Influence of Breath-Mimicking Ventilated Incubation on Three-Dimensional Bioprinted Respiratory Tissue Scaffolds.

Development of respiratory tissue constructs is challenging due to the complex structure of native respiratory tissue and the unique biomechanical conditions induced by breathing. While studies have shown that the inclusion of biomechanical stimulus mimicking physiological conditions greatly benefits the development of engineered tissues, to our knowledge no studies investigating the influence of biomechanical stimulus on the development of respiratory tissue models produced through three-dimensional (3D) bioprinting have been reported. This paper presents a study on the utilization of a novel breath-mimicking ventilated incubator to impart biomechanical stimulus during the culture of 3D respiratory bioprinted constructs. Constructs were bioprinted using an alginate/collagen hydrogel containing human primary pulmonary fibroblasts with further seeding of human primary bronchial epithelial cells. Biomechanical stimulus was then applied via a novel ventilated incubator capable of mimicking the pressure and airflow conditions of multiple breathing conditions: standard incubation, shallow breathing, normal breathing, and heavy breathing, over a two-week time period. At time points between 1 and 14 days, constructs were characterized in terms of mechanical properties, cell proliferation, and morphology. The results illustrated that incubation conditions mimicking normal and heavy breathing led to greater and more continuous cell proliferation and further indicated a more physiologically relevant respiratory tissue model.

Synthesis of Alginate/Collagen Bioink for Bioprinting Respiratory Tissue Models.

Synthesis of bioinks for bioprinting of respiratory tissue requires considerations related to immunogenicity, mechanical properties, printability, and cellular compatibility. Biomaterials can be tailored to provide the appropriate combination of these properties through the synergy of materials with individual pros and cons. Sodium alginate, a water-soluble polymer derived from seaweed, is a cheap yet printable biomaterial with good structural properties; however, it lacks physiological relevance and cell binding sites. Collagen, a common component in the extra cellular matrix of many tissues, is expensive and lacks printability; however, it is highly biocompatible and exhibits sites for cellular binding. This paper presents our study on the synthesis of bioinks from alginate and collagen for use in bioprinting respiratory tissue models. Bioinks were synthesized from 40 mg/mL (4%) alginate and 3 mg/mL (0.3%) collagen in varying ratios (1:0, 4:1, 3:1, 2:1, and 1:1); then examined in terms of rheological properties, printability, compressive, and tensile properties and cellular compatibility. The results illustrate that the ratio of alginate to collagen has a profound impact on bioink performance and that, among the examined ratios, the 3:1 ratio is the most appropriate for use in bioprinting respiratory tissue scaffolds.

Development of a Nanoparticle System for Controlled Release in Bioprinted Respiratory Scaffolds.

The use of nanoparticle systems for the controlled release of growth factors is a promising approach to mimicking of the biochemical environment of native tissues in tissue engineering. However, sustaining growth factor release inside an appropriate therapeutic window is a challenge, particularly in bioprinted scaffolds. In this study, a chitosan-coated alginate-based nanoparticle system loaded with hepatocyte growth factor was developed and then incorporated into bioprinted scaffolds. The release kinetics were investigated with a focus on identifying the impact of the chitosan coating and culture conditions. Our results demonstrated that the chitosan coating decreased the release rate and lessened the initial burst release, while culturing in dynamic conditions had no significant impact compared to static conditions. The nanoparticles were then incorporated into bioinks at various concentrations, and scaffolds with a three-dimensional (3D) structure were bioprinted from the bioinks containing human pulmonary fibroblasts and bronchial epithelial cells to investigate the potential use of a controlled release system in respiratory tissue engineering. It was found that the bioink loaded with a concentration of 4 µg/mL of nanoparticles had better printability compared to other concentrations, while the mechanical stability of the scaffolds was maintained over a 14-day culture period. The examination of the incorporated cells demonstrated a high degree of viability and proliferation with visualization of the beginning of an epithelial barrier layer. Taken together, this study demonstrates that a chitosan-coated alginate-based nanoparticle system allows the sustained release of growth factors in bioprinted respiratory tissue scaffolds.

Investigation into relationships between design parameters and mechanical properties of 3D printed PCL/nHAp bone scaffolds.

BACKGROUND: Scaffolds are of great importance in tissue engineering applications as they provide a mechanically supportive environment for cellular activity, which is particularly necessary for hard tissues such as bone. Notably, the mechanical properties of a scaffold vary with differing design parameters such as those related to scaffold height and internal structure. Thus, the present study aimed to explore the relationship between design parameters and mechanical properties of composite polycaprolactone (PCL) and nano-hydroxyapatite (nHAp) scaffolds fabricated by three-dimensional (3D) printing. METHODS: We designed and printed scaffolds with different internal structures (lattice and staggered) and varying heights (4, 6, 8 and 10 layers), and consistent porosity (50%) for the purpose of comparison. Then, we examined the scaffold microstructure (pore size and penetration between layers) using scanning electron microscopy (SEM) and mechanical properties (elastic modulus and yield strength) using compressive testing. RESULTS: Our results illustrated that the microstructural parameters were related to scaffold design. At higher heights, pore size increased while penetration between layers decreased; thus, mechanical properties were affected. Results of mechanical testing demonstrated that for lattice scaffolds, elastic modulus was similar for 6 vs 4, and 8 vs 4 layers but ~33% lower for 10 layers vs 4 layers. Similarly, yield strength was comparable for 6 vs 4, and 8 vs 4 layers but ~27% lower for 10 layers vs 4 layers. With staggered scaffolds, when compared to 4-layer results, elastic modulus was similar for 6 layers but was ~43% lower for 8 layers and ~38% lower for 10 layers. Staggered scaffolds had ~38%, ~51%, and ~76% lower yield strength when the number of layers were increased from 4 to 6, 8, and 10 layers, respectively. When comparing lattice and staggered scaffolds with the same layer number, elastic modulus was similar, apart from 8-layer scaffolds where the staggered design was ~42% lower than lattice. Yield strength was similar between 4-layer staggered and lattice scaffolds, while staggered scaffolds with 6, 8, and 10 number of layers showed ~43%, ~45%, ~68% lower strength, respectively, than those found in lattice scaffolds with the same layer numbers. CONCLUSIONS: Mechanical properties of 3D printed scaffolds depended on scaffold height for both lattice and staggered internal structures. Staggered scaffolds had lower mechanical properties than the lattice scaffolds with the same height and were more sensitive to the change in scaffold height. Taken together, lattice scaffolds demonstrated the advantages of more stable mechanical properties over staggered scaffolds. Also, scaffolds with lower height were more promising in terms of mechanical properties compared to scaffolds with greater height.

Radiochemical, Computational, and Spectroscopic Evaluation of High-Denticity Desferrioxamine Derivatives DFO2 and DFO2p toward an Ideal Zirconium-89 Chelate Platform.

Desferrioxamine (DFO) has long been considered the gold standard chelator for incorporating [89Zr]Zr4+ in radiopharmaceuticals for positron emission tomography (PET) imaging. To improve the stability of DFO with zirconium-89 and to expand its coordination sphere to enable binding of large therapeutic radiometals, we have synthesized the highest denticity DFO derivatives to date: dodecadentate DFO2 and DFO2p. In this study, we describe the synthesis and characterization of a novel DFO-based chelator, DFO2p, which is comprised of two DFO strands connected by an p-NO2-phenyl linker and therefore contains double the chelating moieties of DFO (potential coordination number up to 12 vs 6). The chelator DFO2p offers an optimized synthesis comprised of only a single reaction step and improves water solubility relative to DFO2, but the shorter linker reduces molecular flexibility. Both DFO2 and DFO2p, each with 6 potential hydroxamate ligands, are able to reach a more energetically favorable 8-coordinate environment for Zr(IV) than DFO. The zirconium(IV) coordination environment of these complexes were evaluated by a combination of density functional theory (DFT) calculations and synchrotron spectroscopy (extended X-ray absorption fine structure), which suggest the inner-coordination sphere of zirconium(IV) to be comprised of the outermost four hydroxamate ligands. These results also confirm a single Zr(IV) in each chelator, and the hydroxide ligands which complete the coordination sphere of Zr(IV)-DFO are absent from Zr(IV)-DFO2 and Zr(IV)-DFO2p. Radiochemical stability studies with zirconium-89 revealed the order of real-world stability to be DFO2 > DFO2p ≫ DFO. The zirconium-89 complexes of these new high-denticity chelators were found to be far more stable than DFO, and the decreased molecular flexibility of DFO2p, relative to DFO2, could explain its decreased stability, relative to DFO2.

3D printing PCL/nHA bone scaffolds: exploring the influence of material synthesis techniques.

BACKGROUND: It is known that a number of parameters can influence the post-printing properties of bone tissue scaffolds. Previous research has primarily focused on the effect of parameters associated with scaffold design (e.g., scaffold porosity) and specific scaffold printing processes (e.g., printing pressure). To our knowledge, no studies have investigated variations in post-printing properties attributed to the techniques used to synthesize the materials for printing (e.g., melt-blending, powder blending, liquid solvent, and solid solvent). METHODS: Four material preparation techniques were investigated to determine their influence on scaffold properties. Polycaprolactone/nano-hydroxyapatite 30% (wt.) materials were synthesized through melt-blending, powder blending, liquid solvent, and solid solvent techniques. The material printability and the properties of printed scaffolds, in terms of swelling/degradation, mechanical strength, morphology, and thermal properties, were examined and compared to one another using Kruskal-Wallis nonparametric statistical analysis. RESULTS: Material prepared through the liquid solvent technique was found to have limited printability, while melt-blended material demonstrated the highest degree of uniformity and lowest extent of swelling and degradation. Scaffolds prepared with powder-blended material demonstrated the highest Young's modulus, yield strength, and modulus of resilience; however, they also demonstrated the highest degree of variability. The higher degree of inhomogeneity in the material was further supported by thermal gravimetric analysis. While scaffolds printed from melt-blended, powder-blended, and solid solvent materials demonstrated a high degree of micro-porosity, the liquid solvent material preparation technique resulted in minimal micro-porosity. CONCLUSIONS: Study results indicate that specific techniques used to prepare materials influence the printing process and post-printing scaffold properties. Among the four techniques examined, melt-blended materials were found to be the most favorable, specifically when considering the combination of printability, consistent mechanical properties, and efficient preparation. Techniques determined to be favourable based on the properties investigated should undergo further studies related to biological properties and time-dependent properties beyond 21-days.

Innovation and possible long-term impact driven by COVID-19: Manufacturing, personal protective equipment and digital technologies.

It is known that discrete events causing extreme societal and economic pressures as well as technological opportunity are major driving factors of innovation. Due to the presence of both of these factors during the COVID-19 pandemic it was hypothesized that there would be significant on-going innovation throughout society during the pandemic, with many of the innovations having the ability to have long-term societal impact. Analysis of literature and patent databases determined sectors of accelerated innovation to include manufacturing, personal protective equipment and digital technologies. The ability of flexible and advanced manufacturing technologies to provide more adaptable production capabilities that are less susceptible to disruption, make it likely that these technologies will be incorporated further, changing the way many manufacturing firms operate. Collaboration has increased, demonstrating increases in problem-solving efficiency; however, concerns around intellectual property is likely to reduce the long-term impact of these procedural changes. Advancements in personal protective equipment and disinfection technologies may have the long-term impact of reducing waste production and triggering changes in cleaning protocols throughout society. Digital technologies such as telemedicine, data collection, artificial intelligence and communication technologies were found to have undergone significant innovation, with possible impacts such as large-scale systemic shifts, and changes in how governments, corporations, the scientific community and the public interact.

Structural Characterization of the Solution Chemistry of Zirconium(IV) Desferrioxamine: A Coordination Sphere Completed by Hydroxides.

Positron emission tomography (PET) using radiolabeled, monoclonal antibodies has become an effective, noninvasive method for tumor detection and is a critical component of targeted radionuclide therapy. Metal ion chelator and bacterial siderophore desferrioxamine (DFO) is the gold standard compound for incorporation of zirconium-89 in radiotracers for PET imaging because it is thought to form a stable chelate with [89Zr]Zr4+. However, DFO may not bind zirconium-89 tightly in vivo, with free zirconium-89 reportedly liberated into the bones of experimental mouse models. Although high bone uptake has not been observed to date in humans, this potential instability has been proposed to be related to the unsaturated coordination sphere of [89Zr]Zr-DFO, which is thought to consist of the 3 hydroxamate groups of DFO and 1 or 2 water molecules. In this study, we have used a combination of X-ray absorption spectroscopy and density functional theory (DFT) geometry optimization calculations to further probe the coordination chemistry of this complex in solution. We find the extended X-ray absorption fine structure (EXAFS) curve fitting of an aqueous solution of Zr(IV)-DFO to be consistent with an 8-coordinate Zr with oxygen ligands. DFT calculations suggest that the most energetically favorable Zr(IV) coordination environment in DFO likely consists of the 3 hydroxamate ligands from DFO, each with bidentate coordination, and 2 hydroxide ligands. Further EXAFS curve fitting provides additional support for this model. Therefore, we propose that the coordination sphere of Zr(IV)-DFO is most likely completed by 2 hydroxide ligands rather than 2 water molecules, forming Zr(DFO)(OH)2.

Bioprinting for combating infectious diseases.

Infectious diseases have the ability to impact health on a global scale, as is being demonstrated by the current coronavirus disease 2019 (COVID-19) pandemic. The strenuous circumstances related to this global health crisis have been highlighting the challenges faced by the biomedical field in combating infectious diseases. Notably, printing technologies have advanced rapidly over the last decades, allowing for the incorporation of living cells in the printing process (or bioprinting) to create constructs that are able to serve as in vitro tissue or virus-disease models in combating infectious diseases. This paper describes applications of bioprinting in addressing the challenges faced in combating infectious diseases, with a specific focus on in vitro modelling and on development of therapeutic agents and vaccines. Integration of these technologies may allow for a more efficient and effective response to current and future pandemics.