Using decellularized extracellular matrix (dECM) hydrogels as bioinks has been an important step forward for bioprinting of functional tissue constructs, considering their rich microenvironment and their high degree of biomimicry. However, directly using dECM hydrogels as bioinks may not be suitable for bioprinting processes because of the loss of shape fidelity and geometrical precision of bioprinted structure due to their slow gelation kinetics. In this article, the development and direct bioprinting of dECM hydrogel bioink from bovine Achilles tendon were presented. The developed bioink is used for a microcapillary-based bioprinting process without any support structure and/or any additional cross-linker components. The reported decellularization and solubilization methods yield dECM pre-gels which turn into stable hydrogels in a short time at physiological conditions. The gelation kinetics and mechanical strength of bioinks with different concentrations and digestion times are characterized. A support structure-free 3D bioprinting of the developed bioink is shown by aspirating dECM bioinks and then in situ gelation and extrusion through a fine microcapillary nozzle. The viability assays indicate that the developed dECM bioink has no cytotoxic effect on encapsulated NIH 3T3 cells and the cells show lineage-specific morphology in the early days of culture as well.
Multimaterial additive manufacturing or three-dimensional (3D) printing of hydrogel structures provides the opportunity to engineer geometrically dependent functionalities. However, current fabrication methods are mostly limited to one type of material or only provide one type of functionality. In this paper, we report a novel method of multimaterial deposition of hydrogel structures based on an aspiration-on-demand protocol, in which the constitutive multimaterial segments of extruded filaments were first assembled in liquid state by sequential aspiration of inks into a glass capillary, followed by in situ gel formation. We printed different patterned objects with varying chemical, electrical, mechanical, and biological properties by tuning process and material related parameters, to demonstrate the abilities of this method in producing heterogeneous and multi-functional hydrogel structures. Our results show the potential of proposed method in producing heterogeneous objects with spatially controlled functionalities while preserving structural integrity at the switching interface between different segments. We anticipate that this method would introduce new opportunities in multimaterial additive manufacturing of hydrogels for diverse applications such as biosensors, flexible electronics, tissue engineering and organ printing.
Three-dimensional (3D) bioprinting is a rapidly advancing tissue engineering technology that holds great promise for the regeneration of several tissues, including bone. However, to generate a successful 3D bone tissue engineering construct, additional complexities should be taken into account such as nutrient and oxygen delivery, which is often insufficient after implantation in large bone defects. We propose that a well-designed tissue engineering construct, that is, an implant with a specific spatial pattern of cells in a matrix, will improve the healing outcome. By using a computational model of bone regeneration we show that particular cell patterns in tissue engineering constructs are able to enhance bone regeneration compared to uniform ones. We successfully bioprinted one of the most promising cell-gradient patterns by using cell-laden hydrogels with varying cell densities and observed a high cell viability for three days following the bioprinting process. In summary, we present a novel strategy for the biofabrication of bone tissue engineering constructs by designing cell-gradient patterns based on a computational model of bone regeneration, and successfully bioprinting the chosen design. This integrated approach may increase the success rate of implanted tissue engineering constructs for critical size bone defects and also can find a wider application in the biofabrication of other types of tissue engineering constructs.
Three-dimensional (3D) bioprinting is an additive manufacturing process in which the combination of biomaterials and living cells, referred to as a bioink, is deposited layer-by-layer to form biologically active 3D tissue constructs. Recent advancements in the field show that the success of this technology requires the development of novel biomaterials or the improvement of existing bioinks. Polyethylene glycol (PEG) is one of the well-known synthetic biomaterials and has been commonly used as a photocrosslinkable bioink for bioprinting; however, other types of cell-friendly crosslinking mechanisms to form PEG hydrogels need to be explored for bioprinting and tissue engineering. In this work, we proposed micro-capillary based bioprinting of a novel molecularly engineered PEG-based bioink that transiently incorporates low molecular weight gelatin (LMWG) fragments. The rheological properties and release profile of the LMWG fragments were characterized, and their presence during hydrogel formation had no effect on the swelling ratio or sol fraction when compared to PEG hydrogels formed without the LMWG fragments. For bioprinting, PEG was first functionalized with cell-adhesive RGD ligands and was then crosslinked using protease-sensitive peptides via a Michael-type addition reaction inside the micro-capillary. The printability was assessed by the analysis of extrudability, shape fidelity, and printing accuracy of the hydrogel filaments after the optimization of the gelation conditions of the PEG-based bioink. The LMWG fragments supplemented into the bioink allowed the extrusion of smooth and uniform cylindrical strands of the hydrogel and improved shape fidelity and printing accuracy. Encapsulated cells in both bioprinted and non-bioprinted PEG-based hydrogels showed high viability and continued to proliferate over time in culture with a well-defined cell morphology depending on the presence of the cell adhesive peptide RGD. The presented micro-capillary based bioprinting process for a novel PEG-based bioink can be promising to construct complex 3D structures with micro-scale range and spatiotemporal variations without using any cytotoxic photoinitiator, UV light, or polymer support.
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