Co-culture system of hepatocytes and endothelial cells: two in vitro approaches for enhancing liver-specific functions of hepatocytes

Wang, Gaoxiong; Zheng, Youshi; Wang, Yingchao; Cai, Zhixiong; Liao, Naishun; Liu, Jingfeng; Zhang, Wenmin

doi:10.1007/s10616-018-0219-3

Cited by 12 publications

(12 citation statements)

References 24 publications

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“…Thus, we evaluated the effects of iEndo and 3D culture on maturation of iHep by developing a 3D w/ iEndo model in which an endothelial layer was interconnected with a hydrogel-based 3D hepatic model. Markers of hepatocyte function, such as albumin and urea secretion and CYP3A4 expression, were significantly higher into the gel in the presence of iEndo, as reported by others. ,− Consequently, we observed that a 3D environment and cell–cell interactions have been crucial also in our model for maintaining hepatocyte viability and function.…”

Section: Discussionsupporting

confidence: 85%

“…iHep and iEndo were co-cultured in commercial 12-well Transwell inserts (Greiner Bio-One, 665641) having a PET membrane with a pore diameter of 0.4 μm, density of 2 × 10 6 pores/cm 2 . The iHep to iEndo cell ratio (4:1) was close to what was observed in vivo …”

Section: Methodssupporting

confidence: 86%

“…The iHep to iEndo cell ratio (4:1) was close to what was observed in vivo. 12 iEndo were plated on the bottom side of the Transwell-like insert membrane pre-coated with Fibronectin (Promocell) according to the manufacturer's protocol. Afterward, the 12-well plates were placed in the incubator upside down to allow the cells to firmly attach to the microporous membrane.…”

Section: ■ Introductionmentioning

confidence: 99%

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Development of an Induced Pluripotent Stem Cell-Based Liver-on-a-Chip Assessed with an Alzheimer’s Disease Drug

Fanizza

Boeri

Donnaloja

et al. 2023

ACS Biomater. Sci. Eng.

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Liver-related drug metabolism is a key aspect of pharmacokinetics and possible toxicity. From this perspective, the availability of advanced in vitro models for drug testing is still an open need, also to the end of reducing the burden of in vivo experiments. In this scenario, organ-on-a-chip is gaining attention as it couples a state-of-the art in vitro approach to the recapitulation of key in vivo physiological features such as fluidodynamics and a tri-dimensional cytoarchitecture. We implemented a novel liver-on-a-chip (LoC) device based on an innovative dynamic device (MINERVA 2.0) where functional hepatocytes (iHep) have been encapsulated into a 3D hydrogel matrix interfaced through a porous membrane with endothelial cells (iEndo)]. Both lines were derived from human-induced pluripotent stem cells (iPSCs), and the LoC was functionally assessed with donepezil, a drug approved for Alzheimer’s disease therapy. The presence of iEndo and a 3D microenvironment enhanced the expression of liver-specific physiologic functions as in iHep, after 7 day perfusion, we noticed an increase of albumin, urea production, and cytochrome CYP3A4 expression compared to the iHep static culture. In particular, for donepezil kinetics, a computational fluid dynamic study conducted to assess the amount of donepezil diffused into the LoC indicated that the molecule should be able to pass through the iEndo and reach the target iHep construct. Then, we performed experiments of donepezil kinetics that confirmed the numerical simulations. Overall, our iPSC-based LoC reproduced the in vivo physiological microenvironment of the liver and was suitable for potential hepatotoxic screening studies.

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Section: Discussionsupporting

confidence: 85%

Section: Methodssupporting

confidence: 86%

Section: ■ Introductionmentioning

confidence: 99%

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Development of an Induced Pluripotent Stem Cell-Based Liver-on-a-Chip Assessed with an Alzheimer’s Disease Drug

Fanizza

Boeri

Donnaloja

et al. 2023

ACS Biomater. Sci. Eng.

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“…Urea levels in cocultures exhibited the opposite trend of the levels in monocultures, as there was a continuous increase in co-cultures over time ( Figure 4C). Our results are in accordance with those of a previous study (Table 1) that reported higher levels of urea and albumin secretion in hepatocytes co-cultured with endothelial cells, HSCs, and adipose cells than in a monoculture (Wang et al, 2018). Another study that examined the co-culture of HepaRG cells (cell line), LX2 cells, HUVECs, and monocytes (Table 1) also reported more urea secretion in the co-culture than in a monoculture (Rennert et al, 2015).…”

Section: Figuresupporting

confidence: 92%

“…Hepatocytes are co-cultured with an HSC line (e.g., LX2 cells), umbilical cord-mesenchymal stem cells (UC-MSCs), and umbilical cord blood (UCB)-CD34 hematopoietic stem/progenitor cells with the aim of producing a microenvironment matching the liver's in vivo microenvironment. A previous study reported the formation of a vessel-like structure in a co-culture of hepatocytes with endothelial cells and HSCs, with increased urea and albumin secretion as well as Cyp3A4 expression (Wang et al, 2018). Meanwhile, in a co-culture of MSCs and an endothelial cell line (HUVEC EA.…”

Section: Introductionmentioning

confidence: 93%

3D Co-Culture of Hepatocyte, a Hepatic Stellate Cell Line, and Stem Cells for Developing a Bioartificial Liver Prototype

Sibuea¹,

Pawitan²,

Antarianto³

et al. 2020

IJTech

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A liver organoid is an in vitro reconstruction of the liver that mimics the in vivo liver microstructure and performs liver functions. Liver organoids can be used for drug testing, as a model of liver disease pathogenesis, and as a bioartificial liver prototype material to develop promising alternative therapies for liver failure. In this study, we reconstructed liver organoids using primary rat hepatocytes, a hepatic stellate cell line (LX2), human umbilical cord-mesenchymal stem cells (UC-MSCs), and human umbilical cord blood (UCB)-CD34+ hematopoietic stem/progenitor cells. Suspensions of primary rat hepatocytes, LX2 cells, UC-MSCs, and UCB-CD34+ cells were co-cultured using 11 ratio sets, and spheroid formation was evaluated for 2 days. Ratio sets with a positive liver organoid appearance were cultured in four different culture media, and after they were harvested, their viability was compared with that of a hepatocyte monoculture. Liver organoids were further analyzed for 14 days to assess albumin and urea production as well as relative gene expression. We found that a 5:1:2:2 cellular density ratio of hepatocytes:LX2 cells:UC-MSCs:UCB-CD34+ cells, respectively, and Williams E medium supplemented with platelet lysate, ITS, and dexamethasone were the most suitable conditions for liver organoid reconstruction. Expression of the albumin and GPT1 genes and CD31 in the liver organoid increased until day 14, while urea secretion increased until day 5. Liver organoids reconstructed through the 3D co-culture of primary rat hepatocytes, LX2 cells, UC-MSCs, and UCB-CD34+ cells at a specific cellular ratio using an economical medium with a simple composition maintained their functions until day 14. As a future direction, these organoids can be used to develop a bioartificial liver.

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Self‐Healing Hydrogel Containing Decellularized Liver Matrix and Endothelial Cell‐Covered Hepatocyte Spheroids for Rescue of Injured Hepatocytes

Chou,

Cheng,

Yin

et al. 2024

Macromolecular Bioscience

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Liver fibrosis occurs in many chronic liver diseases, while severe fibrosis can lead to liver failure. A chitosan‐phenol based self‐healing hydrogel (CP) integrated with decellularized liver matrix (DLM) is proposed in this study as a 3D gel matrix to carry hepatocytes for possible therapy of liver fibrosis. To mimic the physiological liver microenvironment, DLM is extracted from pigs and mixed with CP hydrogel to generate DLM‐CP self‐healing hydrogel. Hepatocyte spheroids coated with endothelial cells (ECs) are fabricated using a customized method and embedded in the hydrogel. Hepatocytes injured by exposure to CCl4‐containing medium are used as the in vitro toxin‐mediated liver fibrosis model, where the EC‐covered hepatocyte spheroids embedded in the hydrogel are co‐cultured with the injured hepatocytes. The urea synthesis of the injured hepatocytes reaches 91% of the normal level after 7 days of co‐culture, indicating that the hepatic function of injured hepatocytes is rescued by the hybrid spheroid‐laden DLM‐CP hydrogel. Moreover, the relative lactate dehydrogenase activity of the injured hepatocytes is decreased 49% by the hybrid spheroid‐laden DLM‐CP hydrogel after 7 days of co‐culture, suggesting reduced damage in the injured hepatocytes. The combination of hepatocyte/EC hybrid spheroids and DLM‐CP hydrogel presents a promising therapeutic strategy for hepatic fibrosis.

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Co-culture system of hepatocytes and endothelial cells: two in vitro approaches for enhancing liver-specific functions of hepatocytes

Cited by 12 publications

References 24 publications

Development of an Induced Pluripotent Stem Cell-Based Liver-on-a-Chip Assessed with an Alzheimer’s Disease Drug

Development of an Induced Pluripotent Stem Cell-Based Liver-on-a-Chip Assessed with an Alzheimer’s Disease Drug

3D Co-Culture of Hepatocyte, a Hepatic Stellate Cell Line, and Stem Cells for Developing a Bioartificial Liver Prototype

Self‐Healing Hydrogel Containing Decellularized Liver Matrix and Endothelial Cell‐Covered Hepatocyte Spheroids for Rescue of Injured Hepatocytes

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