Liquid-embedded (bio)printing of alginate-free, standalone, ultrafine, and ultrathin-walled cannular structures.

While there has been considerable success in the three-dimensional bioprinting of relatively large standalone filamentous tissues, the fabrication of solid fibers with ultrafine diameters or those cannular featuring ultrathin walls remains a particular challenge. Here, an enabling strategy for (bio)printing of solid and hollow fibers whose size ranges could be facilely adjusted across a broad spectrum, is reported, using an aqueous two-phase embedded (bio)printing approach combined with specially designed cross-linking and extrusion methods. The generation of standalone, alginate-free aqueous architectures using this aqueous two-phase strategy allowed freeform patterning of aqueous bioinks, such as those composed of gelatin methacryloyl, within the immiscible aqueous support bath of poly(ethylene oxide). Our (bio)printing strategy revealed the fabrication of standalone solid or cannular structures with diameters as small as approximately 3 or 40 µm, respectively, and wall thicknesses of hollow conduits down to as thin as <5 µm. With cellular functions also demonstrated, we anticipate the methodology to serve as a platform that may satisfy the needs for the different types of potential biomedical and other applications in the future, especially those pertaining to cannular tissues of ultrasmall diameters and ultrathin walls used toward regenerative medicine and tissue model engineering.

Conformation-driven strategy for resilient and functional protein materials.

The exceptional elastic resilience of some protein materials underlies essential biomechanical functions with broad interest in biomedical fields. However, molecular design of elastic resilience is restricted to amino acid sequences of a handful of naturally occurring resilient proteins such as resilin and elastin. Here, we exploit non-resilin/elastin sequences that adopt kinetically stabilized, random coil-dominated conformations to achieve near-perfect resilience comparable with that of resilin and elastin. We also show a direct correlation between resilience and Raman-characterized protein conformations. Furthermore, we demonstrate that metastable conformation of proteins enables the construction of mechanically graded protein materials that exhibit spatially controlled conformations and resilience. These results offer insights into molecular mechanisms of protein elastomers and outline a general conformation-driven strategy for developing resilient and functional protein materials.

Photonic control of image-guided ferroptosis cancer nanomedicine.

Photonic nanomaterials, characterized by their remarkable photonic tunability, empower a diverse range of applications, including cutting-edge advances in cancer nanomedicine. Recently, ferroptosis has emerged as a promising alternative strategy for effectively killing cancer cells with minimizing therapeutic resistance. Novel design of photonic nanomaterials that can integrate photoresponsive-ferroptosis inducers, -diagnostic imaging, and -synergistic components provide significant benefits to effectively trigger local ferroptosis. This review provides a comprehensive overview of recent advancements in photonic nanomaterials for image-guided ferroptosis cancer nanomedicine, offering insights into their strengths, constraints, and their potential as a future paradigm in cancer treatment.

Reversed-engineered human alveolar lung-on-a-chip model.

Here, we present a physiologically relevant model of the human pulmonary alveoli. This alveolar lung-on-a-chip platform is composed of a three-dimensional porous hydrogel made of gelatin methacryloyl with an inverse opal structure, bonded to a compartmentalized polydimethylsiloxane chip. The inverse opal hydrogel structure features well-defined, interconnected pores with high similarity to human alveolar sacs. By populating the sacs with primary human alveolar epithelial cells, functional epithelial monolayers are readily formed. Cyclic strain is integrated into the device to allow biomimetic breathing events of the alveolar lung, which, in addition, makes it possible to investigate pathological effects such as those incurred by cigarette smoking and severe acute respiratory syndrome coronavirus 2 pseudoviral infection. Our study demonstrates a unique method for reconstitution of the functional human pulmonary alveoli in vitro, which is anticipated to pave the way for investigating relevant physiological and pathological events in the human distal lung.

Nanofiber Aerogels with Precision Macrochannels and LL-37-Mimic Peptides Synergistically Promote Diabetic Wound Healing.

Fast healing of diabetic wounds remains a major clinical challenge. Herein, this work reports a strategy to combine nanofiber aerogels containing precision macrochannels and the LL-37-mimic peptide W379 for rapid diabetic wound healing. Nanofiber aerogels consisting of poly(glycolide-co-lactide) (PGLA 90:10)/gelatin and poly-p-dioxanone (PDO)/gelatin short electrospun fiber segments were prepared by partially anisotropic freeze-drying, crosslinking, and sacrificial templating with three-dimensional (3D)-printed meshes, exhibiting nanofibrous architecture and precision micro-/macrochannels. Like human cathelicidin LL-37, W379 peptide at a concentration of 3 µg/mL enhanced the migration and proliferation of keratinocytes and dermal fibroblasts in a cell scratch assay and a proliferation assay. In vivo studies show that nanofiber aerogels with precision macrochannels can greatly promote cell penetration compared to aerogels without macrochannels. Relative to control and aerogels with and without macrochannels, adding W379 peptides to aerogels with precision macrochannels shows the best efficacy in healing diabetic wounds in mice in terms of cell infiltration, neovascularization, and re-epithelialization. The fast re-epithelization could be due to upregulation of phospho-extracellular signal-regulated kinase (p38 MAPK) after treatment with W379. Together, the approach developed in this work could be promising for the treatment of diabetic wounds and other chronic wounds.

An Adhesive Bioink toward Biofabrication under Wet Conditions.

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.

A Natural Hydrogel with Prohealing Properties Enhances Tendon Regeneration.

Tendon regeneration and reduction of peritendinous adhesion remain major clinical challenges. This study addresses these challenges by adopting a unique hydrogel derived from the skin secretion of Andrias davidianus (SSAD) and taking advantage of its biological effects, adhesiveness, and controllable microstructures. The SSAD-derived hydrogel contains many cytokines, which could promote tendon healing. In vitro, leach liquid of SSAD powder could promote tendon stem/progenitor cells migration. In vivo, the SSAD-derived hydrogel featuring double layers possesses strong adhesiveness and could reconnect ruptured Achilles tendons of Sprague-Dawley rats without suturing. The intimal SSAD-derived hydrogel, with a pore size of 241.7 ± 21.0 µm, forms the first layer of the hydrogel to promote tendon healing, and the outer layer SSAD-derived hydrogel, with a pore size of 3.3 ± 1.4 µm, reducing peritendinous adhesion by serving as a dense barrier. Additionally, the SSAD-derived hydrogel exhibits antioxidant and antibacterial characteristics, which further contribute to the reduction of peritendinous adhesion. In vivo studies suggest that the SSAD-derived hydrogel reduces peritendinous adhesion, increases collagen fiber deposition, promotes cell proliferation, and improves the biomechanical properties of the regenerated tendons, indicating better functional restoration. The SSAD-derived bilayer hydrogel may be a feasible biomaterial for tendon repair in the future.

A Bioinspired Hemostatic Powder Derived from the Skin Secretion of Andrias davidianus for Rapid Hemostasis and Intraoral Wound Healing.

High-performance hemostasis has become increasingly essential in treating various traumas. However, available topical hemostats still have various drawbacks and side-effects. Herein, hemostatic powders derived from the skin secretion of Andrias davidianus (SSAD) with controllable particle size are prepared using feasible frozen-ball milling following lyophilization for hemorrhage-control. Scanning electron microscopy, rheometry, and Brunauer-Emmett-Teller test are used to characterize the coagulation-promoting surface topography, rheological properties, and porous structure of the SSAD particles. The blood-coagulation assays showed that the SSAD powders can induce blood-absorption in a particle size-dependent manner. Particle sizes of the SSAD powders larger than 200 µm and smaller than 800 µm greatly affect the blood-clotting rate. Associated with the thromboelastography (TEG) and amino acid/protein composition analyses, the accessibility and diffusion of blood are mainly dependent on the wettability, adhesivity, and clotting factors of the SSAD particles. Rapid hemostasis in vivo further involves three hemorrhage models (liver, femoral artery, and tail) as well as an oral wound model, which suggest favorable hemostatic and simultaneous regenerative effects of the SSAD hemostatic powder. Considering its degradability and good biocompatibility, SSAD can be an optimal candidate for a new class of inexpensive, natural, and promising hemostatic and wound-dressing agent.

Micropore-Forming Gelatin Methacryloyl (GelMA) Bioink Toolbox 2.0: Designable Tunability and Adaptability for 3D Bioprinting Applications.

It is well-known that tissue engineering scaffolds that feature highly interconnected and size-adjustable micropores are oftentimes desired to promote cellular viability, motility, and functions. Unfortunately, the ability of precise control over the microporous structures within bioinks in a cytocompatible manner for applications in 3D bioprinting is generally lacking, until a method of micropore-forming bioink based on gelatin methacryloyl (GelMA) was reported recently. This bioink took advantage of the unique aqueous two-phase emulsion (ATPE) system, where poly(ethylene oxide) (PEO) droplets are utilized as the porogen. Considering the limitations associated with this very initial demonstration, this article has furthered the understanding of the micropore-forming GelMA bioinks by conducting a systematic investigation into the additional GelMA types (porcine and fish, different methacryloyl-modification degrees) and porogen types (PEO, poly(vinyl alcohol), and dextran), as well as the effects of the porogen concentrations and molecular weights on the properties of the GelMA-based ATPE bioink system. This article exemplifies not only the significantly wider range of micropore sizes achievable and better emulsion stability, but also the improved suitability for both extrusion and digital light processing bioprinting with favorable cellular responses.

Drawn-on-Skin Sensors from Fully Biocompatible Inks toward High-Quality Electrophysiology.

The need to develop wearable devices for personal health monitoring, diagnostics, and therapy has inspired the production of innovative on-demand, customizable technologies. Several of these technologies enable printing of raw electronic materials directly onto biological organs and tissues. However, few of them have been thoroughly investigated for biocompatibility of the raw materials on the cellular, tissue, and organ levels or with different cell types. In addition, highly accurate multiday in vivo monitoring using such on-demand, in situ fabricated devices has yet to be done. Presented herein is the first fully biocompatible, on-skin fabricated electronics for multiple cell types and tissues that can capture electrophysiological signals with high fidelity. While also demonstrating improved mechanical and electrical properties, the drawn-on-skin ink retains its properties under various writing conditions, which minimizes the variation in electrical performance. Furthermore, the drawn-on-skin ink shows excellent biocompatibility with cardiomyocytes, neurons, mice skin tissue, and human skin. The high signal-to-noise ratios of the electrophysiological signals recorded with the DoS sensor over multiple days demonstrate its potential for personalized, long-term, and accurate electrophysiological health monitoring.

Investigating lymphangiogenesis in a sacrificially bioprinted volumetric model of breast tumor tissue.

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.

3D Printing of Monolithic Proteinaceous Cantilevers Using Regenerated Silk Fibroin.

Silk fibroin, regenerated from Bombyx mori, has shown considerable promise as a printable, aqueous-based ink using a bioinspired salt-bath system in our previous work. Here, we further developed and characterized silk fibroin inks that exhibit concentration-dependent fluorescence spectra at the molecular level. These insights supported extrusion-based 3D printing using concentrated silk fibroin solutions as printing inks. 3D monolithic proteinaceous structures with high aspect ratios were successfully printed using these approaches, including cantilevers only supported at one end. This work provides further insight and broadens the utility of 3D printing with silk fibroin inks for the microfabrication of proteinaceous structures.

A Heart-Breast Cancer-on-a-Chip Platform for Disease Modeling and Monitoring of Cardiotoxicity Induced by Cancer Chemotherapy.

Cardiotoxicity is one of the most serious side effects of cancer chemotherapy. Current approaches to monitoring of chemotherapy-induced cardiotoxicity (CIC) as well as model systems that develop in vivo or in vitro CIC platforms fail to notice early signs of CIC. Moreover, breast cancer (BC) patients with preexisting cardiac dysfunctions may lead to different incident levels of CIC. Here, a model is presented for investigating CIC where not only induced pluripotent stem cell (iPSC)-derived cardiac tissues are interacted with BC tissues on a dual-organ platform, but electrochemical immuno-aptasensors can also monitor cell-secreted multiple biomarkers. Fibrotic stages of iPSC-derived cardiac tissues are promoted with a supplement of transforming growth factor-ß 1 to assess the differential functionality in healthy and fibrotic cardiac tissues after treatment with doxorubicin (DOX). The production trend of biomarkers evaluated by using the immuno-aptasensors well-matches the outcomes from conventional enzyme-linked immunosorbent assay, demonstrating the accuracy of the authors' sensing platform with much higher sensitivity and lower detection limits for early monitoring of CIC and BC progression. Furthermore, the versatility of this platform is demonstrated by applying a nanoparticle-based DOX-delivery system. The proposed platform would potentially help allow early detection and prediction of CIC in individual patients in the future.

Studying endothelial cell shedding and orientation using adaptive perfusion-culture in a microfluidic vascular chip.

Most tissue-engineered blood vessels are endothelialized by static cultures in vitro. However, it has not been clear whether endothelial cell-shedding and local damage may occur in an endothelial layer formed by static cultures under the effect of blood flow shear postimplantation. In this study, we report a bionic and cost-effective vascular chip platform, and proved that a static culture of endothelialized tissue-engineered blood vessels had the problem of a large number of endothelial cells falling off under the condition imitating the human arterial blood flow, and we addressed this challenge by regulating the flow field in a vascular chip. Electrospun membranes made of highly oriented or randomly distributed poly(ε-caprolactone) fibers were used as the vascular scaffolds, on which endothelial cells proliferated well and eventually formed dense intima layers. We noted that the monolayers gradually adapted to the artery-like microenvironment through the regulation of chip flow field, which also revealed improved cellular orientations. In conclusion, we have proposed a vascular chip with adaptive flow patterns to gradually accommodate the statically cultured vascular endothelia to the shear environment of arterial flow field and enhanced the orientation of the endothelial cells. This strategy may find numerous potential applications such as screening of vascular engineering biomaterials and maturation parameters, studying of vascular biology and pathology, and construction of vessel-on-a-chip models for drug analysis, among others.

Deep Learning-Enabled Resolution-Enhancement in Mini- and Regular Microscopy for Biomedical Imaging.

Artificial intelligence algorithms that aid mini-microscope imaging are attractive for numerous applications. In this paper, we optimize artificial intelligence techniques to provide clear, and natural biomedical imaging. We demonstrate that a deep learning-enabled super-resolution method can significantly enhance the spatial resolution of mini-microscopy and regular-microscopy. This data-driven approach trains a generative adversarial network to transform low-resolution images into super-resolved ones. Mini-microscopic images and regular-microscopic images acquired with different optical microscopes under various magnifications are collected as our experimental benchmark datasets. The only input to this generative-adversarial-network-based method are images from the datasets down-sampled by the Bicubic interpolation. We use independent test set to evaluate this deep learning approach with other deep learning-based algorithms through qualitative and quantitative comparisons. To clearly present the improvements achieved by this generative-adversarial-network-based method, we zoom into the local features to explore and highlight the qualitative differences. We also employ the peak signal-to-noise ratio and the structural similarity, to quantitatively compare alternative super-resolution methods. The quantitative results illustrate that super-resolution images obtained from our approach with interpolation parameter α=0.25 more closely match those of the original high-resolution images than to those obtained by any of the alternative state-of-the-art method. These results are significant for fields that use microscopy tools, such as biomedical imaging of engineered living systems. We also utilize this generative adversarial network-based algorithm to optimize the resolution of biomedical specimen images and then generate three-dimensional reconstruction, so as to enhance the ability of three-dimensional imaging throughout the entire volumes for spatial-temporal analyses of specimen structures.

Bioprinted Injectable Hierarchically Porous Gelatin Methacryloyl Hydrogel Constructs with Shape-Memory Properties.

Direct injection of cell-laden hydrogels shows high potentials in tissue regeneration for translational therapy. The traditional cell-laden hydrogels are often used as bulk space fillers to tissue defects after injection, likely limiting their structural controllability. On the other hand, patterned cell-laden hydrogel constructs often necessitate invasive surgical procedures. To overcome these problems, herein, we report a unique strategy for encapsulating living human cells in a pore-forming gelatin methacryloyl (GelMA)-based bioink to ultimately produce injectable hierarchically macro-micro-nanoporous cell-laden GelMA hydrogel constructs through three-dimensional (3D) extrusion bioprinting. The hydrogel constructs can be fabricated into various shapes and sizes that are defect-specific. Due to the hierarchically macro-micro-nanoporous structures, the cell-laden hydrogel constructs can readily recover to their original shapes, and sustain high cell viability, proliferation, spreading, and differentiation after compression and injection. Besides, in vivo studies further reveal that the hydrogel constructs can integrate well with the surrounding host tissues. These findings suggest that our unique 3D-bioprinted pore-forming GelMA hydrogel constructs are promising candidates for applications in minimally invasive tissue regeneration and cell therapy.

Faithful Fabrication of Biocompatible Multicompartmental Memomicrospheres for Digitally Color-Tunable Barcoding.

Barcodes have attracted widespread attention, especially for the multiplexed bioassays and anti-counterfeiting used toward medical and biomedical applications. An enabling gas-shearing approach is presented for generating 10-faced microspherical barcodes with precise control over the properties of each compartment. As such, the color of each compartment could be programmatically adjusted in the 10-faced memomicrospheres by using pregel solutions containing different combinations of fluorescent nanoparticles. During the process, three primary colors (red, green, and blue) are adopted to obtain up to seven merged fluorescent colors for constituting a large amount of coding as well as a magnetic compartment, capable of effective and robust high-throughput information-storage. More importantly, by using the biocompatible sodium alginate to construct the multicolor microspherical barcodes, the proposed technology is likely to advance the fields of food and pharmaceutics anti-counterfeiting. These remarkable properties point to the potential value of gas-shearing in engineering microspherical barcodes for biomedical applications in the future.

Multisensor-integrated organs-on-chips platform for automated and continual in situ monitoring of organoid behaviors.

Organ-on-a-chip systems are miniaturized microfluidic 3D human tissue and organ models designed to recapitulate the important biological and physiological parameters of their in vivo counterparts. They have recently emerged as a viable platform for personalized medicine and drug screening. These in vitro models, featuring biomimetic compositions, architectures, and functions, are expected to replace the conventional planar, static cell cultures and bridge the gap between the currently used preclinical animal models and the human body. Multiple organoid models may be further connected together through the microfluidics in a similar manner in which they are arranged in vivo, providing the capability to analyze multiorgan interactions. Although a wide variety of human organ-on-a-chip models have been created, there are limited efforts on the integration of multisensor systems. However, in situ continual measuring is critical in precise assessment of the microenvironment parameters and the dynamic responses of the organs to pharmaceutical compounds over extended periods of time. In addition, automated and noninvasive capability is strongly desired for long-term monitoring. Here, we report a fully integrated modular physical, biochemical, and optical sensing platform through a fluidics-routing breadboard, which operates organ-on-a-chip units in a continual, dynamic, and automated manner. We believe that this platform technology has paved a potential avenue to promote the performance of current organ-on-a-chip models in drug screening by integrating a multitude of real-time sensors to achieve automated in situ monitoring of biophysical and biochemical parameters.

Generation of Cost-Effective Paper-Based Tissue Models through Matrix-Assisted Sacrificial 3D Printing.

Due to the combined advantages of cellulose and nanoscale (diameter 20-60 nm), bacterial cellulose possesses a series of attractive features including its natural origin, moderate biosynthesis process, good biocompatibility, and cost-effectiveness. Moreover, bacterial cellulose nanofibers can be conveniently processed into three-dimensional (3D) intertwined structures and form stable paper devices after simple drying. These advantages make it suitable as the material for construction of organ-on-a-chip devices using matrix-assisted sacrificial 3D printing. We successfully fabricated various microchannel structures embedded in the bulk bacterial cellulose hydrogels and retained their integrity after the drying process. Interestingly, these paper-based devices containing hollow microchannels could be rehydrated and populated with relevant cells to form vascularized tissue models. As a proof-of-concept demonstration, we seeded human umbilical vein endothelial cells (HUVECs) into the microchannels to obtain the vasculature and inoculated the MCF-7 cells onto the surrounding matrix of the paper device to build a 3D paper-based vascularized breast tumor model. The results showed that the microchannels were perfusable, and both HUVECs and MCF-7 cells exhibited favorable proliferation behaviors. This study may provide a new strategy for constructing simple and low-cost in vitro tissue models, which may find potential applications in drug screening and personalized medicine.

A Tumor-on-a-Chip System with Bioprinted Blood and Lymphatic Vessel Pair.

Current in vitro anti-tumor drug screening strategies are insufficiently portrayed lacking true perfusion and draining microcirculation systems, which may post significant limitation in reproducing the transport kinetics of cancer therapeutics explicitly. Herein, we report the fabrication of an improved tumor model consisting of bioprinted hollow blood vessel and lymphatic vessel pair, hosted in a three-dimensional (3D) tumor microenvironment-mimetic hydrogel matrix, termed as the tumor-on-a-chip with bioprinted blood and lymphatic vessel pair (TOC-BBL). The bioprinted blood vessel was perfusable channel with opening on both ends while the bioprinted lymphatic vessel was blinded on one end, both of which were embedded in a hydrogel tumor mass, with vessel permeability individually tunable through optimization of the composition of the bioinks. We demonstrated that systems with different combinations of these bioprinted blood/lymphatic vessels exhibited varying levels of diffusion profiles for biomolecules and anti-cancer drugs. Our TOC-BBL platform mimicking the natural pathway of drug-tumor interactions would have the drug introduced through the perfusable blood vessel, cross the vascular wall into the tumor tissue via diffusion, and eventually drained into the lymphatic vessel along with the carrier flow. Our results suggested that this unique in vitro tumor model containing the bioprinted blood/lymphatic vessel pair may have the capacity of simulating the complex transport mechanisms of certain pharmaceutical compounds inside the tumor microenvironment, potentially providing improved accuracy in future cancer drug screening.