver health and disease. The International Society of Hepatic Sinusoidal Research (ISHSR) commissioned this working group to review the currently available in vitro liver models and describe the advantages and disadvantages of each in the context of evaluating their use for the study of liver functionality, disease modeling, therapeutic discovery, and clinical applicability. SummaryThis review provides a glimpse on the most advanced methods for studying liver sinusoidal biology in vitro to uncover liver disease pathophysiology focusing on liver-on-a-chip and microfluidic devices, liver scaffolds and matrices, mechanobiological cues, spheroids/organoids, liver slices and omics. This review provides a glimpse on the most advanced methods for studying liver sinusoidal biology in vitro to uncover liver disease pathophysiology focusing on liver-on-a-chip and microfluidic devices, liver scaffolds and matrices, mechanobiological cues, spheroids/organoids, liver slices and omics. Liver disease represents one of the leading causes of death worldwide, and the incidence of some pathologies, such as nonalcoholic fatty liver disease (NAFLD), nonalcoholic steatohepatitis (NASH), and liver cancer, continues to increase.1Karlsen T.H. Sheron N. Zelber-Sagi S. et al.The EASL–Lancet Liver Commission: protecting the next generation of Europeans against liver disease complications and premature mortality.Lancet. 2022; 399: 61-116Abstract Full Text Full Text PDF PubMed Google Scholar Despite years of research, liver diseases still have limited treatment options in the clinic. This paucity of treatments is explained partly by the limitations of traditional in vitro tools and animal models that do not accurately mimic the clinical pathophysiology of diseases and have a low accuracy for drug discovery purposes. Indeed, several studies have shown that traditional cell culture methodologies do not reflect the complexity of a human liver in vivo and thus cannot predict drug sensitivity. In contrast, animal models differ in biology compared with human pathologies, which explains why promising therapies tested in animal models often fail when tested in human beings and, unfortunately, the field of hepatology has numerous recent examples of failures in clinical phases.2Nevzorova Y.A. Boyer-Diaz Z. Cubero F.J. Gracia-Sancho J. Animal models for liver disease – a practical approach for translational research.J Hepatol. 2020; 73: 423-440Abstract Full Text Full Text PDF PubMed Scopus (87) Google Scholar With the advent of precision medicine, which offers much hope for individual patient outcomes, there is increased demand for robust and patient-specific tools to better improve our understanding and treatment of complex and multifactorial diseases such as liver diseases. Advances in vascular biology, microfluidics, and bioengineering have led to the development of sophisticated in vitro models that could fill this gap (Figure 1). In addition, omics techniques provide further insight to preclinical research in hepatology. In this review, we discuss the benefits and limitations of advanced in vitro research techniques that presently are being applied to the study of liver diseases and further critique how these tools may provide insight into the prediction of patient responses to a therapy. During the 21st century, the development of biology-inspired devices aimed at mimicking the sinusoidal niche integrating microfluidics led to the rapidly evolving liver-on-a-chip (LoC) technology.3Polidoro M.A. Ferrari E. Marzorati S. et al.Experimental liver models: from cell culture techniques to microfluidic organs-on-chip.Liver Int. 2021; 41: 1744-1761Crossref PubMed Scopus (13) Google Scholar The design of these in vitro liver-resembling tools, which have been reviewed extensively by Ortega-Ribera et al4Ortega-Ribera M. Yeste J. Villa R. Gracia-Sancho J. Nanoengineered biomaterials for the treatment of liver diseases.in: Mozafari M. Rajadas J. Kaplan D. Nanoengineered biomaterials for regenerative medicine. Elsevier, 2018: 417-441Google Scholar and is out of the scope of the present review, is inspired in sinusoidal cell biology, architecture, and hemodynamics, but materialized under each research teams’ eyes in terms of appearance, size, fabrication procedures, costs, and microfluidics integration, leading to significant variation in the finalized product. The latest advances in the field include chronic liver disease–specific devices, LoC models designed to study key pathophysiological processes in the development of liver disease, and to understand the interconnection with other organs-on-chip to better depict liver functions and systemic implications (Figure 2). Multi-organ chips, for instance, liver-, adipose-, and gut-on-a-chip connected, may be particularly suitable to understand organ-crosstalk in chronic liver disease (CLD), such as NAFLD/NASH or cholangiopathies.5Shroff T. Aina K. Maass C. et al.Studying metabolism with multi-organ chips: new tools for disease modelling, pharmacokinetics and pharmacodynamics.Open Biol. 2022; 12210333Crossref PubMed Scopus (0) Google Scholar In recent years, disease-focused LoC devices mimicking some of the landmark etiological characteristics of CLD have been developed. Fat accumulation in hepatocytes, occurring in NAFLD/NASH, has been represented in LoC under the combination of glucose and free fatty acids (usually a 2:1 ratio of oleic and palmitic acid).6Hassan S. Sebastian S. Maharjan S. et al.Liver-on-a-chip models of fatty liver disease.Hepatology. 2020; 71: 733-740Crossref PubMed Scopus (39) Google Scholar Antifibrotic compounds such as obeticholic acid, elafibranor,7Du K. Li S. Li C. et al.Modeling nonalcoholic fatty liver disease on a liver lobule chip with dual blood supply.Acta Biomater. 2021; 134: 228-239Crossref PubMed Scopus (7) Google Scholar pioglitazone, or metformin8Teng Y. Zhao Z. 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Petropolis D.B. Manatakis D.V. et al.Modeling alcohol-associated liver disease in a human Liver-Chip.Cell Rep. 2021; 36109393Abstract Full Text Full Text PDF PubMed Scopus (10) Google Scholar Importantly, Ortega-Prieto et al15Ortega-Prieto A.M. Skelton J.K. Wai S.N. et al.3D microfluidic liver cultures as a physiological preclinical tool for hepatitis B virus infection.Nat Commun. 2018; 9: 682Crossref PubMed Scopus (133) Google Scholar developed a model for hepatitis B virus (HBV) long-term infection in primary human hepatocytes that recapitulates virus–host interactions and its associated immune effectors. LoC devices using primary cells isolated through standardized protocols from preclinical models of CLD and from patients' liver tissue also have been developed.16Ortega-Ribera M. Fernández-Iglesias A. Illa X. et al.Resemblance of the human liver sinusoid in a fluidic device with biomedical and pharmaceutical applications.Biotechnol Bioeng. 2018; 115: 2585-2594Crossref PubMed Scopus (25) Google Scholar,17Fernandez-Igleasias A. Ortega Ribera M. Guixé-Muntet S. Gracia-Sancho J. 4 In 1: antibody-free protocol for isolating the main hepatic cells from healthy and cirrhotic single rat livers.J Cell Mol Med. 2019; 23: 877-886Crossref PubMed Scopus (8) Google Scholar These specialized LoC settings may widen the current knowledge on disease dynamics and provide potential applicability as in vitro preclinical models for drug screening. Even though LoC complexity has increased outstandingly since the initial models/prototypes, the intricacy of the whole liver still is under-represented. In this regard, several scientists brought the attention and focused their studies on specific processes or structures. For example, the essential features of the bile duct containing primary mouse cholangiocytes,18Du Y. Khandekar G. Llewellyn J. et al.A bile duct-on-a-chip with organ-level functions.Hepatology. 2020; 71: 1350-1363Crossref PubMed Scopus (31) Google Scholar the unique vasculature organization of the liver,19Ya S. Ding W.G. Li S. et al.On-chip construction of liver lobules with self-assembled perfusable hepatic sinusoid networks.ACS Appl Mater Interfaces. 2021; 13: 32640-32652Crossref PubMed Scopus (6) Google Scholar the sinusoidal zonation within LoC devices,20Kang Y.B.A. Eo J. Mert S. et al.Metabolic patterning on a chip: towards in vitro liver zonation of primary rat and human hepatocytes.Sci Rep. 2018; 8: 8951Crossref PubMed Scopus (60) Google Scholar neutrophil recruitment and interaction with liver sinusoidal endothelial cells (LSECs) after lipopolysaccharide stimulation,21Du Y. Li N. 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This model provided a promising tool for screening and characterization of hepatotoxicity in a 3D spheroid-embedded hydrogel system that more closely resembles conditions in vivo. In 2013, Takebe et al59Takebe T. Zhang R.R. Koike H. et al.Generation of a vascularized and functional human liver from an iPSC-derived organ bud transplant.Nat Protoc. 2014; 9: 396-409Crossref PubMed Scopus (261) Google Scholar described the in vitro generation of 3D liver bud organoids from human induced pluripotent stem cell–derived hepatic endoderm cells co-cultured with endothelial and mesenchymal lineages. Interestingly, when these liver buds were ectopically transplanted at various sites including the cranium, subrenal capsule, and distal and proximal-mesentery in immunodeficient mice they were able to rescue the drug-induced lethal liver failure model.61Takebe T. Sekine K. 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