Stiffness assisted cell-matrix remodeling trigger 3D mechanotransduction regulatory programs.

Engineered matrices provide a valuable platform to understand the impact of biophysical factors on cellular behavior such as migration, proliferation, differentiation, and tissue remodeling, through mechanotransduction. While recent studies have identified some mechanisms of 3D mechanotransduction, there is still a critical knowledge gap in comprehending the interplay between 3D confinement, ECM properties, and cellular behavior. Specifically, the role of matrix stiffness in directing cellular fate in 3D microenvironment, independent of viscoelasticity, microstructure, and ligand density remains poorly understood. To address this gap, we designed a nanoparticle crosslinker to reinforce collagen-based hydrogels without altering their chemical composition, microstructure, viscoelasticity, and density of cell-adhesion ligand and utilized it to understand cellular dynamics. This crosslinking mechanism utilizes nanoparticles as crosslink epicenter, resulting in 10-fold increase in mechanical stiffness, without other changes. Human mesenchymal stem cells (hMSCs) encapsulated in 3D responded to mechanical stiffness by displaying circular morphology on soft hydrogels (5 kPa) and elongated morphology on stiff hydrogels (30 kPa). Stiff hydrogels facilitated the production and remodeling of nascent extracellular matrix (ECM) and activated mechanotransduction cascade. These changes were driven through intracellular PI3AKT signaling, regulation of epigenetic modifiers and activation of YAP/TAZ signaling. Overall, our study introduces a unique biomaterials platform to understand cell-ECM mechanotransduction in 3D for regenerative medicine as well as disease modelling.

Dynamically Cross-Linked Granular Hydrogels for 3D Printing and Therapeutic Delivery.

Granular hydrogels have recently emerged as promising biomaterials for tissue engineering and 3D-printing applications, addressing the limitations of bulk hydrogels while exhibiting desirable properties such as injectability and high porosity. However, their structural stability can be improved with post-injection interparticle cross-linking. In this study, we developed granular hydrogels with interparticle cross-linking through reversible and dynamic covalent bonds. We fragmented photo-cross-linked bulk hydrogels to produce aldehyde or hydrazide-functionalized microgels using chondroitin sulfate. Mixing these microgels facilitated interparticle cross-linking through reversible hydrazone bonds, providing shear-thinning and self-healing properties for injectability and 3D printing. The resulting granular hydrogels displayed high mechanical stability without the need for secondary cross-linking. Furthermore, the porosity and sustained release of growth factors from these hydrogels synergistically enhanced cell recruitment. Our study highlights the potential of reversible interparticle cross-linking for designing injectable and 3D printable therapeutic delivery scaffolds using granular hydrogels. Overall, our study highlights the potential of reversible interparticle cross-linking to improve the structural stability of granular hydrogels, making them an effective biomaterial for use in tissue engineering and 3D-printing applications.

Nanobio Interface Between Proteins and 2D Nanomaterials.

Two-dimensional (2D) nanomaterials have significantly contributed to recent advances in material sciences and nanotechnology, owing to their layered structure. Despite their potential as multifunctional theranostic agents, the biomedical translation of these materials is limited due to a lack of knowledge and control over their interaction with complex biological systems. In a biological microenvironment, the high surface energy of nanomaterials leads to diverse interactions with biological moieties such as proteins, which play a crucial role in unique physiological processes. These interactions can alter the size, surface charge, shape, and interfacial composition of the nanomaterial, ultimately affecting its biological activity and identity. This review critically discusses the possible interactions between proteins and 2D nanomaterials, along with a wide spectrum of analytical techniques that can be used to study and characterize such interplay. A better understanding of these interactions would help circumvent potential risks and provide guidance toward the safer design of 2D nanomaterials as a platform technology for various biomedical applications.

Designing Cost-Effective Open-Source Multihead 3D Bioprinters.

For the past decade, additive manufacturing has resulted in significant advances toward fabricating anatomic-size patient-specific scaffolds for tissue models and regenerative medicine. This can be attributed to the development of advanced bioinks capable of precise deposition of cells and biomaterials. The combination of additive manufacturing with advanced bioinks is enabling researchers to fabricate intricate tissue scaffolds that recreate the complex spatial distributions of cells and bioactive cues found in the human body. However, the expansion of this promising technique has been hampered by the high cost of commercially available bioprinters and proprietary software. In contrast, conventional three-dimensional (3D) printing has become increasingly popular with home hobbyists and caused an explosion of both low-cost thermoplastic 3D printers and open-source software to control the printer. In this study, we bring these benefits into the field of bioprinting by converting widely available and cost-effective 3D printers into fully functional, open-source, and customizable multihead bioprinters. These bioprinters utilize computer controlled volumetric extrusion, allowing bioinks with a wide range of flow properties to be bioprinted, including non-Newtonian bioinks. We demonstrate the practicality of this approach by designing bioprinters customized with multiple extruders, automatic bed leveling, and temperature controls for ∼$400 USD. These bioprinters were then used for in vitro and ex vivo bioprinting to demonstrate their utility for tissue engineering.

Nanoengineered Ink for Designing 3D Printable Flexible Bioelectronics.

Flexible electronics require elastomeric and conductive biointerfaces with native tissue-like mechanical properties. The conventional approaches to engineer such a biointerface often utilize conductive nanomaterials in combination with polymeric hydrogels that are cross-linked using toxic photoinitiators. Moreover, these systems frequently demonstrate poor biocompatibility and face trade-offs between conductivity and mechanical stiffness under physiological conditions. To address these challenges, we developed a class of shear-thinning hydrogels as biomaterial inks for 3D printing flexible bioelectronics. These hydrogels are engineered through a facile vacancy-driven gelation of MoS2 nanoassemblies with naturally derived polymer-thiolated gelatin. Due to shear-thinning properties, these nanoengineered hydrogels can be printed into complex shapes that can respond to mechanical deformation. The chemically cross-linked nanoengineered hydrogels demonstrate a 20-fold rise in compressive moduli and can withstand up to 80% strain without permanent deformation, meeting human anatomical flexibility. The nanoengineered network exhibits high conductivity, compressive modulus, pseudocapacitance, and biocompatibility. The 3D-printed cross-linked structure demonstrates excellent strain sensitivity and can be used as wearable electronics to detect various motion dynamics. Overall, the results suggest that these nanoengineered hydrogels offer improved mechanical, electronic, and biological characteristics for various emerging biomedical applications including 3D-printed flexible biosensors, actuators, optoelectronics, and therapeutic delivery devices.

Nano-bio interactions of 2D molybdenum disulfide.

Two-dimensional (2D) molybdenum disulfide (MoS2) is an ultrathin nanomaterial with a high degree of anisotropy, surface-to-volume ratio, chemical functionality and mechanical strength. These properties together enable MoS2 to emerge as a potent nanomaterial for diverse biomedical applications including drug delivery, regenerative medicine, biosensing and bioelectronics. Thus, understanding the interactions of MoS2 with its biological interface becomes indispensable. These interactions, referred to as "nano-bio" interactions, play a key role in determining the biocompatibility and the pathways through which the nanomaterial influences molecular, cellular and biological function. Herein, we provide a critical overview of the nano-bio interactions of MoS2 and emphasize on how these interactions dictate its biomedical applications including intracellular trafficking, biodistribution and biodegradation. Also, a critical evaluation of the interactions of MoS2 with proteins and specific cell types such as immune cells and progenitor/stem cells is illustrated which governs the short-term and long-term compatibility of MoS2-based biomedical devices.

2D Covalent Organic Framework Direct Osteogenic Differentiation of Stem Cells.

2D covalent organic frameworks (COFs) are an emerging class of crystalline porous organic polymers with a wide-range of potential applications. However, poor processability, aqueous instability, and low water dispersibility greatly limit their practical biomedical implementation. Herein, a new class of hydrolytically stable 2D COFs for sustained delivery of drugs to direct stem cell fate is reported. Specifically, a boronate-based COF (COF-5) is stabilized using amphiphilic polymer Pluronic F127 (PLU) to produce COF-PLU nanoparticles with thickness of ≈25 nm and diameter ≈200 nm. These nanoparticles are internalized via clathrin-mediated endocytosis and have high cytocompatibility (half-inhibitory concentration ≈1 mg mL-1 ). Interestingly, the 2D COFs induce osteogenic differentiation in human mesenchymal stem cells, which is unique. In addition, an osteogenic agent-dexamethasone-is able to be loaded within the porous structure of COFs for sustained delivery which further enhances the osteoinductive ability. These results demonstrate for the first time the fabrication of hydrolytically stable 2D COFs for sustained delivery of dexamethasone and demonstrate its osteoinductive characteristics.

Emerging 2D Nanomaterials for Biomedical Applications.

Two-dimensional (2D) nanomaterials are an emerging class of biomaterials with remarkable potential for biomedical applications. The planar topography of these nanomaterials confers unique physical, chemical, electronic and optical properties, making them attractive candidates for therapeutic delivery, biosensing, bioimaging, regenerative medicine, and additive manufacturing strategies. The high surface-to-volume ratio of 2D nanomaterials promotes enhanced interactions with biomolecules and cells. A range of 2D nanomaterials, including transition metal dichalcogenides (TMDs), layered double hydroxides (LDHs), layered silicates (nanoclays), 2D metal carbides and nitrides (MXenes), metal-organic framework (MOFs), covalent organic frameworks (COFs) and polymer nanosheets have been investigated for their potential in biomedical applications. Here, we will critically evaluate recent advances of 2D nanomaterial strategies in biomedical engineering and discuss emerging approaches and current limitations associated with these nanomaterials. Due to their unique physical, chemical, and biological properties, this new class of nanomaterials has the potential to become a platform technology in regenerative medicine and other biomedical applications.

Generalizing hydrogel microparticles into a new class of bioinks for extrusion bioprinting.

Hydrogel microparticles (HMPs) are an emerging bioink that can allow three-dimensional (3D) printing of most soft biomaterials by improving physical support and maintaining biological functions. However, the mechanisms of HMP jamming within printing nozzles and yielding to flow remain underexplored. Here, we present an in-depth investigation via both experimental and computational methods on the HMP dissipation process during printing as a result of (i) external resistance from the printing apparatus and (ii) internal physicochemical properties of HMPs. In general, a small syringe opening, large or polydisperse size of HMPs, and less deformable HMPs induce high resistance and closer HMP packing, which improves printing fidelity and stability due to increased interparticle adhesion. However, smooth extrusion and preserving viability of encapsulated cells require low resistance during printing, which is associated with less shear stress. These findings can be used to improve printability of HMPs and facilitate their broader use in 3D bioprinting.

Nanoengineered Light-Activatable Polybubbles for On-Demand Therapeutic Delivery.

Vaccine coverage is severely limited in developing countries due to inefficient protection of vaccine functionality as well as lack of patient compliance to receive the additional booster doses. Thus, there is an urgent need to design a thermostable vaccine delivery platform that also enables release of the bolus after predetermined time. Here, the formation of injectable and light-activatable polybubbles for vaccine delivery is reported. In vitro studies show that polybubbles enable delayed burst release, irrespective of cargo types, namely small molecule and antigen. The extracorporeal activation of polybubbles is achieved by incorporating near-infrared (NIR)-sensitive gold nanorods (AuNRs). Interestingly, light-activatable polybubbles can be used for on-demand burst release of cargo. In vitro, ex vivo, and in vivo studies demonstrate successful activation of AuNR-loaded polybubbles. Overall, the light-activatable polybubble technology can be used for on-demand delivery of various therapeutics including small molecule drugs, immunologically relevant protein, peptide antigens, and nucleic acids.

Bioprinting 101: Design, Fabrication, and Evaluation of Cell-Laden 3D Bioprinted Scaffolds.

3D bioprinting is an additive manufacturing technique that recapitulates the native architecture of tissues. This is accomplished through the precise deposition of cell-containing bioinks. The spatiotemporal control over bioink deposition permits for improved communication between cells and the extracellular matrix, facilitates fabrication of anatomically and physiologically relevant structures. The physiochemical properties of bioinks, before and after crosslinking, are crucial for bioprinting complex tissue structures. Specifically, the rheological properties of bioinks determines printability, structural fidelity, and cell viability during the printing process, whereas postcrosslinking of bioinks are critical for their mechanical integrity, physiological stability, cell survival, and cell functions. In this review, we critically evaluate bioink design criteria, specifically for extrusion-based 3D bioprinting techniques, to fabricate complex constructs. The effects of various processing parameters on the biophysical and biochemical characteristics of bioinks are discussed. Furthermore, emerging trends and future directions in the area of bioinks and bioprinting are also highlighted. Graphical abstract [Figure: see text] Impact statement Extrusion-based 3D bioprinting is an emerging additive manufacturing approach for fabricating cell-laden tissue engineered constructs. This review critically evaluates bioink design criteria to fabricate complex tissue constructs. Specifically, pre- and post-printing evaluation approaches are described, as well as new research directions in the field of bioink development and functional bioprinting are highlighted.