Mammalian Cell Viability Methods in 3D Scaffolds for Tissue Engineering

Gantenbein, Benjamin; Croft, Andreas S.; Larraillet, Marie

doi:10.5772/intechopen.93078

Cited by 7 publications

(6 citation statements)

References 69 publications

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“…25 Clinical data showed that the stiffness of native liver tissue increases drastically up to 4.1− 21.9 kPa during fibrosis due to aberrant ECM deposition, thus distorting angiogenesis and hepatic regeneration. 34 In our study, the compressive modulus was reduced in scaffolds possessing liver ECM compared to that in blank silk scaffolds and was similar to the healthy liver (1.5−4.5 kPa) (Figure 1H,I).…”

Section: Discussionsupporting

confidence: 68%

Mimicking Physiologically Relevant Hepatocyte Zonation Using Immunomodulatory Silk Liver Extracellular Matrix Scaffolds toward a Bioartificial Liver Platform

Janani

Mandal

2021

ACS Appl. Mater. Interfaces

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Mimicking nativelike metabolic zonation is indispensable to develop an efficient bioartificial liver model, as it facilitates physiological cues, hepatocyte polarity, and phenotypic functions. The present study shows the first evidence of hepatocyte metabolic heterogeneity in an in vitro liver model encompassing liver extracellular matrix (ECM)-functionalized silk scaffolds (LECM-SF) by altering ECM proportion. Upon static culture, individual LECM-SF scaffold supports differential synthetic and metabolic functions of cultured primary neonatal rat hepatocytes (PNRHs), owing to discrete biophysical attributes. A single in vitro liver system comprising PNRHs seeded LECM-SF scaffolds assisting periportal to pericentral gradient functions is stacked and matured in a perfusion bioreactor to simulate oxygen gradient. The scaffold with high ECM supports periportal-specific albumin synthesis, urea secretion, and bile duct formation, albeit scaffold with low ECM supports pericentral-specific cytochrome P450 activity. Extensive physicochemical characterizations confirmed the stability and interconnected porous network of scaffolds, signifying cellular infiltration and bidirectional nutrient diffusion. Furthermore, scaffolds demonstrate minimal thrombogenicity, reduced foreign-body response, and enhanced pro-remodeling macrophage activation, supporting constructive tissue remodeling. The developed liver model with zone-specific functions would be a promising avenue in bioartificial liver and drug screening.

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Section: Discussionsupporting

confidence: 68%

Mimicking Physiologically Relevant Hepatocyte Zonation Using Immunomodulatory Silk Liver Extracellular Matrix Scaffolds toward a Bioartificial Liver Platform

Janani

Mandal

2021

ACS Appl. Mater. Interfaces

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“…Cytocompatibility of the matrix is typically assessed using live-dead staining. 92,93 Longer term continuous culture and estimation of proliferation rates by assays such as bromodeoxyuridine (BrdU), along with morphological assessments benchmarked against well-established 3D scaffolds or physiological samples, is also used to verify the suitability of a synthetic scaffold for 3D cell culture. 92,94…”

Section: B Biopolymers As Monolithic Bioink Substratesmentioning

confidence: 99%

Design approaches for 3D cell culture and 3D bioprinting platforms

Sreepadmanabh,

Arun,

Bhattacharjee

2024

Biophysics Reviews

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The natural habitat of most cells consists of complex and disordered 3D microenvironments with spatiotemporally dynamic material properties. However, prevalent methods of in vitro culture study cells under poorly biomimetic 2D confinement or homogeneous conditions that often neglect critical topographical cues and mechanical stimuli. It has also become increasingly apparent that cells in a 3D conformation exhibit dramatically altered morphological and phenotypical states. In response, efforts toward designing biomaterial platforms for 3D cell culture have taken centerstage over the past few decades. Herein, we present a broad overview of biomaterials for 3D cell culture and 3D bioprinting, spanning both monolithic and granular systems. We first critically evaluate conventional monolithic hydrogel networks, with an emphasis on specific experimental requirements. Building on this, we document the recent emergence of microgel-based 3D growth media as a promising biomaterial platform enabling interrogation of cells within porous and granular scaffolds. We also explore how jammed microgel systems have been leveraged to spatially design and manipulate cellular structures using 3D bioprinting. The advent of these techniques heralds an unprecedented ability to experimentally model complex physiological niches, with important implications for tissue bioengineering and biomedical applications.

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“…Due to the impossibility of performing the Trypan Blue exclusion method on the printed cells, due to the hardening of the constructs by the crosslinking agent, the viability was evaluated using the LIVE/DEAD Viability Assay (ThermoFisher Scientific Inc., USA), which has already been used in different 3D bioprinting studies [52][53][54][55]. Constructs were incubated in 0.75 µM calcein AM (Invitrogen, Carlsbad, CA, USA) (green) and in 1 µM ethidium homodimer-1 (Invitrogen, Carlsbad, CA, USA) (red), dissolved in cold 1X PBS (Sigma-Aldrich, Milan, Italy).…”

Section: Viability Assaymentioning

confidence: 99%

Patients’ Stem Cells Differentiation in a 3D Environment as a Promising Experimental Tool for the Study of Amyotrophic Lateral Sclerosis

Scarian

Bordoni²,

Fantini

et al. 2022

IJMS

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Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease (NDD) that affects motor neurons, causing weakness, muscle atrophy and spasticity. Unfortunately, there are only symptomatic treatments available. Two important innovations in recent years are three-dimensional (3D) bioprinting and induced pluripotent stem cells (iPSCs). The aim of this work was to demonstrate the robustness of 3D cultures for the differentiation of stem cells for the study of ALS. We reprogrammed healthy and sALS peripheral blood mononuclear cells (PBMCs) in iPSCs and differentiated them in neural stem cells (NSCs) in 2D. NSCs were printed in 3D hydrogel-based constructs and subsequently differentiated first in motor neuron progenitors and finally in motor neurons. Every step of differentiation was tested for cell viability and characterized by confocal microscopy and RT-qPCR. Finally, we tested the electrophysiological characteristics of included NSC34. We found that NSCs maintained good viability during the 3D differentiation. Our results suggest that the hydrogel does not interfere with the correct differentiation process or with the electrophysiological features of the included cells. Such evidence confirmed that 3D bioprinting can be considered a good model for the study of ALS pathogenesis.

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Mammalian Cell Viability Methods in 3D Scaffolds for Tissue Engineering

Cited by 7 publications

References 69 publications

Mimicking Physiologically Relevant Hepatocyte Zonation Using Immunomodulatory Silk Liver Extracellular Matrix Scaffolds toward a Bioartificial Liver Platform

Mimicking Physiologically Relevant Hepatocyte Zonation Using Immunomodulatory Silk Liver Extracellular Matrix Scaffolds toward a Bioartificial Liver Platform

Design approaches for 3D cell culture and 3D bioprinting platforms

Patients’ Stem Cells Differentiation in a 3D Environment as a Promising Experimental Tool for the Study of Amyotrophic Lateral Sclerosis

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