Design of a novel bioink suitable for the 3D printing of lymphoid cells

Abstract

Introduction: For decades, in vitro 2D cell culture techniques have been employed in research, but they fail to recapitulate the complexity of natural tissues. 3D bioprinting could potentially overcome this drawback due to the possibility to control the spatial disposition of living cells and the geometry of the 3D scaffold.Materials and methods: This study reports the design and characterization of a novel bioink for extrusion bioprinting, analyzing different blend formulations composed of alginate, gelatin, and methylcellulose, suitable as cell-laden bioink for lymphoid cells, in particular those isolated from patients with Chronic Lymphocytic Leukemia (CLL). The rheological properties as a function of temperature and the printability of the formulations were investigated to define the optimal printing parameters. In vitro stability of the printed scaffolds was investigated under culture conditions and compression tests were performed on printed and bioprinted scaffolds to compare their mechanical properties with those of fresh lymphoid tissue. Finally, MEC1, a CLL cell line, was bioprinted to investigate cell viability, cell density, and cell capability to be released from the scaffold over time.Results and discussion: Results showed that, for the selected blends, good shape fidelity and printing accuracy were achieved with a limitation on the number of printed layers. Scaffolds withstood culture conditions showing stability for up to 3 weeks and their mechanical properties were similar to those of lymphoid tissues already reported in the literature. High cell viability after 21 days was observed for both MEC1 and primary peripheral mononuclear cells, confirming the possibility to use the selected formulation to successfully bioprint lymphoid cells by possibly mimicking their native lymphoid microenvironment.

Shaping Synthetic Multicellular and Complex Multimaterial Tissues via Embedded Extrusion‐Volumetric Printing of Microgels

“…The second bioink was designed to allow for the delivery of high cell densities with good shape fidelity, even in absence of stably incorporated hydrogel carriers. For this purpose, we produced a bioink based on methylcellulose (MC), due to its ability to both modulate the viscosity of the cell suspension, [62][63][64] and rapidly dissolve post printing in an aqueous environment.…”

Crosslinking strategies in modulating methylcellulose hydrogel properties

“…Owing to the reversibility of the fibril formation process during MC physical gelation, MC hydrogels possess some distinctive properties, e.g., reversibility of the sol-gel transition, reduced cytotoxicity (due to the absence of chemical crosslinkers), and low costs. 1,17 In this regard, the reversible nature of the physical gelation process can be smartly exploited, e.g., when MC is used as a ''nongelling'' -or a sacrificial -component in different bioprinting approaches, [18][19][20][21] and its selective dissolution is achieved simply by lowering the temperature.…”

“…In this regard, the reversible nature of the physical gelation process can be smartly exploited, e.g. , when MC is used as a “non-gelling” – or a sacrificial – component in different bioprinting approaches, 18–21 and its selective dissolution is achieved simply by lowering the temperature.…”

Mechanical Properties Affect Primary T Cell Activation in 3D Bioprinted Hydrogels

“…In 3D bioprinting, cells or molecules are directly printed in a layer-by-layer fashion in a set pattern such that the cells hold together to form the required 3D construct . 3D bioprinted matrices prepared using natural polymers, such as alginate, gelatin, chitosan, silk, hyaluronic acid, etc., provide a hydrated environment that mimics the physical and mechanical properties of the extracellular matrices. − Furthermore, some natural polymers like gelatin contain cell-binding sites that confer better biocompatibility and enable cell proliferation or differentiation . The technology also has other advantages, such as spatial control of multicellular patterning, tunable cellular densities, reproducibility, agility, short work time, and precise control of geometries .…”

“…Although several reports have demonstrated the use of 3D bioprinting in fabricating tissue scaffolds, there are only a few reports on the bioprinting of immune cells and organs. ,, Furthermore, to the best of our knowledge, there are no reports of ex vivo tissue-mimetic constructs of immune cells by using DLP-based printing. One of the first reports of immune cell bioprinting was of human regulatory T cells in alginate–gelatin methacryloyl hydrogel using extrusion-based bioprinting .…”