Search

The integration of light-driven technologies into biofabrication has revolutionized the field of tissue engineering and regenerative medicine, with numerous breakthroughs in the last few years. Light-based bioprinting approaches (lithography, multiphoton and volumetric bioprinting) have shown the potential to fabricate large scale tissue engineering constructs of high resolution, with great flexibility and control over the cellular organization. Given the unprecedented degree of freedom in fabricating convoluted structures, key challenges in regenerative medicine, such as introducing complex channels and pre-vascular networks in 3D constructs have also been addressed. Light has also been proven as a powerful tool, leading to novel photo-chemistry in designing bioinks, but also able to impart spatial-temporal control over cellular functions through photo-responsive chemistry. For instance, smart constructs able to undergo remotely controlled shape changes, stiffening, softening and degradation can be produced. The non-invasive nature of light stimulation also enables to trigger such responses post-fabrication, during the maturation phase of a construct. Such unique ability can be used to mimic the dynamic processes occurring in tissue regeneration, as well as in disease progression and degenerative processes in vivo. Bringing together these novel multidisciplinary expertise, the present Special Issue aims to discuss the most recent trends, strategies and novel light-based technologies in the field of biofabrication. These include: 1) using light-based bioprinting to develop in vitro models for drug screening, developmental biology models, disease models, and also functional tissues for implantation; 2) novel light-based biofabrication technologies; 3) development of new photo-responsive bioinks or biomaterial inks.

Bone regeneration of critical-sized bone defects, bone fractures or joint replacements remains a significant clinical challenge. Although there has been rapid advancement in both the fields of bone tissue engineering and additive manufacturing, functional bone implants with rapid vascularization capacity to ensure osseointegration and long-term biological fixation in large bone defects remains limited in clinics. In this study, we developed an in vitro vascularized bone implant by combining cell-laden hydrogels with direct metal printed (DMP) porous titanium alloys (Ti–6Al–4V). A 5 wt% allylated gelatin (GelAGE), was utilized to co-encapsulate human mesenchymal stromal cells (hMSCs) and human umbilical vein endothelial cells (HUVECs) to investigate concurrent osteogenic and vasculogenic performance. DMP macro-porous Ti–6Al–4V scaffolds were subsequently infused/enriched with cell-laden GelAGE to examine the feasibility to deliver cells and engineer vascular-like networks in the hybrid implant. Furthermore, as a proof of concept, a full-scale porous Ti–6Al–4V acetabular cup was impregnated with cell-laden hydrogel to validate the clinical potential of this strategy. The vasculogenic potential was evaluated by examining micro-capillary formation coupled with capillary network maturation and stabilization. Osteogenic differentiation was assessed via alkaline phosphatase activity as well as osteocalcin and osteopontin expression. Our results suggested that GelAGE supported HUVECs spreading and vascular-like network formation, along with osteogenesis of hMSCs. Titanium hybrid constructs with cell-laden hydrogel demonstrated enhanced osteogenesis with similar vasculogenic capability compared to the cell-laden hydrogel alone constructs. The full-scale implant with cell-laden hydrogel coating similarly showed cell distribution and spreading, implying the potential for further clinical application. Our study presents the feasibility of integrating bio-functional hydrogels with porous titanium implants to fabricate a vascularized hybrid construct with both mechanical support and preferable biological functionality (osteogenesis/vasculogenesis), which paves the way for improved strategies to enhance bone regeneration in complex large bone defects achieving long-term bone-implant fixation.

Recently developed modular bioassembly techniques hold tremendous potential in tissue engineering and regenerative medicine, due to their ability to recreate the complex microarchitecture of native tissue. Here, we developed a novel approach to fabricate hybrid tissue-engineered constructs adopting high-throughput microfluidic and 3D bioassembly strategies. Osteochondral tissue fabrication was adopted as an example in this study, because of the challenges in fabricating load bearing osteochondral tissue constructs with phenotypically distinct zonal architecture. By developing cell-instructive chondrogenic and osteogenic bioink microsphere modules in high-throughput, together with precise manipulation of the 3D bioassembly process, we successfully fabricated hybrid engineered osteochondral tissue in vitro with integrated but distinct cartilage and bone layers. Furthermore, by encapsulating allogeneic umbilical cord blood-derived mesenchymal stromal cells, and demonstrating chondrogenic and osteogenic differentiation, the hybrid biofabrication of hydrogel microspheres in this 3D bioassembly model offers potential for an off-the-shelf, single-surgery strategy for osteochondral tissue repair.

Bioprinting of living cells is rapidly developing as an advanced biofabrication approach to engineer tissues. Bioinks can be extruded in three-dimensions (3D) to fabricate complex and hierarchical constructs for implantation. However, lack of functionality can often be attributed to poor bioink properties. Indeed, advanced bioinks encapsulating living cells should: (i) present optimal rheological properties and retain 3D structure post-fabrication, (ii) promote cell viability and support cell differentiation, (iii) localise proteins of interest (e.g. vascular endothelial growth factor (VEGF)) to stimulate encapsulated cell activity and tissue ingrowth upon implantation. In this study, we present the results of the inclusion of a synthetic nanoclay, Laponite (LPN) together with a gelatin methacryloyl (GelMA) bioink and the development of a functional cell-instructive bioink. A nanocomposite bioink displaying enhanced shape fidelity retention and interconnected porosity within extrusion-bioprinted fibres was observed. Human bone marrow stromal cell (HBMSC) viability within the nanocomposite showed no significant changes over 21 days of culture in LPN-GelMA (85.60 ± 10.27 %), compared to a significant decrease in GelMA from 7 (95.88 ± 2.90 %) to 21 days (55.54 ± 14.72 %) (p

Bottom-up biofabrication approaches combining micro-tissue fabrication techniques with extrusion-based 3D printing of thermoplastic polymer scaffolds are emerging strategies in tissue engineering. These biofabrication strategies support native self-assembly mechanisms observed in developmental stages of tissue or organoid growth as well as promoting cell-cell interactions and cell differentiation capacity. Few technologies have been developed to automate the precise assembly of micro-tissues or tissue modules into structural scaffolds. We describe an automated 3D bioassembly platform capable of fabricating simple hybrid constructs via a two-step bottom-up bioassembly strategy, as well as complex hybrid hierarchical constructs via a multistep bottom-up bioassembly strategy. The bioassembly system consisted of a fluidic-based singularisation and injection module incorporated into a commercial 3D bioprinter. The singularisation module delivers individual micro-tissues to an injection module, for insertion into precise locations within a 3D plotted scaffold. To demonstrate applicability for cartilage tissue engineering, human chondrocytes were isolated and micro-tissues of 1 mm diameter were generated utilising a high throughput 96-well plate format. Micro-tissues were singularised with an efficiency of 96.0 ± 5.1%. There was no significant difference in size, shape or viability of micro-tissues before and after automated singularisation and injection. A layer-by-layer approach or aforementioned bottom-up bioassembly strategy was employed to fabricate a bilayered construct by alternatively 3D plotting a thermoplastic (PEGT/PBT) polymer scaffold and inserting pre-differentiated chondrogenic micro-tissues or cell-laden gelatin-based (GelMA) hydrogel micro-spheres, both formed via high-throughput fabrication techniques. No significant difference in viability between the construct assembled utilising the automated bioassembly system and manually assembled construct was observed. Bioassembly of pre-differentiated micro-tissues as well as chondrocyte-laden hydrogel micro-spheres demonstrated the flexibility of the platform while supporting tissue fusion, long-term cell viability, and deposition of cartilage-specific extracellular matrix proteins. This technology provides an automated and scalable pathway for bioassembly of both simple and complex 3D tissue constructs of clinically relevant shape and size, with demonstrated capability to facilitate direct spatial organisation and hierarchical 3D assembly of micro-tissue modules, ranging from biomaterial free cell pellets to cell-laden hydrogel formulations.