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Droplet‐based bioprinting has shown remarkable potential in tissue engineering and regenerative medicine. However, it requires bioinks with low viscosities, which makes it challenging to create complex 3D structures and spatially pattern them with different materials. This study introduces a novel approach to bioprinting sophisticated volumetric objects by merging droplet‐based bioprinting and cryobioprinting techniques. By leveraging the benefits of cryopreservation, we fabricated, for the first time, intricate, self‐supporting cell‐free or cell‐laden structures with single or multiple materials in a simple droplet‐based bioprinting process that is facilitated by depositing the droplets onto a cryoplate followed by crosslinking during revival. The feasibility of this approach is demonstrated by bioprinting several cell types, with cell viability increasing to 80%–90% after up to 2 or 3 weeks of culture. Furthermore, the applicational capabilities of this approach are showcased by bioprinting an endothelialized breast cancer model. The results indicate that merging droplet and cryogenic bioprinting complements current droplet‐based bioprinting techniques and opens new avenues for the fabrication of volumetric objects with enhanced complexity and functionality, presenting exciting potential for biomedical applications. 

Abstract 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. 

Abstract 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 sophisticated in vitro models 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 for in vitro studies of various vascular and other diseases.