Generation of Hepatic Tissue Structures Using Multicellular Spheroid Culture

Tao, Fumiya; Mihara, Hirotaka; Kojima, Naoya

doi:10.1007/978-1-4939-8961-4_14

Cited by 7 publications

(7 citation statements)

References 7 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Spheroids were produced using a method described in a previous report [17] and protocol [18]. Two ml of the MC medium was poured into a 35 mm Petri dish using Microman (Gilson, Middleton, WI, USA), because of the high viscosity of the MC medium.…”

Section: Methodsmentioning

confidence: 99%

Improved Oxygen Supply to Multicellular Spheroids Using A Gas-permeable Plate and Embedded Hydrogel Beads

Mihara

Kugawa

Sayo

et al. 2019

Cells

Self Cite

View full text Add to dashboard Cite

Culture systems for three-dimensional tissues, such as multicellular spheroids, are indispensable for high-throughput screening of primary or patient-derived xenograft (PDX)-expanded cancer tissues. Oxygen supply to the center of such spheroids is particularly critical for maintaining cellular functions as well as avoiding the development of a necrotic core. In this study, we evaluated two methods to enhance oxygen supply: (1) using a culture plate with a gas-permeable polydimethylsiloxane (PDMS) membrane on the bottom, and; (2) embedding hydrogel beads in the spheroids. Culturing spheroids on PDMS increased cell growth and affected glucose/lactate metabolism and CYP3A4 mRNA expression and subsequent enzyme activity. The spheroids, comprised of 5000 Hep G2 cells and 5000 20 µm-diameter hydrogel beads, did not develop a necrotic core for nine days when cultured on a gas-permeable sheet. In contrast, central necrosis in spheroids lacking hydrogel beads was observed after day 3 of culture, even when using PDMS. These results indicate that the combination of gas-permeable culture equipment and embedded hydrogel beads improves culture 3D spheroids produced from primary or PDX-expanded tumor cells.

show abstract

Section: Methodsmentioning

confidence: 99%

Improved Oxygen Supply to Multicellular Spheroids Using A Gas-permeable Plate and Embedded Hydrogel Beads

Mihara

Kugawa

Sayo

et al. 2019

Cells

Self Cite

View full text Add to dashboard Cite

show abstract

“…We previously established a rapid aggregation system that allows cell aggregation cells by using 3% methylcellulose (MC; Sigma) medium 60 . MC medium was poured into a dish with a positive-displacement pipette (Gilson), because 3% MC medium is highly viscous.…”

Section: Generation Of Spheroids Containing Cells and Ecm Components/mentioning

confidence: 99%

Development of a tunable method to generate various three-dimensional microstructures by replenishing macromolecules such as extracellular matrix components and polysaccharides

Tao

Sayo

Sugimoto

et al. 2020

Sci Rep

Self Cite

View full text Add to dashboard Cite

Multicellular spheroids (spheroids) are expected to be a promising approach to mimic in vivo organ functions and cell microenvironments. However, conventional spheroids do not fully consider the existence of extracellular matrices (ecMs). in this study, we developed a tunable method for replenishing macromolecules, including ecM components and polysaccharides, into spheroids without compromising cell viability by injecting a microvolume cell suspension into a high density of methylcellulose dissolved in the culture medium. Adjusting the ecM concentration in the cell suspension enabled the generation of different three-dimensional microstructures, such as "ECM gel capsules", which contained individually separated cells, and "ECM-loaded spheroids", which had thin ECM layers between cells. ECM-loaded spheroids with a 30-fold dilution of Matrigel (0.3 mg/ml) showed significantly higher albumin secretion than control spheroids composed of Hep G2 or HuH-7 cells. Additionally, the expression levels of major cYp genes were decreased in ecM gel capsules with undiluted Matrigel (9 mg/ml) compared to those in control spheroids. However, 0.3 mg/ml Matrigel did not disrupt gene expression. furthermore, cell polarity associated with tight junction proteins (ZO-1 and Claudin-1) and the transporter protein MRP2 was markedly induced by using 0.3 mg/ml Matrigel. thus, high-performance three-dimensional tissues fabricated by this method are applicable to increasing the efficiency of drug screening and to regenerative medicine.

show abstract

“…Spheroids were produced using a method described in a previous report [16] and protocol [17]. cell suspension (1×10 7 cells/ml) and hydrogel beads (1×10 7 beads/ml) in growth medium to produce a suspension of 5×10 6 cells/ml and 5×10 6 beads/ml.…”

Section: Spheroid Productionmentioning

confidence: 99%

Improvement of Oxygen Supply to the Multicellular Spheroids Using a Gas-Permeable Plate and Embedded Hydrogel Beads

Mihara¹,

Kugawa²,

Sayo³

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

Culture systems for 3-dimensional tissues, such as multicellular spheroids, are indispensable for high-throughput screening of primary or patient-derived xenograft (PDX)-expanded cancer tissues. Oxygen supply to the center of such spheroids is particularly critical for maintaining cellular functions as well as avoiding the development of a necrotic core. In this study, we evaluated 2 methods to enhance oxygen supply: (1) using culture plate with gas-permeable polydimethylsiloxane (PDMS) membrane at its bottom and (2) embedding hydrogel beads in the spheroids. Culturing spheroids on PDMS increased cell growth and affected glucose/lactate metabolism and CYP3A4 mRNA expression and subsequent enzyme activity. The spheroids comprised 5000 Hep G2 cells and 5000 20 µm-diameter hydrogel beads did not develop a necrotic core for 9 days when cultured on a gas-permeable sheet. In contrast, central necrosis in spheroids lacking hydrogel beads was observed after day 3 of culture, even when using PDMS. These results indicate that the combination of gas-permeable culture equipment and embedded hydrogel beads improves culture 3D spheroids produced from primary or PDX-expanded tumor cells.

show abstract

Generation of Hepatic Tissue Structures Using Multicellular Spheroid Culture

Cited by 7 publications

References 7 publications

Improved Oxygen Supply to Multicellular Spheroids Using A Gas-permeable Plate and Embedded Hydrogel Beads

Improved Oxygen Supply to Multicellular Spheroids Using A Gas-permeable Plate and Embedded Hydrogel Beads

Development of a tunable method to generate various three-dimensional microstructures by replenishing macromolecules such as extracellular matrix components and polysaccharides

Improvement of Oxygen Supply to the Multicellular Spheroids Using a Gas-Permeable Plate and Embedded Hydrogel Beads

Contact Info

Product

Resources

About