3D printed microfluidics: advances in strategies, integration, and applications.

The ability to construct multiplexed micro-systems for fluid regulation could substantially impact multiple fields, including chemistry, biology, biomedicine, tissue engineering, and soft robotics, among others. 3D printing is gaining traction as a compelling approach to fabricating microfluidic devices by providing unique capabilities, such as 1) rapid design iteration and prototyping, 2) the potential for automated manufacturing and alignment, 3) the incorporation of numerous classes of materials within a single platform, and 4) the integration of 3D microstructures with prefabricated devices, sensing arrays, and nonplanar substrates. However, to widely deploy 3D printed microfluidics at research and commercial scales, critical issues related to printing factors, device integration strategies, and incorporation of multiple functionalities require further development and optimization. In this review, we summarize important figures of merit of 3D printed microfluidics and inspect recent progress in the field, including ink properties, structural resolutions, and hierarchical levels of integration with functional platforms. Particularly, we highlight advances in microfluidic devices printed with thermosetting elastomers, printing methodologies with enhanced degrees of automation and resolution, and the direct printing of microfluidics on various 3D surfaces. The substantial progress in the performance and multifunctionality of 3D printed microfluidics suggests a rapidly approaching era in which these versatile devices could be untethered from microfabrication facilities and created on demand by users in arbitrary settings with minimal prior training.

A Bionic Testbed for Cardiac Ablation Tools.

Bionic-engineered tissues have been proposed for testing the performance of cardiovascular medical devices and predicting clinical outcomes ex vivo. Progress has been made in the development of compliant electronics that are capable of monitoring treatment parameters and being coupled to engineered tissues; however, the scale of most engineered tissues is too small to accommodate the size of clinical-grade medical devices. Here, we show substantial progress toward bionic tissues for evaluating cardiac ablation tools by generating a centimeter-scale human cardiac disk and coupling it to a hydrogel-based soft-pressure sensor. The cardiac tissue with contiguous electromechanical function was made possible by our recently established method to 3D bioprint human pluripotent stem cells in an extracellular matrix-based bioink that allows for in situ cell expansion prior to cardiac differentiation. The pressure sensor described here utilized electrical impedance tomography to enable the real-time spatiotemporal mapping of pressure distribution. A cryoablation tip catheter was applied to the composite bionic tissues with varied pressure. We found a close correlation between the cell response to ablation and the applied pressure. Under some conditions, cardiomyocytes could survive in the ablated region with more rounded morphology compared to the unablated controls, and connectivity was disrupted. This is the first known functional characterization of living human cardiomyocytes following an ablation procedure that suggests several mechanisms by which arrhythmia might redevelop following an ablation. Thus, bionic-engineered testbeds of this type can be indicators of tissue health and function and provide unique insight into human cell responses to ablative interventions.

3D Printed Skin-Interfaced UV-Visible Hybrid Photodetectors.

Photodetectors that are intimately interfaced with human skin and measure real-time optical irradiance are appealing in the medical profiling of photosensitive diseases. Developing compliant devices for this purpose requires the fabrication of photodetectors with ultraviolet (UV)-enhanced broadband photoresponse and high mechanical flexibility, to ensure precise irradiance measurements across the spectral band critical to dermatological health when directly applied onto curved skin surfaces. Here, a fully 3D printed flexible UV-visible photodetector array is reported that incorporates a hybrid organic-inorganic material system and is integrated with a custom-built portable console to continuously monitor broadband irradiance in-situ. The active materials are formulated by doping polymeric photoactive materials with zinc oxide nanoparticles in order to improve the UV photoresponse and trigger a photomultiplication (PM) effect. The ability of a stand-alone skin-interfaced light intensity monitoring system to detect natural irradiance within the wavelength range of 310-650 nm for nearly 24 h is demonstrated.

3D-printed flexible organic light-emitting diode displays.

The ability to fully 3D-print active electronic and optoelectronic devices will enable unique device form factors via strategies untethered from conventional microfabrication facilities. Currently, the performance of 3D-printed optoelectronics can suffer from nonuniformities in the solution-deposited active layers and unstable polymer-metal junctions. Here, we demonstrate a multimodal printing methodology that results in fully 3D-printed flexible organic light-emitting diode displays. The electrodes, interconnects, insulation, and encapsulation are all extrusion-printed, while the active layers are spray-printed. Spray printing leads to improved layer uniformity via suppression of directional mass transport in the printed droplets. By exploiting the viscoelastic oxide surface of the printed cathode droplets, a mechanical reconfiguration process is achieved to increase the contact area of the polymer-metal junctions. The uniform cathode array is intimately interfaced with the top interconnects. This hybrid approach creates a fully 3D-printed flexible 8 × 8 display with all pixels turning on successfully.

Conduction Cooling and Plasmonic Heating Dramatically Increase Droplet Vitrification Volumes for Cell Cryopreservation.

Droplet vitrification has emerged as a promising ice-free cryopreservation approach to provide a supply chain for off-the-shelf cell products in cell therapy and regenerative medicine applications. Translation of this approach requires the use of low concentration (i.e., low toxicity) permeable cryoprotectant agents (CPA) and high post cryopreservation viability (>90%), thereby demanding fast cooling and warming rates. Unfortunately, with traditional approaches using convective heat transfer, the droplet volumes that can be successfully vitrified and rewarmed are impractically small (i.e., 180 picoliter) for <2.5 m permeable CPA. Herein, a novel approach to achieve 90-95% viability in micro-liter size droplets with 2 m permeable CPA, is presented. Droplets with plasmonic gold nanorods (GNRs) are printed onto a cryogenic copper substrate for improved cooling rates via conduction, while plasmonic laser heating yields >400-fold improvement in warming rates over traditional convective approach. High viability cryopreservation is then demonstrated in a model cell line (human dermal fibroblasts) and an important regenerative medicine cell line (human umbilical cord blood stem cells). This approach opens a new paradigm for cryopreservation and rewarming of dramatically larger volume droplets at lower CPA concentration for cell therapy and other regenerative medicine applications.

3D printed self-supporting elastomeric structures for multifunctional microfluidics.

Microfluidic devices fabricated via soft lithography have demonstrated compelling applications such as lab-on-a-chip diagnostics, DNA microarrays, and cell-based assays. These technologies could be further developed by directly integrating microfluidics with electronic sensors and curvilinear substrates as well as improved automation for higher throughput. Current additive manufacturing methods, such as stereolithography and multi-jet printing, tend to contaminate substrates with uncured resins or supporting materials during printing. Here, we present a printing methodology based on precisely extruding viscoelastic inks into self-supporting microchannels and chambers without requiring sacrificial materials. We demonstrate that, in the submillimeter regime, the yield strength of the as-extruded silicone ink is sufficient to prevent creep within a certain angular range. Printing toolpaths are specifically designed to realize leakage-free connections between channels and chambers, T-shaped intersections, and overlapping channels. The self-supporting microfluidic structures enable the automatable fabrication of multifunctional devices, including multimaterial mixers, microfluidic-integrated sensors, automation components, and 3D microfluidics.

3D printed patient-specific aortic root models with internal sensors for minimally invasive applications.

Minimally invasive surgeries have numerous advantages, yet complications may arise from limited knowledge about the anatomical site targeted for the delivery of therapy. Transcatheter aortic valve replacement (TAVR) is a minimally invasive procedure for treating aortic stenosis. Here, we demonstrate multimaterial three-dimensional printing of patient-specific soft aortic root models with internally integrated electronic sensor arrays that can augment testing for TAVR preprocedural planning. We evaluated the efficacies of the models by comparing their geometric fidelities with postoperative data from patients, as well as their in vitro hemodynamic performances in cases with and without leaflet calcifications. Furthermore, we demonstrated that internal sensor arrays can facilitate the optimization of bioprosthetic valve selections and in vitro placements via mapping of the pressures applied on the critical regions of the aortic anatomies. These models may pave exciting avenues for mitigating the risks of postoperative complications and facilitating the development of next-generation medical devices.

3D printed deformable sensors.

The ability to directly print compliant biomedical devices on live human organs could benefit patient monitoring and wound treatment, which requires the 3D printer to adapt to the various deformations of the biological surface. We developed an in situ 3D printing system that estimates the motion and deformation of the target surface to adapt the toolpath in real time. With this printing system, a hydrogel-based sensor was printed on a porcine lung under respiration-induced deformation. The sensor was compliant to the tissue surface and provided continuous spatial mapping of deformation via electrical impedance tomography. This adaptive 3D printing approach may enhance robot-assisted medical treatments with additive manufacturing capabilities, enabling autonomous and direct printing of wearable electronics and biological materials on and inside the human body.

In Situ Expansion, Differentiation, and Electromechanical Coupling of Human Cardiac Muscle in a 3D Bioprinted, Chambered Organoid.

RATIONALE: One goal of cardiac tissue engineering is the generation of a living, human pump in vitro that could replace animal models and eventually serve as an in vivo therapeutic. Models that replicate the geometrically complex structure of the heart, harboring chambers and large vessels with soft biomaterials, can be achieved using 3-dimensional bioprinting. Yet, inclusion of contiguous, living muscle to support pump function has not been achieved. This is largely due to the challenge of attaining high densities of cardiomyocytes-a notoriously nonproliferative cell type. An alternative strategy is to print with human induced pluripotent stem cells, which can proliferate to high densities and fill tissue spaces, and subsequently differentiate them into cardiomyocytes in situ. OBJECTIVE: To develop a bioink capable of promoting human induced pluripotent stem cell proliferation and cardiomyocyte differentiation to 3-dimensionally print electromechanically functional, chambered organoids composed of contiguous cardiac muscle. METHODS AND RESULTS: We optimized a photo-crosslinkable formulation of native ECM (extracellular matrix) proteins and used this bioink to 3-dimensionally print human induced pluripotent stem cell-laden structures with 2 chambers and a vessel inlet and outlet. After human induced pluripotent stem cells proliferated to a sufficient density, we differentiated the cells within the structure and demonstrated function of the resultant human chambered muscle pump. Human chambered muscle pumps demonstrated macroscale beating and continuous action potential propagation with responsiveness to drugs and pacing. The connected chambers allowed for perfusion and enabled replication of pressure/volume relationships fundamental to the study of heart function and remodeling with health and disease. CONCLUSIONS: This advance represents a critical step toward generating macroscale tissues, akin to aggregate-based organoids, but with the critical advantage of harboring geometric structures essential to the pump function of cardiac muscle. Looking forward, human chambered organoids of this type might also serve as a test bed for cardiac medical devices and eventually lead to therapeutic tissue grafting.

3D Printed Neural Regeneration Devices.

Neural regeneration devices interface with the nervous system and can provide flexibility in material choice, implantation without the need for additional surgeries, and the ability to serve as guides augmented with physical, biological (e.g., cellular), and biochemical functionalities. Given the complexity and challenges associated with neural regeneration, a 3D printing approach to the design and manufacturing of neural devices could provide next-generation opportunities for advanced neural regeneration via the production of anatomically accurate geometries, spatial distributions of cellular components, and incorporation of therapeutic biomolecules. A 3D printing-based approach offers compatibility with 3D scanning, computer modeling, choice of input material, and increasing control over hierarchical integration. Therefore, a 3D printed implantable platform could ultimately be used to prepare novel biomimetic scaffolds and model complex tissue architectures for clinical implants in order to treat neurological diseases and injuries. Further, the flexibility and specificity offered by 3D printed in vitro platforms have the potential to be a significant foundational breakthrough with broad research implications in cell signaling and drug screening for personalized healthcare. This progress report examines recent advances in 3D printing strategies for neural regeneration as well as insight into how these approaches can be improved in future studies.

Corrigendum to "3D printed bionic nanodevices" [Nano Today 11 (2016) 330-350].

[This corrects the article PMC5016035.].

3D Bioprinted In Vitro Metastatic Models via Reconstruction of Tumor Microenvironments.

The development of 3D in vitro models capable of recapitulating native tumor microenvironments could improve the translatability of potential anticancer drugs and treatments. Here, 3D bioprinting techniques are used to build tumor constructs via precise placement of living cells, functional biomaterials, and programmable release capsules. This enables the spatiotemporal control of signaling molecular gradients, thereby dynamically modulating cellular behaviors at a local level. Vascularized tumor models are created to mimic key steps of cancer dissemination (invasion, intravasation, and angiogenesis), based on guided migration of tumor cells and endothelial cells in the context of stromal cells and growth factors. The utility of the metastatic models for drug screening is demonstrated by evaluating the anticancer efficacy of immunotoxins. These 3D vascularized tumor tissues provide a proof-of-concept platform to i) fundamentally explore the molecular mechanisms of tumor progression and metastasis, and ii) preclinically identify therapeutic agents and screen anticancer drugs.

3D Printed Polymer Photodetectors.

Extrusion-based 3D printing, an emerging technology, has been previously used in the comprehensive fabrication of light-emitting diodes using various functional inks, without cleanrooms or conventional microfabrication techniques. Here, polymer-based photodetectors exhibiting high performance are fully 3D printed and thoroughly characterized. A semiconducting polymer ink is printed and optimized for the active layer of the photodetector, achieving an external quantum efficiency of 25.3%, which is comparable to that of microfabricated counterparts and yet created solely via a one-pot custom built 3D-printing tool housed under ambient conditions. The devices are integrated into image sensing arrays with high sensitivity and wide field of view, by 3D printing interconnected photodetectors directly on flexible substrates and hemispherical surfaces. This approach is further extended to create integrated multifunctional devices consisting of optically coupled photodetectors and light-emitting diodes, demonstrating for the first time the multifunctional integration of multiple semiconducting device types which are fully 3D printed on a single platform. The 3D-printed optoelectronic devices are made without conventional microfabrication facilities, allowing for flexibility in the design and manufacturing of next-generation wearable and 3D-structured optoelectronics, and validating the potential of 3D printing to achieve high-performance integrated active electronic materials and devices.

3D Printed Organ Models with Physical Properties of Tissue and Integrated Sensors.

The design and development of novel methodologies and customized materials to fabricate patient-specific 3D printed organ models with integrated sensing capabilities could yield advances in smart surgical aids for preoperative planning and rehearsal. Here, we demonstrate 3D printed prostate models with physical properties of tissue and integrated soft electronic sensors using custom-formulated polymeric inks. The models show high quantitative fidelity in static and dynamic mechanical properties, optical characteristics, and anatomical geometries to patient tissues and organs. The models offer tissue-mimicking tactile sensation and behavior and thus can be used for the prediction of organ physical behavior under deformation. The prediction results show good agreement with values obtained from simulations. The models also allow the application of surgical and diagnostic tools to their surface and inner channels. Finally, via the conformal integration of 3D printed soft electronic sensors, pressure applied to the models with surgical tools can be quantitatively measured.

3D Printed Functional and Biological Materials on Moving Freeform Surfaces.

Conventional 3D printing technologies typically rely on open-loop, calibrate-then-print operation procedures. An alternative approach is adaptive 3D printing, which is a closed-loop method that combines real-time feedback control and direct ink writing of functional materials in order to fabricate devices on moving freeform surfaces. Here, it is demonstrated that the changes of states in the 3D printing workspace in terms of the geometries and motions of target surfaces can be perceived by an integrated robotic system aided by computer vision. A hybrid fabrication procedure combining 3D printing of electrical connects with automatic pick-and-placing of surface-mounted electronic components yields functional electronic devices on a free-moving human hand. Using this same approach, cell-laden hydrogels are also printed on live mice, creating a model for future studies of wound-healing diseases. This adaptive 3D printing method may lead to new forms of smart manufacturing technologies for directly printed wearable devices on the body and for advanced medical treatments.

3D Printed Organ Models for Surgical Applications.

Medical errors are a major concern in clinical practice, suggesting the need for advanced surgical aids for preoperative planning and rehearsal. Conventionally, CT and MRI scans, as well as 3D visualization techniques, have been utilized as the primary tools for surgical planning. While effective, it would be useful if additional aids could be developed and utilized in particularly complex procedures involving unusual anatomical abnormalities that could benefit from tangible objects providing spatial sense, anatomical accuracy, and tactile feedback. Recent advancements in 3D printing technologies have facilitated the creation of patient-specific organ models with the purpose of providing an effective solution for preoperative planning, rehearsal, and spatiotemporal mapping. Here, we review the state-of-the-art in 3D printed, patient-specific organ models with an emphasis on 3D printing material systems, integrated functionalities, and their corresponding surgical applications and implications. Prior limitations, current progress, and future perspectives in this important area are also broadly discussed.

3D Printed Stem-Cell Derived Neural Progenitors Generate Spinal Cord Scaffolds.

A bioengineered spinal cord is fabricated via extrusion-based multi-material 3D bioprinting, in which clusters of induced pluripotent stem cell (iPSC)-derived spinal neuronal progenitor cells (sNPCs) and oligodendrocyte progenitor cells (OPCs) are placed in precise positions within 3D printed biocompatible scaffolds during assembly. The location of a cluster of cells, of a single type or multiple types, is controlled using a point-dispensing printing method with a 200 µm center-to-center spacing within 150 µm wide channels. The bioprinted sNPCs differentiate and extend axons throughout microscale scaffold channels, and the activity of these neuronal networks is confirmed by physiological spontaneous calcium flux studies. Successful bioprinting of OPCs in combination with sNPCs demonstrates a multicellular neural tissue engineering approach, where the ability to direct the patterning and combination of transplanted neuronal and glial cells can be beneficial in rebuilding functional axonal connections across areas of central nervous system (CNS) tissue damage. This platform can be used to prepare novel biomimetic, hydrogel-based scaffolds modeling complex CNS tissue architecture in vitro and harnessed to develop new clinical approaches to treat neurological diseases, including spinal cord injury.

3D Printed Electrically-Driven Soft Actuators.

Soft robotics is an emerging field enabled by advances in the development of soft materials with properties commensurate to their biological counterparts, for the purpose of reproducing locomotion and other distinctive capabilities of active biological organisms. The development of soft actuators is fundamental to the advancement of soft robots and bio-inspired machines. Among the different material systems incorporated in the fabrication of soft devices, ionic hydrogel-elastomer hybrids have recently attracted vast attention due to their favorable characteristics, including their analogy with human skin. Here, we demonstrate that this hybrid material system can be 3D printed as a soft dielectric elastomer actuator (DEA) with a unimorph configuration that is capable of generating high bending motion in response to an applied electrical stimulus. We characterized the device actuation performance via applied (i) ramp-up electrical input, (ii) cyclic electrical loading, and (iii) payload masses. A maximum vertical tip displacement of 9.78 ± 2.52 mm at 5.44 kV was achieved from the tested 3D printed DEAs. Furthermore, the nonlinear actuation behavior of the unimorph DEA was successfully modeled using analytical energetic formulation and a finite element method (FEM).