Cholangiocarcinoma (CCA) is characterized by heterogeneous mutations and a refractory nature. Thus, the development of a model for effective drug screening is urgently needed. As the established therapeutic testing models for CCA are often ineffective, we fabricated an enabling three-dimensional (3D)-bioprinted CCA-on-a-chip model that to a good extent resembled the multicellular microenvironment and the anatomical microstructure of the hepato-vascular–biliary system to perform high-content antitumor drug screening. Specifically, cholangiocytes, hepatocytes, and vascular endotheliocytes were employed for 3D bioprinting of the models, allowing for a high degree of spatial and tube-like microstructural control. Interestingly, it was possible to observe CCA cells attached to the surfaces of the gelatin methacryloyl (GelMA) hydrogel-embedded microchannels and overgrown in a thickening manner, generating bile duct stenosis, which was expected to be analogous to the in vivo configuration. Over 4000 differentially expressed genes were detected in the CCA cells in our 3D coculture model compared to the traditional two-dimensional (2D) monoculture. Further screening revealed that the CCA cells grown in the 3D traditional model were more sensitive to the antitumoral prodrug than those in the 2D monoculture due to drug biotransformation by the neighboring functional hepatocytes. This study provides proof-of-concept validation of our bioprinted CCA-on-a-chip as a promising drug screening model for CCA treatment and paves the way for potential personalized medicine strategies for CCA patients in the future.

Three-dimensional (3D) bioprinting is driving significant innovations in biomedicine over recent years. Under certain scenarios such as in intraoperative bioprinting, the bioinks used should exhibit not only cyto/biocompatibility but also adhesiveness in wet conditions. Herein, an adhesive bioink composed of gelatin methacryloyl, gelatin, methacrylated hyaluronic acid, and skin secretion of Andrias davidianus is designed. The bioink exhibits favorable cohesion to allow faithful extrusion bioprinting in wet conditions, while simultaneously showing good adhesion to a variety of surfaces of different chemical properties, possibly achieved through the diverse bonds presented in the bioink formulation. As such, this bioink is able to fabricate sophisticated planar and volumetric constructs using extrusion bioprinting, where the dexterity is further enhanced using ergonomic handheld bioprinters to realize in situ bioprinting. In vitro experiments reveal that cells maintain high viability; further in vivo studies demonstrate good integration and immediate injury sealing. The characteristics of the bioink indicate its potential widespread utility in extrusion bioprinting and will likely broaden the applications of bioprinting toward situations such as in situ dressing and minimally invasive tissue regeneration.

The role of the human intestinal tract in host-microbe interactions has been highlighted in recent years. Several 3-dimensional (3D) models have been developed to reproduce the physiological characteristics of the human gut and to investigate the function of the gut microbiota. One challenge for 3D models is to recapitulate the low oxygen concentrations in the intestinal lumen. Moreover, most earlier 3D culture systems used a membrane to physically separate bacteria from the intestinal epithelium, which has sometimes made the studies of bacteria adhering to or invading cells less feasible. We report the establishment of a 3D gut epithelium model and cultured it at high cell viability under an anaerobic condition. We further cocultured intestinal bacteria including both commensal and pathogen directly with epithelial cells in the established 3D model under the anaerobic condition. We subsequently compared the gene expression differences of aerobic and anaerobic conditions for cell and bacterial growth via dual RNA sequencing. Our study provides a physiologically relevant 3D gut epithelium model that mimics the anaerobic condition in the intestinal lumen and supplies a powerful system for future in-depth gut-microbe interactional investigations.

Three-dimensional (3D) bioprinting of vascular tissues that are mechanically and functionally comparable to their native counterparts is an unmet challenge. Here, we developed a tough double-network hydrogel (bio)ink for microfluidic (bio)printing of mono-and dual-layered hollow conduits to recreate vein-and artery-like tissues, respectively. The tough hydrogel consisted of energy-dissipative ionically cross-linked alginate and elastic enzyme-cross-linked gelatin. The 3D bioprinted venous and arterial conduits exhibited key functionalities of respective vessels including relevant mechanical properties, perfusability, barrier performance, expressions of specific markers, and susceptibility to severe acute respiratory syndrome coronavirus 2 pseudo-viral infection. Notably, the arterial conduits revealed physiological vasoconstriction and vasodilatation responses. We further explored the feasibility of these conduits for vascular anastomosis. Together, our study presents biofabrication of mechanically and functionally relevant vascular conduits, showcasing their potentials as vascular models for dis-ease studies in vitro and as grafts for vascular surgeries in vivo, possibly serving broad biomedical applications in the future.

Thrombosis in the circulation system can lead to major myocardial infarction and cardiovascular deaths. Understanding thrombosis formation is necessary for developing safe and effective treatments. In this work, using digital light processing (DLP)-based 3D printing, we fabricated sophisticatedin vitromodels of blood vessels with internal microchannels that can be used for thrombosis studies. In this regard, photoacoustic microscopy (PAM) offers a unique advantage for label-free visualization of the 3D-printed vessel models, with large penetration depth and functional sensitivity. We compared the imaging performances of two PAM implementations: optical-resolution PAM and acoustic-resolution PAM, and investigated 3D-printed vessel structures with different patterns of microchannels. Our results show that PAM can provide clear microchannel structures at depths up to 3.6 mm. We further quantified the blood oxygenation in the 3D-printed vascular models, showing that thrombi had lower oxygenation than the normal blood. We expect that PAM can find broad applications in 3D printing and bioprinting forin vitrostudies of various vascular and other diseases.

The architecture of the colloidal photonic crystals (CPCs) is of paramount importance to their functionality and applications. Nevertheless, the realization of CPCs with arbitrarily designed, volumetrically sophisticated structures at the macroscale remains challenging. In this work, a printable CPC ink was developed. By combining this ink with a digital light processing (DLP)-based three-dimensional (3D) printing system, we were able to fabricate the CPC superstructures featuring both digitally defined macroscale geometries and structural colors originating from the ordered structures at the sub-micron scale. Moreover, besides the arbitrarily adjustable and precisely designable architectures, the optical properties, mechanical performances, and stimuli-responsiveness of the printed objects could also be facilely tuned by the composition of the ink or the printing parameters of the DLP system. This technology endows us the ability to fabricate CPCs with a multitude of desirable functions as well as mimic the hierarchical structures and color-manipulation strategies of some creatures in nature, which paves the way for potentially broad applications in intelligent color displays, 3D integrated sensors, biomimetic color-morphing soft robots, and smart anti-counterfeiting labels, among others.

Growth and environmental responses are essential for living organisms to survive and adapt to constantly changing environments. In order to simulate new conditions and capture dynamic responses to environmental shifts in a developing whole-cell model of E. coli, we incorporated additional regulation, including dynamics of the global regulator guanosine tetraphosphate (ppGpp), along with dynamics of amino acid biosynthesis and translation. With the model, we show that under perturbed ppGpp conditions, small molecule feedback inhibition pathways, in addition to regulation of expression, play a role in ppGpp regulation of growth. We also found that simulations with dysregulated amino acid synthesis pathways provide average amino acid concentration predictions that are comparable to experimental results but on the single-cell level, concentrations unexpectedly show regular fluctuations. Additionally, during both an upshift and downshift in nutrient availability, the simulated cell responds similarly with a transient increase in the mRNA:rRNA ratio. This additional simulation functionality should support a variety of new applications and expansions of the E. coli Whole-Cell Modeling Project.

The use of biomimetic models of the glomerulus has the potential to improve our understanding of the pathogenesis of kidney diseases and to enable progress in therapeutics. Current in vitro models comprise organ-on-a-chip, scaffold-based and organoid approaches. Glomerulus-on-a-chip designs mimic components of glomerular microfluidic flow but lack the inherent complexity of the glomerular filtration barrier. Scaffold-based 3D culture systems and organoids provide greater microenvironmental complexity but do not replicate fluid flows and dynamic responses to fluidic stimuli. As the available models do not accurately model the structure or filtration function of the glomerulus, their applications are limited. An optimal approach to glomerular modelling is yet to be developed, but the field will probably benefit from advances in biofabrication techniques. In particular, 3D bioprinting technologies could enable the fabrication of constructs that recapitulate the complex structure of the glomerulus and the glomerular filtration barrier. The next generation of in vitro glomerular models must be suitable for high(er)-content or/and high(er)-throughput screening to enable continuous and systematic monitoring. Moreover, coupling of glomerular or kidney models with those of other organs is a promising approach to enable modelling of partial or full-body responses to drugs and prediction of therapeutic outcomes.

3D printing has emerged as an enabling approach in a variety of different fields. However, the bulk volume of printing systems limits the expansion of their applications. In this study, a portable 3D Digital Light Processing (DLP) printer is built based on a smartphone-powered projector and a custom-written smartphone-operated app. Constructs with detailed surface architectures, porous features, or hollow structures, as well as sophisticated tissue analogs, are successfully printed using this platform, by utilizing commercial resins as well as a range of hydrogel-based inks, including poly(ethylene glycol)-diacrylate, gelatin methacryloyl, or allylated gelatin. Moreover, due to the portability of the unique DLP printer, medical implants can be fabricated for point-of-care usage, and cell-laden tissues can be produced in situ, achieving a new milestone for mobile-health technologies. Additionally, the all-in-one printing system described herein enables the integration of the 3D scanning smartphone app to obtain object-derived 3D digital models for subsequent printing. Along with further developments, this portable, modular, and easy-to-use smartphone-enabled DLP printer is anticipated to secure exciting opportunities for applications in resource-limited and point-of-care settings not only in biomedicine but also for home and educational purposes.

3D Printing In article number 2102153, Khoon S. Lim, Yu Shrike Zhang, and co-workers present a smartphone-enabled Digital Light Processing (DLP) 3D printer based on a smartphone-powered projector and a customized touchscreen smartphone app, featuring portability, modularity, and an easy-to-use interface. Through printing with commercial resins and hydrogel-based (bio)inks, it is applied to the fabrication of medical implants, in situ bioprinting, and integration with 3D object-scanning capability.

Recapitulation of complex tissues signifies a remarkable challenge and, to date, only a few approaches have emerged that can efficiently reconstruct necessary gradients in 3D constructs. This is true even though mimicry of these gradients is of great importance to establish the functionality of engineered tissues and devices. Here, a composable-gradient Digital Light Processing (DLP)-based (bio)printing system is developed, utilizing the unprecedented integration of a microfluidic mixer for the generation of either continual or discrete gradients of desired (bio)inks in real time. Notably, the precisely controlled gradients are composable on-the-fly by facilely by adjusting the (bio)ink flow ratios. In addition, this setup is designed in such a way that (bio)ink waste is minimized when exchanging the gradient (bio)inks, further enhancing this time- and (bio)ink-saving strategy. Various planar and 3D structures exhibiting continual gradients of materials, of cell densities, of growth factor concentrations, of hydrogel stiffness, and of porosities in horizontal and/or vertical direction, are exemplified. The composable fabrication of multifunctional gradients strongly supports the potential of the unique bioprinting system in numerous biomedical applications.

Organs-on-chips have emerged as viable platforms for drug screening and personalized medicine. While a wide variety of human organ-on-a-chip models have been developed, rarely have there been reports on the inclusion of sensors, which are critical in continually measuring the microenvironmental parameters and the dynamic responses of the microtissues to pharmaceutical compounds over extended periods of time. In addition, automation capacity is strongly desired for chronological monitoring. To overcome this major hurdle, in this protocol we detail the fabrication of electrochemical affinity-based biosensors and their integration with microfluidic chips to achieve in-line microelectrode functionalization, biomarker detection and sensor regeneration, allowing continual, in situ and noninvasive quantification of soluble biomarkers on organ-on-a-chip platforms. This platform is almost universal and can be applied to in-line detection of a majority of biomarkers, can be connected with existing organ-on-a-chip devices and can be multiplexed for simultaneous measurement of multiple biomarkers. Specifically, this protocol begins with fabrication of the electrochemically competent microelectrodes and the associated microfluidic devices (~3 d). The integration of electrochemical biosensors with the chips and their further combination with the rest of the platform takes ~3 h. The functionalization and regeneration of the microelectrodes are subsequently described, which require ~7 h in total. One cycle of sampling and detection of up to three biomarkers accounts for ~1 h.

3D printing and bioprinting have become a key component in precision medicine. They have been used toward the fabrication of medical devices with patient-specific shapes, production of engineered tissues for in vivo regeneration, and preparation of in vitro tissue models used for screening therapeutics. In particular, vat polymerization-based 3D (bio)printing as a unique strategy enables more sophisticated architectures to be rapidly built. This progress report aims to emphasize the recent advances made in vat polymerization-based 3D printing and bioprinting, including new biomaterial ink formulations and novel vat polymerization system designs. While some of these approaches have not been utilized toward the combination with biomaterial inks, it is anticipated their rapid translation into biomedical applications.

Lymphatic vessels, as a means to metastasize, are frequently recruited by tumor tissues during their progression. However, reliable in vitro models to dissect the intricate crosstalk between lymphatic vessels and tumors are still in urgent demand. Here, we describe a tissue-engineering method based on sacrificial bioprinting, to develop an enabling model of the human breast tumor with embedded multiscale lymphatic vessels, which is compatible with existing microscopy to examine the processes of lymphatic vessel sprouting and breast tumor cell migration in a physiologically relevant volumetric microenvironment. This platform will potentially help shed light on the complex biology of the tumor microenvironment, tumor lymphangiogenesis, lymphatic metastasis, as well as tumor anti-lymphangiogenic therapy in the future. We further anticipate wide adoption of the method to the production of various tissues and their models with incorporation of lymphatics vessels towards relevant applications.

In this study, we present the photosynthetic oxygen (O2) supply to mammalian cells within a volumetric extracellular matrix-like construct, whereby a three-dimensional (3D)-bioprinted fugitive pattern encapsulating unicellular green algae, Chlamydomonas reinhardtii (C. reinhardtii), served as a natural photosynthetic O2-generator. The presence of bioprinted C. reinhardtii enhanced the viability and functionality of mammalian cells while reducing the hypoxic conditions within the tissues. We were able to subsequently endothelialize the hollow perfusable microchannels formed after enzymatic removal of the bioprinted C. reinhardtii-laden patterns from the matrices following the initial oxygenation period, to obtain biologically relevant vascularized mammalian tissue constructs. The feasibility of co-culture of C. reinhardtii with human cells, the printability and the enzymatic degradability of the fugitive bioink, as well as the exploration of C. reinhardtii as a natural, eco-friendly, cost-effective, and sustainable source of O2 would likely promote the development of engineered tissues, tissue models, and food for various applications.