Rat multicellular 3D liver microtissues to explore TGF-β1 induced effects

Prestigiacomo, Vincenzo; Weston, Anna; Suter‐Dick, Laura

doi:10.1016/j.vascn.2019.106650

Cited by 10 publications

(8 citation statements)

References 21 publications

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“…For the stellate cells model, we present, as already described by Norona et al, the maintenance for several weeks, of the LX-2 cell line (a cell line obtained from activated human primary stellate cells), in a "quiescent"-like phenotype. Unlike LX-2 cultured in 2D, the expression and synthesis of two markers of the activated HSC phenotype, respectively ACTA2 and type I collagen, are strongly inhibited when the cells are embedded in a GelMa matrix, confirming the capacity of a flexible matrix to support the quiescence of stellate cells in vitro (Norona et al, 2016;Prestigiacomo et al, 2020;Sohara et al, 2002).…”

Section: J O U R N a L P R E -P R O O Fmentioning

confidence: 98%

“…Models where different liver cell types are co-cultured in 3D partially recreate the complex interactions between parenchymal and non-parenchymal cells, allowing to explore physiological pathways in a context closer to that of in vivo organs (Leite et al, 2016;Messner et al, 2013;Prestigiacomo et al, 2017Prestigiacomo et al, , 2020.…”

Section: J O U R N a L P R E -P R O O Fmentioning

confidence: 99%

“…They become proliferating, contractile and activate tissue healing by J o u r n a l P r e -p r o o f synthesizing components of the extracellular matrix (ECM) such as type I collagen. The in vitro study of mechanisms of development of hepatic fibrosis such as activation and matrix remodeling by stellate cells requires the establishment of cocultures between, inter alia, hepatic cells and hepatocytes (Loréal et al, 1993;Prestigiacomo et al, 2017Prestigiacomo et al, , 2020Théret et al, 1997). However, the culture of stellate cells remains today a technological obstacle in obtaining relevant hepatic fibrosis models.…”

Section: Introductionmentioning

confidence: 99%

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3D culture of HepaRG cells in GelMa and its application to bioprinting of a multicellular hepatic model

et al. 2021

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Bioprinting is an emergent technology that has already demonstrated the capacity to create complex and/or vascularized multicellular structures with defined and organized architectures, in a reproducible and high throughput way. Here, we present the implementation of a complex liver model by the development of a three-dimensional extrusion bioprinting process, including parameters for matrix polymerization of methacrylated gelatin, using two hepatic cell lines, Huh7 and HepaRG. The printed structures exhibited long-term viability (28 days), proliferative ability, a relevant hepatocyte phenotype and functions equivalent to or better than those of their 2D counterparts using standard DMSO treatment.This work served as a basis for the bioprinting of complex multicellular models associating the hepatic parenchymal cells, HepaRG, with stellate cells (LX-2) and endothelial cells (HUVECs), able of colonizing the surface of the structure and thus recreating a pseudo endothelial barrier. When bioprinted in 3D monocultures, LX-2 expression was modulated by TGF-β-1 toward the induction of myofibroblastic genes such as ACTA2 and COL1A1. In 3D multicellular bioprinted structures comprising HepaRG, LX-2 and endothelial cells, we evidenced fibrillar collagen deposition, which is never observed in monocultures of either HepaRG or LX-2 alone. These observations indicate that a precise control of cellular communication is required to recapitulate key steps of fibrogenesis. Bioprinted 3D co-cultures therefore open up new perspectives in studying the molecular and cellular basis of fibrosis development and provide better access to potential inducers and inhibitors of collagen expression and deposition.

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Section: J O U R N a L P R E -P R O O Fmentioning

confidence: 98%

Section: J O U R N a L P R E -P R O O Fmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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3D culture of HepaRG cells in GelMa and its application to bioprinting of a multicellular hepatic model

et al. 2021

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“…Thus it is essential to elucidate the mechanisms controlling HSC activation. TGF-β1 is a canonical HSC activator after liver injury (Prestigiacomo et al 2020 ). In the present study, we found that miR-223 was highly expressed in NK-Exo and inhibition of miR-223 expression in NK-Exo significantly abrogated the TGF-β1-induced HSC activation, as demonstrated by suppressed proliferation and decreased expression of α-SMA and CoL1A1 in human HSC-LX-2 cells.…”

Section: Discussionmentioning

confidence: 99%

Exosomal miR-223 derived from natural killer cells inhibits hepatic stellate cell activation by suppressing autophagy

Wang

Quan

2020

Mol Med

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Background: Activation of hepatic stellate cells (HSCs) is a prominent driver of liver fibrosis. We previously demonstrated that exosomes derived from natural killer (NK) cells (NK-Exo) attenuated TGF-β1-induced HSC activation. Herein, this study was designed to investigate the mechanism underlying the action of NK-Exo. Methods: NK-Exo was isolated from NK-92MI cells and then administered into TGF-β1-treated LX-2 (human HSC line) cells. MiR-223 expression in NK-Exo was downregulated by transfecting NK-92MI cells with miR-223 inhibitor followed by exosome isolation. The HSC activation was evaluated by determining cell proliferation using CCK-8 assay and measuring the protein levels of α-SMA and CoL1A1 using western blot in LX-2 cells. The expression of miR-223 was detected by qRT-PCR. The interaction between miR-223 and ATG7 was analyzed by a dual-luciferase activity assay. The autophagy was evaluated by measuring the autophagy-related proteins using western blot. Results: miR-223 was highly expressed in NK-Exo and inhibition of miR-223 expression in NK-Exo abrogated the inhibitory effect of NK-Exo on TGF-β-induced HSC activation. ATG7 was confirmed as a direct target of miR-223. Furthermore, treatment with the autophagy activator rapamycin and ATG7 overexpression in LX-2 cells abolished the HSC activation-suppressive effect of NK-Exo. Conclusion: NK-Exo attenuated TGF-β-induced HSC activation by transferring miR-223 that inhibited autophagy via targeting ATG7.

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“…Under gravity, cells slowly settle to the lowest point of the drop, aggregating and forming spheroids ( Figure 1a). Coculture of different cell types and size control of the spheroids are easily achievable by varying the content and volume of the cell solution (Prestigiacomo et al, 2020).…”

Section: Hanging Dropmentioning

confidence: 99%

A review of manufacturing capabilities of cell spheroid generation technologies and future development

Liú

Chen

Naing

2020

Biotech & Bioengineering

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Spheroid culture provides cells with a three‐dimensional environment that can better mimic physiological conditions compared to monolayer culture. Technologies involved in the generation of cell spheroids are continuously being innovated to produce spheroids with enhanced properties. In this paper, we review the manufacturing capabilities of current cell spheroid generation technologies. We propose that spheroid generation technologies should enable tight and robust process controls to produce spheroids of consistent and repeatable quality. Future technology development for the generation of cell spheroids should look into improvement in process control, standardization, scalability and monitoring, in addition to advanced methods of spheroid transfer and characterization.

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Rat multicellular 3D liver microtissues to explore TGF-β1 induced effects

Cited by 10 publications

References 21 publications

3D culture of HepaRG cells in GelMa and its application to bioprinting of a multicellular hepatic model

3D culture of HepaRG cells in GelMa and its application to bioprinting of a multicellular hepatic model

Exosomal miR-223 derived from natural killer cells inhibits hepatic stellate cell activation by suppressing autophagy

A review of manufacturing capabilities of cell spheroid generation technologies and future development

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