3D spheroid culture of hESC/hiPSC-derived hepatocyte-like cells for drug toxicity testing

Takayama, Kazuo; Kono, Kenji; Nagamoto, Yasuhito; Kishimoto, Keisuke; Tashiro, Kentaro; Sakurai, Fuminori; Tachibana, Masashi; Kanda, Katsuhiro; Hayakawa, Tetsuo; Furue, Miho; Mizuguchi, Hiroyuki

doi:10.1016/j.biomaterials.2012.11.029

Cited by 251 publications

(192 citation statements)

References 34 publications

Supporting

Mentioning

177

Contrasting

Order By: Relevance

“…hiPSC-derived hepatic cells, despite their potential to have malignant transformation following in vivo transplantations, have been widely recognized as the most promising candidate for developing patient-specific human hepatic models in vitro (4,7). Current in vitro liver models using hiPSC-derived hepatic cells are largely limited by their lack of biomimicry (3,7,(10)(11)(12)(13)(14). The 3D triculture model presented here highlights the successful application of DLP-based bioprinting technology to liver tissue engineering and thus the progress of the field to a level where the complex liver microarchitecture and cell composition can be studied in a physiologically relevant model.…”

Section: Discussionmentioning

confidence: 99%

“…Cells at both hepatic progenitor and maturation stages, i.e., HPCs and HLCs, respectively, have demonstrated their potentials in organizing into vascularized liver buds (11,13) and in the construction of in vitro models with simple geometry (10,12,14). To determine the stage of cells along the differentiation pathway that is more suitable for hydrogel encapsulation, HPCs and the relatively more mature HLCs were compared in terms of their albumin secretion time course following encapsulation in GelMA (Fig.…”

Section: D Hydrogel Encapsulation At Hepatic Progenitor Stage Demonsmentioning

confidence: 99%

“…Major liver functions are tightly linked to the 3D assembly of hepatocytes with the supporting cell types from both endodermal and mesodermal origins in a hexagonal lobule unit. Current approaches to use HLCs for an in vitro liver model, however, are mostly limited to 2D culture or simple 3D spheroid cultures (3,7,(10)(11)(12)(13)(14). The lack of a biomimetic microenvironment provided from the 3D interactions of parenchymal and nonparenchymal cell types along the hepatic differentiation stages may potentially be one of the limiting factors to functional maturation of the hepatic progenitor cells (HPCs), as well as the functional preservation of HLCs in vitro (15,16).…”

mentioning

confidence: 99%

See 2 more Smart Citations

Deterministically patterned biomimetic human iPSC-derived hepatic model via rapid 3D bioprinting

Zhu

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

744

661

View full text Add to dashboard Cite

The functional maturation and preservation of hepatic cells derived from human induced pluripotent stem cells (hiPSCs) are essential to personalized in vitro drug screening and disease study. Major liver functions are tightly linked to the 3D assembly of hepatocytes, with the supporting cell types from both endodermal and mesodermal origins in a hexagonal lobule unit. Although there are many reports on functional 2D cell differentiation, few studies have demonstrated the in vitro maturation of hiPSC-derived hepatic progenitor cells (hiPSC-HPCs) in a 3D environment that depicts the physiologically relevant cell combination and microarchitecture. The application of rapid, digital 3D bioprinting to tissue engineering has allowed 3D patterning of multiple cell types in a predefined biomimetic manner. Here we present a 3D hydrogel-based triculture model that embeds hiPSC-HPCs with human umbilical vein endothelial cells and adiposederived stem cells in a microscale hexagonal architecture. In comparison with 2D monolayer culture and a 3D HPC-only model, our 3D triculture model shows both phenotypic and functional enhancements in the hiPSC-HPCs over weeks of in vitro culture. Specifically, we find improved morphological organization, higher liver-specific gene expression levels, increased metabolic product secretion, and enhanced cytochrome P450 induction. The application of bioprinting technology in tissue engineering enables the development of a 3D biomimetic liver model that recapitulates the native liver module architecture and could be used for various applications such as early drug screening and disease modeling.3D bioprinting | in vitro hepatic model | iPSC | tissue engineering | biomaterials T he liver plays a critical role in the synthesis of important proteins and the metabolism of xenobiotic; the failure of these functions is closely related to disease development and drug-induced toxicity (1). For these reasons, in vitro liver models have been extensively developed to serve as platforms for pathophysiological studies and as alternatives to animal models in drug screening and hepatotoxicity prediction (2-4). Human primary hepatocytes, considered one of the most mature liver cell sources, lose many liver-specific functions rapidly when cultured in vitro due to the great discrepancies between the native and culture environments (5, 6). In addition, the practical difficulties in obtaining liver biopsy samples from every patient further hinder their use in personalized liver models. Consequently, hepatocytes derived from human induced pluripotent stem cells (hiPSCs), with the potential to be patient specific and easily accessible, have been widely acknowledged as the most promising cell source for developing personalized human hepatic models (4, 7).Many groups have reported monolayer differentiation of hiPSCs into hepatocyte-like cells (HLCs) and their ability to metabolize drugs (7-9). Nevertheless, hiPSC-derived HLCs are still considered immature in terms of many liver-specific gene expressions, functions, and...

show abstract

Section: Discussionmentioning

confidence: 99%

Section: D Hydrogel Encapsulation At Hepatic Progenitor Stage Demonsmentioning

confidence: 99%

mentioning

confidence: 99%

See 1 more Smart Citation

Deterministically patterned biomimetic human iPSC-derived hepatic model via rapid 3D bioprinting

Zhu

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

744

661

View full text Add to dashboard Cite

show abstract

“…Therefore, multicellular spheroids are considered useful in vitro systems for drug screening, including tests of drug metabolism and toxicity, [1][2][3][4][5][6] and for studying tumor metastasis. 7,8) Furthermore, such spheroids have attracted much attention for application in cell-based therapy [9][10][11] and as a building block for the construction of large tissues.…”

mentioning

confidence: 99%

Optimization of Albumin Secretion and Metabolic Activity of Cytochrome P450 1A1 of Human Hepatoblastoma HepG2 Cells in Multicellular Spheroids by Controlling Spheroid Size

Nishikawa

Tanaka

Nishikawa

et al. 2017

Biological & Pharmaceutical Bulletin

View full text Add to dashboard Cite

Key words albumin; CYP1A1; human hepatoblastoma HepG2 cell; size control; spheroid Three-dimensional multicellular spheroids possess greater in vivo tissue-like morphology and function than two-dimensional monolayered cells. Therefore, multicellular spheroids are considered useful in vitro systems for drug screening, including tests of drug metabolism and toxicity, [1][2][3][4][5][6] and for studying tumor metastasis. 7,8) Furthermore, such spheroids have attracted much attention for application in cell-based therapy [9][10][11] and as a building block for the construction of large tissues. 12,13) In a previous study, we demonstrated that spheroid formation greatly prolonged the survival of insulinsecreting NIT-1 cells after transplantation in mice. 9)Spheroid size is a key parameter to determine the functions and properties of spheroids. It is reported that oxygen concentration is decreased in the core of the spheroids of human hepatoblastoma HepG2 cells, hepatocytes, and tumor cells. [14][15][16][17] This affects the expression of various molecules, including hypoxia inducible factor (HIF)-1. In addition, paracellular or transcellular transport is required in order for the inner cells to take up extracellular materials and to release secretory products into the culture media. Despite their importance, details regarding the relationship between spheroid size and cellular functions remain to be elucidated. In particular, few studies have investigated the effect of spheroid sizes over 300 µm on their functions and properties.Investigating the effects of spheroid size requires the preparation of size-controlled, multicellular spheroids. Several techniques for spheroid formation have been reported, including the spinner flask, hanging drop, and liquid overlay methods. [18][19][20] However, a common problem with these techniques is the difficulty in controlling spheroid size. In contrast, microfabrication is suitable for the development of sizecontrolled microwells with narrow size distribution. 10,21,22) In a previous study, we demonstrated that polydimethylsiloxane (PDMS)-based microwells constructed using a microfabrication technique could be used to obtain spheroids with a very narrow size distribution. 23) We also found that coating the microwells with poly(N-isopropylacrylamide) (PNIPAAm), a thermoresponsive polymer, increased the quality of the spheroids, since the coating prevented the cells from adhering to the PDMS-based microwells. 23)In the present study, we developed PDMS-based microwells of different sizes in order to prepare variably sized multicellular spheroids. HepG2 cells were used for the study as they represent one of the most extensively used cell lines to evaluate drug metabolism and toxicity. 3,24,25) Production of various proteins is an index of liver function. Albumin is the most abundant serum protein produced by hepatocytes. 26)Another important liver function is the metabolism of various compounds, including drugs. CYP enzymes are the major enzymes involved in drug metabolism. CYP1A1 is ...

show abstract

“…While technological advancements are needed before this system can realize its potential in high-throughput applications, it has been effective in some lower throughput screening studies. Three-dimensional spheroid cultures using a Nanopillar Plate have already been used for drug toxicity using hepatocyte-like cells derived from human ESCs and human iPSCs [72]. Another example of developing 3D organoid systems for modeling is demonstrated by developing human gastric organoids (hGOs) [73].…”

Section: Figmentioning

confidence: 99%

Induced Pluripotent Stem Cell Models to Enable In Vitro Models for Screening in the Central Nervous System

Hunsberger

Efthymiou

Malik

et al. 2015

Stem Cells and Development

View full text Add to dashboard Cite

There is great need to develop more predictive drug discovery tools to identify new therapies to treat diseases of the central nervous system (CNS). Current nonpluripotent stem cell-based models often utilize non-CNS immortalized cell lines and do not enable the development of personalized models of disease. In this review, we discuss why in vitro models are necessary for translational research and outline the unique advantages of induced pluripotent stem cell (iPSC)-based models over those of current systems. We suggest that iPSC-based models can be patient specific and isogenic lines can be differentiated into many neural cell types for detailed comparisons. iPSC-derived cells can be combined to form small organoids, or large panels of lines can be developed that enable new forms of analysis. iPSC and embryonic stem cell-derived cells can be readily engineered to develop reporters for lineage studies or mechanism of action experiments further extending the utility of iPSC-based systems. We conclude by describing novel technologies that include strategies for the development of diversity panels, novel genomic engineering tools, new three-dimensional organoid systems, and modified high-content screens that may bring toxicology into the 21st century. The strategic integration of these technologies with the advantages of iPSC-derived cell technology, we believe, will be a paradigm shift for toxicology and drug discovery efforts.Disease Modeling D iseases of the central nervous system (CNS) affect a large number of people, but therapeutic intervention is hampered by the lack of useful models for many of these diseases. Current research on human subjects, particularly for drug discovery for CNS diseases, is severely (and appropriately) limited by ethical guidelines. Therefore, surrogate models are needed that share important anatomical, physiological, and genetic features to advance new treatments and therapies for CNS diseases [1].Developing rapid and effective therapies for CNS diseases requires the availability of in vitro models that accurately recapitulate disease phenotypes and predict patient treatment response. A proper model must be both sensitive and predictive while reflecting both normal and disease processes. Equally important, these models should enable the investigation of genetic and environmental risk factors contributing to diseases in a rapid and economical way. Currently used models often do not reflect a typical human response [2-4], despite efforts underway to better characterize these models and increase their preclinical value in predicting safety and efficacy in the clinic [5,6]. Therefore, there is a great need to develop disease-and patient-specific models from cells directly affected in CNS disorders. These cellbased models, we envision, could either replace or supplement current animal models and enable the efficient translation of basic research into the clinical setting. Limitations with current CNS modelsCurrently, drug discovery relies on the use of animalbased or cell-based models, ...

show abstract

3D spheroid culture of hESC/hiPSC-derived hepatocyte-like cells for drug toxicity testing

Cited by 251 publications

References 34 publications

Deterministically patterned biomimetic human iPSC-derived hepatic model via rapid 3D bioprinting

Deterministically patterned biomimetic human iPSC-derived hepatic model via rapid 3D bioprinting

Optimization of Albumin Secretion and Metabolic Activity of Cytochrome P450 1A1 of Human Hepatoblastoma HepG2 Cells in Multicellular Spheroids by Controlling Spheroid Size

Induced Pluripotent Stem Cell Models to Enable In Vitro Models for Screening in the Central Nervous System

Contact Info

Product

Resources

About