Experimental study of the stable droplet formation process during micro-valve-based three-dimensional bioprinting

Biomaterial-based 3D bioprinting strategy for orthopedic tissue engineering.

Three-dimensional (3D) bioprinting technology has emerged as an ideal approach to address the challenges in regenerative dentistry by fabricating 3D tissue constructs with customized complex architecture. The dilemma with current dental treatments has led to the exploration of this technology in restoring and maintaining the function of teeth. This scoping review aims to explore 3D bioprinting technology together with the type of biomaterials and cells used for dental applications. Based on PRISMA-ScR guidelines, this systematic search was conducted by using the following databases: Ovid, PubMed, EBSCOhost and Web of Science. The inclusion criteria were (i) cell-laden 3D-bioprinted construct; (ii) intervention to regenerate dental tissue using bioink, which incorporates living cells or in combination with biomaterial; and (iii) 3D bioprinting for dental applications. A total of 31 studies were included in this review. The main 3D bioprinting technique was extrusion-based approach. Novel bioinks in use consist of different types of natural and synthetic polymers, decellularized extracellular matrix and spheroids with encapsulated mesenchymal stem cells, and have shown promising results for periodontal ligament, dentin, dental pulp and bone regeneration application. However, 3D bioprinting in dental applications, regrettably, is not yet close to being a clinical reality. Therefore, further research in fabricating ideal bioinks with implantation into larger animal models in the oral environment is very much needed for clinical translation.

Organoids provide more realistic in vitro models that closely mimic their corresponding tissues or organs. Three-dimensional (3D) bioprinting allows for precise control over cell distribution, arrangement, and the regulation of cell behavior and interactions, offering flexibility and repeatability. By combining 3D bioprinting with organoid culture, optimal extracellular matrix conditions can be created, better replicating the complex interactions between cells and their environment. The integration of 3D bioprinting amplifies the scalability of organoids, facilitating the development of large-scale organ models that significantly enhance research capabilities. This review explores the crucial role of 3D bioprinting in organoid development, emphasizing its contributions to improving organoid structure and functionality. We discuss how this innovative approach supports the creation of well-defined physiological microenvironments, aids in the development of functional organoids, and enables high-throughput cultivation. Furthermore, we highlight the advantages of optimizing bioprinting strategies and bioinks to advance organoid applications. In addition, we examine the importance of refining 3D bioprinting devices to enhance organoid fabrication and application. Finally, we outline strategies for leveraging 3D bioprinting to further advance organoid research and its implications for regenerative medicine and disease modeling.

Background: An autologous minimally manipulated homologous adipose tissue (AMHAT) fabricated using three-dimensional (3D) bioprinting has shown potential in the treatment of diabetic foot ulcers and other chronic wounds. Methods: This study assessed the efficacy of AMHAT fabricated using 3D bioprinting for treating skin defects after a wide excision of skin cancer lesions where primary closure was not feasible. A total of 10 consecutive patients were included. The wounds were photographed and analyzed using AI, and the fat tissue harvested from the lower abdomen was processed into an AMHAT patch using a 3D bioprinter. The wound area was measured weekly until complete wound closure was achieved. Results: The area of the skin defects ranged from 1.77-6.22 cm2, averaging 2.72 cm2. Complete closure was achieved in 3-5 (average, 4.2) weeks. The residual area decreased from 52% after 1 week to 3% after 4 weeks. The scar appearance was esthetically favorable, with one patient showing mild nostril asymmetry. Furthermore, patient-reported outcome evaluation using the Japanese version of SCAR-Q for postoperative scars showed a very high level of satisfaction. Conclusions: The 3D bioprinted AMHAT accelerates wound healing with minimal scarring, offering an important option for skin defects where esthetics are a priority.

Three-dimensional bioprinting adipose tissue and mammary Organoids feasible for artificial breast structure regeneration

Chapter 3 - Three-dimensional bioprinting in medical surgery

Cancer vasculogenesis is a pivotal focus of cancer research and treatment given its critical role in tumor development, metastasis, and the formation of vasculogenic microenvironments. Traditional approaches to investigating cancer vasculogenesis face significant challenges in accurately modeling intricate microenvironments. Recent advancements in three-dimensional (3D) bioprinting technology present promising solutions to these challenges. This review provides an overview of cancer vasculogenesis and underscores the importance of precise modeling. It juxtaposes traditional techniques with 3D bioprinting technologies, elucidating the advantages of the latter in developing cancer vasculogenesis models. Furthermore, it explores applications in pathological investigations, preclinical medication screening for personalized treatment and cancer diagnostics, and envisages future prospects for 3D bioprinted cancer vasculogenesis models. Despite notable advancements, current 3D bioprinting techniques for cancer vasculogenesis modeling have several limitations. Nonetheless, by overcoming these challenges and with technological advances, 3D bioprinting exhibits immense potential for revolutionizing the understanding of cancer vasculogenesis and augmenting treatment modalities.

In the era of personalised medicine, novel therapeutic approaches raise increasing hopes to address currently unmet medical needs by developing patient-customised treatments. Three-dimensional (3D) bioprinting is rapidly evolving and has the potential to obtain personalised tissue constructs and overcome some limitations of standard tissue engineering approaches. Bioprinting could support a wide range of biomedical applications, such as drug testing, tissue repair or organ transplantation. There is a growing interest for 3D bioprinting in the orthopaedic field, with remarkable scientific and technical advances. However, the full exploitation of 3D bioprinting in medical applications still requires efforts to anticipate the upcoming challenges in translating bioprinted products from bench to bedside. In this review we summarised current trends, advances and challenges in the application of 3D bioprinting for bone and cartilage tissue engineering. Moreover, we provided a detailed analysis of the applicable regulations through the 3D bioprinting process and an overview of available standards covering bioprinting and additive manufacturing.

3D bioprinting is a disruptive, computer-aided, and additive manufacturing technology that allows the obtention, layer-by-layer, of 3D complex structures. This technology is believed to offer tremendous opportunities in several fields including biomedical, pharmaceutical, and food industries. Several bioprinting processes and bio-ink materials have emerged recently. However, there is still a pressing need to develop low-cost sustainable bio-ink materials with superior qualities (excellent mechanical, viscoelastic and thermal properties, biocompatibility, and biodegradability). Marine-derived biomaterials, including polysaccharides and proteins, represent a viable and renewable source for bio-ink formulations. Therefore, the focus of this review centers around the use of marine-derived biomaterials in the formulations of bio-ink. It starts with a general overview of 3D bioprinting processes followed by a description of the most commonly used marine-derived biomaterials for 3D bioprinting, with a special attention paid to chitosan, glycosaminoglycans, alginate, carrageenan, collagen, and gelatin. The challenges facing the application of marine-derived biomaterials in 3D bioprinting within the biomedical and pharmaceutical fields along with future directions are also discussed.

Three-dimensional (3D) bioprinting is a promising technology which utilises additive manufacturing technologies to produce functional tissues and organs. A bioink is the term used to describe any biomaterial, containing cells or other biologics, which is compatible with 3D bioprinting processes. The ideal bioink is one which is biocompatible, biodegradable, can achieve high shape fidelity during and after the bioprinting process and can provide a microenvironment which can accommodate the formation of a functional biological tissue. This chapter will discuss the composition of multiple naturally derived bioinks and synthetic bioinks which are commonly used in 3D bioprinting. It will also discuss how bioink stiffness and composition can influence and direct cell fate. Finally, this chapter will demonstrate how bioinks have advanced the field of tissue engineering by highlighting some key strategies involving the 3D bioprinting of tissues such as bone, vascular tissue and liver.

Three-dimensional (3D) bioprinting combines biomaterials, cells and functional components into complex living tissues. Herein, we assembled function-control modules into cell-laden scaffolds using 3D bioprinting. A customized 3D printer was able to tune the microstructure of printed bone mesenchymal stem cell (BMSC)-laden methacrylamide gelatin scaffolds at the micrometer scale. For example, the pore size was adjusted to 282 ± 32 μm and 363 ± 60 μm. To match the requirements of the printing nozzle, collagen microfibers with a length of 22 ± 13 μm were prepared with a high-speed crusher. Collagen microfibers bound bone morphogenetic protein 2 (BMP2) with a collagen binding domain (CBD) as differentiation-control module, from which BMP2 was able to be controllably released. The differentiation behaviors of BMSCs in the printed scaffolds were compared in three microenvironments: samples without CBD-BMP2-collagen microfibers in the growth medium, samples without microfibers in the osteogenic medium and samples with microfibers in the growth medium. The results indicated that BMSCs showed high cell viability (>90%) during printing; CBD-BMP2-collagen microfibers induced BMSC differentiation into osteocytes within 14 days more efficiently than the osteogenic medium. Our studies suggest that these function-control modules are attractive biomaterials and have potential applications in 3D bioprinting.

Advances in 3D bioprinting technology for cardiac tissue engineering and regeneration

Current Progress of 3D bioprinting in oral and craniomaxillofacial tissue regeneration: A systematic review.

The principle of three-dimensional (3D) bio-printing involves integration of biomaterials, live cells and controlled motor systems for creating complex biomimetic constructs. Bio-ink is one of the most important components in the process of 3D bio-printing and needs to be sufficiently viscous to be dispensed as a free-standing filament but be biocompatible to maintain cell viability and function. Alginate has been used widely for 3D bio-printing due to its biocompatibility, tunable properties, rapid gelation, low cost, and ability to be functionalized to direct cell behavior. By tuning the physiochemical parameters of alginate-based bio-inks, such as viscosity, improvements in print resolution, fidelity and growth characteristics of encapsulated cells can be achieved. This study aimed to improve the printability of low concentration alginate bio-inks by utilizing calcium sulphate (CaSO4) pre-crosslinking. A variety of alginates, differing in their viscosity, molecular weight and b-D-mannuronate and α-L-guluronate residues were investigated by wet spinning and bio-printing. Rheological and structural properties of pre-crosslinked alginates were characterized with the aim of mitigating the resolution problems associated with the use of low percentage alginate bio-inks, more favorable for maintaining cell viability. Pre-crosslinking produced a significant effect on viscosity of biomaterials improving their suitability for the bio-printing process and influencing the final resolution of the printed structure. Medium viscosity high b-D-mannuronate containing alginate (MVM) showed the highest degree of viscosity change compared to the control (p < 0.0001; n = 6), assessed by single value viscometry analysis and shear rheology, after pre-crosslinking and was subsequently used in experiments with cells. The survival of human corneal stromal fibroblasts (CSFs) was assessed using CellTiterGlo metabolic assay and confirmed with Calcein acetoxymethyl and Ethidium homodimer -1 live/dead staining in pre-crosslinked alginate fibers and bio-printed lattices. Encapsulation of CSFs in pre-crosslinked alginate-based bio-inks did not have a detrimental effect on CSF viability compared to the non-pre-crosslinked control over 7 days under standard cell culture conditions (p > 0.05, n = 3). Overall, printability of low percentage alginate bio-inks was improved by pre-crosslinking without affecting the biocompatibility of the bio-inks.

Silk fibroin (SF) is an attractive material for composing bioinks suitable for three-dimensional (3D) bioprinting. However, the low viscosity of SF solutions obtained through common dissolution methods limits 3D-bioprinting applications without the addition of thickeners or partial gelation beforehand. Here, we report a method of 3D bioprinting low-viscosity SF solutions without additives. We combined a method of freeform reversible embedding of suspended hydrogels, known as the FRESH method, with horseradish peroxidase-catalyzed cross-linking. Using this method, we successfully fabricated 3D SF hydrogel constructs from low-viscosity SF ink (10% w/w, 50 mPa s at 1 s-1 shear rate), which does not yield 3D constructs when printed onto a plate in air. Studies using mouse fibroblasts confirmed that the printing process was cell-friendly. Additionally, cells enclosed in printed SF hydrogel constructs maintained > 90% viability for 11 days of culture. These results demonstrate that the 3D bioprinting technique developed in this study enables new 3D bioprinting applications using SF inks and thus has a great potential to contribute to tissue engineering and regenerative medicine.