Bioprinting最新文献

Localized gene delivery via engineered scaffolds offers spatiotemporal control of the gene vector release. Here, we explored the capability of digital light processing based bioprinting to fabricate 3D scaffolds in hydrogels for controlled gene delivery. We demonstrated the compatibility of the method with three representative hydrogel biomaterials for gene delivery. We further investigated the highly tunable release profile with these scaffolds by creating and combining distinct release mechanisms of diffusion and ion exchange. The efficacy of gene delivery of these scaffolds was validated in vitro using 293T cells. Results from this work could potentially facilitate the development of synergistic and personalized gene therapies.

Patients with diseased/damaged bones are increasingly in need of bone replacement, tissue regeneration, and organ repairs. The shape and size of the injury vary from person to person; thus the customized medical implant is a novel technique that has gained interest in recent times which offers personalized implants to each individual. Additive manufacturing has considerable promise as an efficient fabrication technique for fabricating customized implants with complicated shapes or for fabricating implants for different sited inside the human body. Through cost-effectiveness, efficiency, and better patient outcomes, this method is expected to change healthcare in the near future. Researchers are using various biomaterials to fabricate orthopedic implants using different additive manufacturing techniques such as fused deposition modelling (FDM), stereolithography (SLA), selective laser sintering (SLS), selective laser melting (SLM), selective electron beam melting (SEBM), binder jetting printing (BJP), and direct energy deposition (DED) for the fabrication of the customized implants. The biomaterials and various additive manufacturing techniques employed in current bone tissue engineering implants are overviewed herein, along with their challenges and future direction. Moreover, multiple factors such as material compositions, surface properties, or process parameters are discussed, which significantly alters the properties of the fabricated scaffold. Lastly, various commercially available products and devices available for bone and bone joint implants fabricated using conventional techniques have also been discussed in this study. No AM-based implant commercialized products are available in the market to date, which shows the incredible urge for research in such an area. Based on the finding of this study, additive manufacturing has demonstrated enormous potential for providing a pathway for the fabrication of customized implants. However, certain difficulties still need to be resolved to accelerate its translation into the clinics.

Since the existing polymeric hydrogel inks lack printability, shape fidelity and the desired mechanical properties for bone tissue regeneration, a hydrogel comprised of gelatin (G), collagen (C), and hydroxyapatite (H) nanoparticles is utilized for extrusion-based 3D printing. The rheological characterization of the composite GCH inks was performed to evaluate their printability, while the cuboid column model scaffolds were printed at a printing speed of 8 mm/s by using different needle inner diameters (500 μm/21G, 250 μm/25G, and 200 μm/27G), followed by carbodiimide induced crosslinking for 12 or 24 h. The samples were tested for their micro-structure, swelling, compression and stiffness performance, before and after incubation in an HBSS solution for up to 14 days. The wall diameter of a 3D printed scaffold decreases and pore size increases primarily with the decreasing inner diameter of the nozzles, and secondarily by increasing the crosslinking time. This was supported with their swelling capacity and the creation of new CaP crystals on the scaffold walls' top surfaces by the time of incubation, necessary for cells’ adhesion, proliferation and growth.

The compressive modulus and stiffness of the scaffolds increases proportionally with the increase of their wall diameter and the time of crosslinking, and is inversely proportional to their pores size. The scaffold with the smaller pores provides superior modulus and stiffness, even after 14 days of incubation in the physiological solution (i.e. from ∼0.94 to ∼0.71 MPa and from 17 to 20 kPa to 5–9 kPa, respectively). This is comparative with the reported values for gelatin-based composites, and in the range for hard tissue regeneration at non-load bearing sites, as well as cartilage applications.

Pneumatic-based extrusion as a 3D bioprinting technique is used for the fabrication of tissue constructs. Biopolymers are used to create a hydrogel that is used as the biomaterial ink to fabricate intricate tissue scaffolds able to simulate pathophysiological conditions more accurately than 2D models. There is a delicate balance between the parameters facilitating complex structures without affecting the printed scaffold results, and therefore the influence of each parameter should be fully understood. The aim of this study was to systematically optimise the printing parameters required to successfully 3D bioprint a computer-aided design (CAD) model with a preformulated hybrid hydrogel. A commercial bioprinter with a pneumatic printhead the BioX™ was used with conical print nozzles. A hybrid hydrogel with 6% (w/v) alginate and 23% (w/v) Pluronic F-127 (PF127), displayed printability, high porosity, low degradation, non-Newtonian rheology and were used in the printing parameter optimisation part of the study. Parameters that were optimised included: nozzle size, printing speed, extrusion pressure and temperature. The parameter optimisation index (POI), printability and shape fidelity were used to determine the optimal printing parameters. This was used in combination with a newly formulated scoring system to determine printing accuracy of the scaffold. Parameters that yielded a 100% complete scaffold print was a nozzle size of 27G using an extrusion pressure of 70 kPa and printing speed of 30 mm/s at 37 °C. These printing parameters did not yield the best results in all printability indices evaluated. It was concluded that the visual observations in combination with quantitative grading methods of the scaffolds, were a similarly important factor to take into consideration when selecting the optimal printing parameters.

Microextrusion bioprinting enables heterogeneous constructs with high shape fidelity to be fabricated through the deposition of a bioink with the desired physico-chemical and biological characteristics.

In this work, a novel semi-synthetic hydrogel, consisting of gelatin methacrylate and Pluronic F127, has been specifically formulated to match the requirements of microextrusion bioprinting process. By merging the thermosensitive characteristics of Pluronic with the cross-linking features of gelatin methacrylate, the formulation showed a printability window characterized by good shape retention and chemical stability following photo-crosslinking, as demonstrated by a thorough printability assessment, performed employing empirical predictive models. The mechanical properties of the constructs were comparable to those of soft tissues, widening the range of applicability in soft tissue engineering. The bioink was successfully applied to the fabrication of multilayered porous constructs preserving high levels of cell viability. Interestingly, the spatial arrangement of the cells showed a high degree of alignment along the deposition direction. Overall, the manufacturing process developed herein could represent a promising strategy to design three-dimensional models with predetermined cellular alignment.

Angiogenesis plays a pivotal role in development and tissue growth, as well as in pathological conditions such as cancer. Being able to understand the basic mechanisms involved in the vascularization of tissues and angiogenic network formation provides a window to advance the development of in vitro tissue models and enhance tissue engineering applications. In this study, we leveraged a novel microfluidic-based three dimensional (3D) bioprinting technology and alginate-collagen type I (AGC) bioink, to develop a 3D bioprinting strategy to enable the biofabrication of complex angiogenic networks within the 3D structure. These networks were comprised of simian vacuolating virus 40 (SV40) transformed adult rat brain endothelial cell (SV-ARBEC)-laden hydrogel rings. With mechanical properties relevant for vascular tissue engineering applications, these bioprinted constructs formed spontaneous vascular networks, reminiscent of anisotropic tissue-like structures, while retaining high cellular viability. The vascular network formation was accompanied by extracellular matrix (ECM) remodeling, confirming sequential SV-ARBEC mediated collagen type I fiber deposition and reorganization. Treatment with broad spectrum matrix metalloproteinase (MMP) inhibitor supressed SV-ARBEC angiogenic sprouting, highlighting requirements of ECM remodeling in angiogenic network formation. This novel 3D microfluidic bioprinting technology and biocompatible AGC hydrogel fiber rings supported robust SV-ARBEC angiogenesis and corresponding ECM remodeling, allowing us to present a strategy suitable to advancing applications in vascular research and supporting the further development of disease models, novel testing beds for drug discovery and tissue engineering applications.

Epicardial transplantation of 3D bioprinted patches represents a promising protective strategy against infarction-induced myocardial damage. We previously showed that 3D bioprinted tissues containing cardiac spheroids [in alginate/gelatin (AlgGel) hydrogels] promoted cell viability/function and endothelial cell tubular self-assembly. Here, we hypothesise that bioprinted cardiac spheroid patches improve cardiac function after myocardial infarction (MI). To determine treatment effects of hydrogel alone or with cells, MI mice were transplanted with: (i) AlgGel acellular patches, (ii) AlgGel with freely suspended cardiac cells, (iii) AlgGel with cardiac spheroids. We included control MI mice (no treatment) and mice undergoing sham surgery. We performed measurements to 28 days including echocardiography, flow cytometry and transcriptomic analyses. Our results measured median baseline (pre-surgery) left ventricular ejection fraction (LVEF%) for all mice at 66%. Post-surgery, LVEF% was 58% for Sham (non-infarcted) and 41% for MI (no treatment) mice. Patch transplantation increased LVEF%: 55% (acellular; p = 0.012), 59% (cells; p = 0.106), 64% (spheroids; p = 0.010). Flow cytometry demonstrated host cardiac tissue immune cell population changes with treatments. RNAseq transcriptomes demonstrated similar gene expression profiles for Sham and mice treated with cardiac spheroid patches. Extrusion 3D bioprinting permits hydrogel patch generation even preserving microtissue cardiac spheroids directly suspended in the bioink. Inflammatory and genetic mechanisms may play important roles in regulating host responses after patch transplantation in infarcted hearts. Future studies are needed to elucidate the possible immune cell and gene expression-related molecular mechanisms underlying these initial findings.

Branching morphogenesis, a specialized part of morphogenesis, leads to the formation of microstructures (tubes, canals, and glands), source of the active organ functions. The dynamic mechanisms involved are appearance/disappearance of biomolecules morphogens gradients. In the context of angiogenesis, growth factors allow the initiation, regulation, and remodeling of blood vessels. In the particular case of micro-vascularization, it seems essential to reproduce and study the interaction of endothelial cells with their environment but also with other cellular components, including fibroblasts.

To bring understanding here, we developed an angiogenesis 3D bioprinted (microextrusion bioprinting) model based on a proliferative bioink (7.5% (w/v) gelatin, 0.5% (w/v) alginate, 2% (w/v) fibrinogen) populated with fibroblasts and HUVECs. We demonstrated that we were able to recapitulate branching angiogenesis, producing organized microvascularization tissue in 7 days only.

We clearly demonstrated that a bidirectional communication was at stake between the two cell types, evidenced only when both types were culture in a 3D environment. Proteomic results (multiplexed ELISA) consolidated the understanding of this phenomenon, with 11 angiogenic proteins identified in the co-culture supernatant. They were identified as inducers of vasculogenesis and angiogenesis. Through matrix composition and cell organization study, we were able to demonstrate that tissue remodeling, extracellular matrix production (type I collagen), phenotype modification (pericytes) were taking place in our branching morphogenesis model.

Thanks to this breakthrough scientific advance in the field of regenerative medicine, we can imagine the biofabrication of functional tissues and organs models in the coming decades.

The creation of tissue scaffolds complex enough to facilitate acceptable tissue repair is difficult with analogue production methods, but the trend of computer aided design and 3D printing shows promise for rapidly creating customisable, complex scaffolds. This review discusses recent advances in 3D printing biodegradable soft tissue scaffolds, focusing on poly(caprolactone) (PCL) as a major component of scaffolds for dermal, adipose, and muscle repair. PCL is a biodegradable polyester used in bone and other hard tissue scaffolds. However, creating softer blends and copolymers with PCL has enabled wider application in soft tissue-engineered scaffolds. The review begins with the challenges and requirements of soft tissue-engineered scaffolds, followed by new techniques and materials from recent work. The primary methods of printing soft scaffolds are highlighted, such as extrusion-based, liquid polymerisation based and bioprinting, with multi-material printing being a more recent trend featuring combinations of the previous methods.

Nervous system plays a dynamic role in communicating information from the brain to body parts through central and peripheral nerves. Significant destruction to the nerve system instigates loss of sensor and motor functions. The regeneration of such damaged nerve is essential for retaining its functionality. It requires the scaffold, which acts as an aqueduct between the distal and proximal ends during regeneration. The present review is mainly concerned with the design aspects of fabricating nerve guidance conduits (NGCs) for rectifying injured peripheral nerves using advanced materials and manufacturing methods. A detailed review is presented on the biological and structural properties of nerve conduits. The different design features of the NGCs are elaborated concerning biocompatibility, cell adhesion, and proliferation enhancement. The various biocompatible materials and additives used for fabricating nerve conduits are elaborately discussed. The application of machine learning is elaborated at different stages in developing the NGCs. In addition, challenges and futuristic aspects for improving scaffold properties in repairing and regenerating peripheral nerve injuries are explicated.