Bioprinting of three-dimensional culture models and organ-on-a-chip systems

Multimaterial bioprinting technologies offer promising avenues to create mini-organ models with enhanced tissue heterogeneity and complexity. This article focuses on the application of three-dimensional bioprinting to fabricate organ-on-a-chip systems for in vitro drug testing and screening. We illustrate the capabilities and limitations of a bioprinting approach compared to microfabrication in constructing an organ-on-a-chip device. Further, we propose strategies in multimaterial integration for printing microphysiological tissue models. With these analyses, key challenges and future directions are highlighted.D.Z. gratefully acknowledges support from the China Scholarship Council. Y.L. acknowledges the Schlumberger Foundation and Trinity College Cambridge. Y.Y.S.H. thanks EPSRC, the Royal Society London, and the Isaac Newton Trust for funding support

Rapid patterning of 1-D collagenous topography as an ECM protein fibril platform for image cytometry.

Cellular behavior is strongly influenced by the architecture and pattern of its interfacing extracellular matrix (ECM). For an artificial culture system which could eventually benefit the translation of scientific findings into therapeutic development, the system should capture the key characteristics of a physiological microenvironment. At the same time, it should also enable standardized, high throughput data acquisition. Since an ECM is composed of different fibrous proteins, studying cellular interaction with individual fibrils will be of physiological relevance. In this study, we employ near-field electrospinning to create ordered patterns of collagenous fibrils of gelatin, based on an acetic acid and ethyl acetate aqueous co-solvent system. Tunable conformations of micro-fibrils were directly deposited onto soft polymeric substrates in a single step. We observe that global topographical features of straight lines, beads-on-strings, and curls are dictated by solution conductivity; whereas the finer details such as the fiber cross-sectional profile are tuned by solution viscosity. Using these fibril constructs as cellular assays, we study EA.hy926 endothelial cells' response to ROCK inhibition, because of ROCK's key role in the regulation of cell shape. The fibril array was shown to modulate the cellular morphology towards a pre-capillary cord-like phenotype, which was otherwise not observed on a flat 2-D substrate. Further facilitated by quantitative analysis of morphological parameters, the fibril platform also provides better dissection in the cells' response to a H1152 ROCK inhibitor. In conclusion, the near-field electrospun fibril constructs provide a more physiologically-relevant platform compared to a featureless 2-D surface, and simultaneously permit statistical single-cell image cytometry using conventional microscopy systems. The patterning approach described here is also expected to form the basics for depositing other protein fibrils, seen among potential applications as culture platforms for drug screening

Low-Voltage Continuous Electrospinning Patterning

Electrospinning is a versatile technique for the construction of microfibrous and nanofibrous structures with considerable potential in applications ranging from textile manufacturing to tissue engineering scaffolds. In the simplest form, electrospinning uses a high voltage of tens of thousands volts to draw out ultrafine polymer fibers over a large distance. However, the high voltage limits the flexible combination of material selection, deposition substrate, and control of patterns. Prior studies show that by performing electrospinning with a well-defined "near-field" condition, the operation voltage can be decreased to the kilovolt range, and further enable more precise patterning of fibril structures on a planar surface. In this work, by using solution dependent "initiators", we demonstrate a further lowering of voltage with an ultralow voltage continuous electrospinning patterning (LEP) technique, which reduces the applied voltage threshold to as low as 50 V, simultaneously permitting direct fiber patterning. The versatility of LEP is shown using a wide range of combination of polymer and solvent systems for thermoplastics and biopolymers. Novel functionalities are also incorporated when a low voltage mode is used in place of a high voltage mode, such as direct printing of living bacteria; the construction of suspended single fibers and membrane networks. The LEP technique reported here should open up new avenues in the patterning of bioelements and free-form nano- to microscale fibrous structures.Studentship and scholarship funding supports from the China Scholarship Council scholarship, EPSRC doctoral training partnership, Schlumberger Foundation, WD Armstrong Trus

Harnessing Surface-Functionalized Metal-Organic Frameworks for Selective Tumor Cell Capture

A platform based on a metal-organic framework (MOF) bearing free carboxylic acid groups has been developed for tumor cell capture and potential drug screening applications. A zinc-based MOF expressing uncoordinated carboxylic acids (ZnMOF-COOH) was grown on a ZnO substrate. Post-synthetic modification (PSM) of the acid groups gave a composite material that expressed peptide linkages and allowed the immobilization of anti-epithelial cell adhesion molecule (anti-EpCAM) antibody. This strategy offers a universal method for the controllable immobilization of antibodies and even enzymes on the surface of a MOF. The resulting immunotrapper exhibited excellent capture ability, demonstrating high efficiency and selectivity towards EpCAM-positive tumor cells. The promotion of tumor cell adhesion is attributed to the 3-dimentional (3D) structure of the composite, which revealed spine-like microstructures.This work was financially supported by the National Natural Science Foundation of China (Grant Nos. 21527809 and 21575007), the China Scholarship Council and the UK EPSRC (EP/J500380/1)

Near-Field Electrospinning Patterning Polycaprolactone and Polycaprolactone/Collagen Interconnected Fiber Membrane

Solution-based near-field electrospinning is employed to construct polymeric network membranes, made of orderly arranged and interconnected fibers. The narrow tip-to-nozzle separation of the direct-writing process leads to solvent enriched fibers being deposited on the substrate, despite the use of a low boiling point solvent. This results in fibers with low cross-sectional aspect ratio (flattened appearance), but providing a unique opportunity to produce interconnected fiber junctions through in situ, localized solvent etching by subsequent fiber overlays. Orthogonal networks of polycaprolactone (PCL) fibres, or PCL/collagen composite fibres, are fabricated, and then character-ized by microscopy and spectroscopy techniques. This study presents a direct approach to strengthen interfiber junctions, and further the feasibility to inter-weave and interconnect fibers of different properties, leading to networked membranes with potentially tailorable functions for tissue engineering applications and beyond

Bio‐assembling Macro‐Scale, Lumenized Airway Tubes of Defined Shape via Multi‐Organoid Patterning and Fusion

Abstract: Epithelial, stem‐cell derived organoids are ideal building blocks for tissue engineering, however, scalable and shape‐controlled bio‐assembly of epithelial organoids into larger and anatomical structures is yet to be achieved. Here, a robust organoid engineering approach, Multi‐Organoid Patterning and Fusion (MOrPF), is presented to assemble individual airway organoids of different sizes into upscaled, scaffold‐free airway tubes with predefined shapes. Multi‐Organoid Aggregates (MOAs) undergo accelerated fusion in a matrix‐depleted, free‐floating environment, possess a continuous lumen, and maintain prescribed shapes without an exogenous scaffold interface. MOAs in the floating culture exhibit a well‐defined three‐stage process of inter‐organoid surface integration, luminal material clearance, and lumina connection. The observed shape stability of patterned MOAs is confirmed by theoretical modelling based on organoid morphology and the physical forces involved in organoid fusion. Immunofluorescent characterization shows that fused MOA tubes possess an unstratified epithelium consisting mainly of tracheal basal stem cells. By generating large, shape‐controllable organ tubes, MOrPF enables upscaled organoid engineering towards integrated organoid devices and structurally complex organ tubes

Transparent nanostructured electrode by centrifuge coating

We employ a new solution-based coating process, centrifuge coating, to fabricate nanostructured conductive layers over large areas. This coating procedure allows fast quenching of the metastable dispersed state of nanomaterials, which minimizes material wastes by mitigate the effects of particle re-aggregation. Using this method, we fabricate SWNT coatings on different substrates such as PET (polyethylene terephthalate), PDMS (polydimethylsiloxane), and an acrylic elastomer. The effects of the choice of solvents on the morphology and subsequent performance of the coating network are studied. © 2012 IEEE

Centrifuge coating for low-waste solution processing of transparent nanostructured electrodes

Centrifuge coating was implemented to fabricate nanostructured conductive layers through solution processing at room temperature. This coating procedure allows fast evaporation, thereby fixing the nanomaterials in their dispersed state onto a substrate by the centrifuge action. Material wastes were minimized by mitigating the effects of particle reaggregation. Using this method, we fabricate single-wall nanotube coatings on different substrates such as polyethylene terephthalate, polydimethylsiloxane, and an acrylic elastomer with no prior surface modification of the substrate. The effects of the choice of solvents on the morphology and subsequent performance of the coating network are studied. © 2002-2012 IEEE

Bioprinting of three-dimensional culture models and organ-on-a-chip systems

Multimaterial bioprinting technologies offer promising avenues to create mini-organ models with enhanced tissue heterogeneity and complexity. This article focuses on the application of three-dimensional bioprinting to fabricate organ-on-a-chip systems for in vitro drug testing and screening. We illustrate the capabilities and limitations of a bioprinting approach compared to microfabrication in constructing an organ-on-a-chip device. Further, we propose strategies in multimaterial integration for printing microphysiological tissue models. With these analyses, key challenges and future directions are highlighted

Low-Voltage Continuous Electrospinning: A Versatile Protocol for Patterning Nano- and Micro-Scaled Fibers for Cell Interface.

Nano- and micro-scaled fibers have been incorporated in a number of applications in biofabrication and tissue cultures, providing a cell interfacing structure with extracellular matrix-mimicking topography and adhesion sites, and further supporting localized drug release. Here, we describe the low-voltage electrospinning patterning (LEP) protocol, which allows direct and continuous patterning of sub-micron fibers in a controlled fashion. The processable polymers range from protein (e.g., gelatin) to thermoplastic (e.g., polystyrene) polymers, with flexible selections of collecting substrates. The operation voltage for fiber fabrication can be as low as 50 V, which brings the benefits of reducing costs and mild-processing