Special Issue "Biomaterials and Bioprinting".

The emergence of bioprinting in recent years represents a marvellous advancement in 3D printing technology. It expands the range of 3D printable materials from the world of non-living materials into the world of living materials. Biomaterials play an important role in this paradigm shift. This Special Issue focuses on biomaterials and bioprinting and contains eight articles covering a number of recent topics in this emerging area.

A Solvent-Free Surface Suspension Melt Technique for Making Biodegradable PCL Membrane Scaffolds for Tissue Engineering Applications.

In tissue engineering, there is limited availability of a simple, fast and solvent-free process for fabricating micro-porous thin membrane scaffolds. This paper presents the first report of a novel surface suspension melt technique to fabricate a micro-porous thin membrane scaffolds without using any organic solvent. Briefly, a layer of polycaprolactone (PCL) particles is directly spread on top of water in the form of a suspension. After that, with the use of heat, the powder layer is transformed into a melted layer, and following cooling, a thin membrane is obtained. Two different sizes of PCL powder particles (100 µm and 500 µm) are used. Results show that membranes made from 100 µm powders have lower thickness, smaller pore size, smoother surface, higher value of stiffness but lower ultimate tensile load compared to membranes made from 500 µm powder. C2C12 cell culture results indicate that the membrane supports cell growth and differentiation. Thus, this novel membrane generation method holds great promise for tissue engineering.

Thermal Influence of CNT on the Polyamide 12 Nanocomposite for Selective Laser Sintering.

The thermal influence of carbon nanotubes (CNTs) on the PA12 in the laser sintering process was assessed by physical experiments and a three dimensional simulation model. It appears that, by adding the CNTs into the PA12 matrix, the thermal conductivity increased. A double ellipsoidal heat flux model was applied to input a three dimensional, continuous moving, volumetric laser heat source. The predicted three dimensional temperature distributions suggested that the laser heat was conducted wider and deeper in the PA12-CNT sample than PA12. Greater heat conduction can reduce the interspace between two successive layers, and result in the increase of the parts' density and properties.

A 3D biomimetic model of tissue stiffness interface for cancer drug testing.

Contrary to oversimplified preclinical drug screens that derive treatment responses of cancer cells grown on plastic cell culturing surfaces, the actual in vivo scenario for cancer cell invasion is confronted with a diversity of tissue stiffness. After all, the packing of organs and tissues in the body translates to the abundant presence of tissue stiffness interfaces. The invasive dissemination of cancer cells in vivo might be encouraged by favorable tissue stiffness gradients, likely explaining the preferential spread of cancer cells which is subjective to the cancer type and origin of the primary site. Yet these critical tumor microenvironmental influences cannot be recapitulated in 2D preclinical drug screens, hence omitting potentially invaluable in vivo patterns of drug responses that may support safer clinical dosage implementation of cancer drugs. Current attempts to study stiffness implications on cancer cells are largely confined to 2D surfaces of tunable stiffness. While these studies collectively show that cancer cells migrate better on a stiffer matrix, the generation of a biomimetic 3D tissue stiffness interface for cancer cell migration would clearly give a more definitive understanding on the probable push and pull influences of the 3D ECM. Herein, we developed a biomimetic platform which enables the precise placement of spheroids at tissue stiffness interfaces constructed with natural ECM collagen type I. This enables a standardized comparison of spheroid invasion under a 3D stiffness gradient influence. We found that cancer cells in 3D infiltrated more extensively into a softer matrix of 300 Pa while showing significantly reduced invasion into stiffer matrix of 1200 and 6000 Pa. These biomimetic spheroid cultures postinvasion were suitably subjected to paclitaxel treatment and subsequent daily live quantification of apoptotic cells to evaluate the implications of tissue stiffness on chemotherapeutic treatment. We importantly found that cancer cells which more extensively infiltrated the 300 Pa matrix also succumbed to paclitaxel induced apoptosis earlier than cells in stiffer matrices of 1200 and 6000 Pa respectively. This suggests that reduced invasion of cancer cells attributed to increased tissue stiffness barriers may favor their reduced apoptotic susceptibility to chemotherapeutic treatment.

Pressurized Hot Water Extraction of Mangosteen Pericarp and Its Associated Molecular Signatures in Endothelial Cells.

The mangosteen (Garcinia mangostana L.) pericarp is known to be rich in potent bioactive phytochemical compounds such as xanthones, which possess pharmacologically important antioxidant activity and beneficial cardiometabolic properties. Mangosteen pericarp is typically classified as unavoidable food waste and discarded, despite being rich in bioactive phytochemical compounds that therefore present an exciting opportunity for valorization. Thus, this study aims to extract phytochemical compounds from mangosteen pericarp using pressurized hot water extraction (PHWE) and determine its biological effects in endothelial cells using RNA sequencing. Liquid chromatography with MS/MS (LC/MSMS) and UV detection (LC/UV) was subsequently used to identify three key phytochemical compounds extracted from the mangosteen pericarp: α-Mangostin, γ-Mangostin, and Gartanin. Within the tested range of extraction temperatures by PHWE, our results demonstrated that an extraction temperature of 120 °C yielded the highest concentrations of α-Mangostin, γ-Mangostin, and Gartanin with a concomitant improvement in antioxidant capacity compared to other extraction temperatures. Using global transcriptomic profiling and bioinformatic analysis, the treatment of endothelial cells with mangosteen pericarp extracts (120 °C PHWE) for 48 h caused 408 genes to be differentially expressed. Furthermore, our results demonstrated that key biological processes related to "steroid biosynthesis and metabolism", likely involving the activation of the AMPK signaling pathway, were upregulated by mangosteen pericarp extract treatment. In conclusion, our study suggests a green extraction method to valorize phytochemical compounds from mangosteen pericarp as a natural product with potential beneficial effects on cardiometabolic health.

Solvent-free fabrication of three dimensionally aligned polycaprolactone microfibers for engineering of anisotropic tissues.

Fabrication of aligned microfiber scaffolds is critical in successful engineering of anisotropic tissues such as tendon, ligaments and nerves. Conventionally, aligned microfiber scaffolds are two dimensional and predominantly fabricated by electrospinning which is solvent dependent. In this paper, we report a novel technique, named microfiber melt drawing, to fabricate a bundle of three dimensionally aligned polycaprolactone microfibers without using any organic solvent. This technique is simple yet effective. It has been demonstrated that polycaprolactone microfibers of 10 µm fiber diameter can be directly drawn from a 2 mm orifice. Orifice diameter, temperature and take-up speed significantly influence the final linear density and fiber diameter of the microfibers. Mechanical test suggests that mechanical properties such as stiffness and breaking force of microfiber bundles can be easily adjusted by the number of fibers. In vitro study shows that these microfibers are able to support the proliferation of human dermal fibroblasts over 7 days. In vivo result of Achilles tendon repair in a rabbit model shows that the microfibers were highly infiltrated by tendon tissue as early as in 1 month, besides, the repaired tendon have a well-aligned tissue structure under the guidance of aligned microfibers. However whether these three dimensionally aligned microfibers can induce three dimensionally aligned cells remains inconclusive.

A Deep Learning Quality Control Loop of the Extrusion-based Bioprinting Process.

Extrusion-based bioprinting (EBB) represents one of the most used deposition technologies in the field of bioprinting, thanks to key advantages such as the easy-to-use hardware and the wide variety of materials that can be successfully printed. In recent years, research efforts have been focused on implementing a quality control loop for EBB, which can reduce the trial-and-error process necessary to optimize the printing parameters for a specific ink, standardize the results of a print across multiple laboratories, and so accelerate the translation of extrusion bioprinted products to more impactful clinical applications. Due to its capacity to acquire relevant features from a training dataset and generalize to unseen data, machine learning (ML) is currently being studied in literature as a relevant enabling technology for quality control in EBB. In this context, we propose a robust, deep learning-based control loop to automatically optimize the printing parameters and monitor the printing process online. We collected a comprehensive dataset of EBB prints by recording the process with a high-resolution webcam. To model multiple printing scenarios, each video represents a combination of multiple parameters, including printing set-up (either mechanical or pneumatic extrusion), material color, layer height, and infill density. After pre-processing, the collected dataset was used to thoroughly train and evaluate an ad hoc defined convolutional neural network by controlling over-fitting and optimizing the prediction time of the network. Finally, the ML model was used in a control loop to: (i) monitor the printing process and detect if a print with an error could be stopped before completion to save material and time and (ii) automatically optimize the printing parameters by combining the ML model with a previously published mathematical model of the EBB process. Altogether, we demonstrated for the first time how ML techniques can be used to automatize the EBB process, paving the way for a total quality control loop of the printing process.

Three-Dimensional Printing Technologies for Drug Delivery Applications: Processes, Materials, and Effects.

Since the 1930s, new methods of drug delivery, such as implantable devices with drug release control, have been developed. However, manufacturing techniques require bulk due to high initial production costs. Three-dimensional (3D) printing, also known as additive manufacturing or rapid prototyping, allows the fabrication of personalized drug delivery that uses different materials and complex geometries with multiple release profiles, thereby eradicating high initial costs. Different studies have been developed showing the extensive potential of 3D printing for the pharmaceutical industry, and despite in-depth discussions that have been published, there is no comprehensive review of processes, materials, and effects in drug delivery applications thus far. This review aims to fill this gap by presenting the use of 3D printing technology for drug delivery, exposing the different variations of the technique according to the characteristics, material, and dosage form sought. There are seven main categories of 3D printing according to the standards jointly developed by International Organization for Standardization and American Society for Testing and Materials: material jetting, binder jetting, material extrusion, vat photopolymerization, powder bed fusion, sheet lamination, and directed energy deposition. There are different 3D fabrication processes used for drug delivery applications depending on the dosage form and material applied. In this context, polymers, glasses, and hydrogels represent the most frequent materials used. 3D printing allows different forms of drug dosage. Oral, topical, rectal and vaginal, parental and implantable are discussed in this paper, presenting the identification of the type of 3D printing technology, the active pharmaceutical ingredient, formulation, and pharmaceutical effect. The main aim of this paper is to offer insights to people from academy and industry who are interested in the advancement of drug delivery and in knowing the future directions in the development of 3D printing applications in this area.

Green Extraction of Orange Peel Waste Reduces TNFα-Induced Vascular Inflammation and Endothelial Dysfunction.

Orange peel waste (OPW) is known to contain an abundant amount of polyphenols compounds such as flavonoids, well-reported for their antioxidant and anti-inflammatory properties. While OPW is generally regarded as a food waste, the opportunity to extract bioactive compounds from these "wastes" arises due to their abundance, allowing the investigation of their potential effects on endothelial cells. Hence, this study aims to use a green extraction method and pressurized hot water extraction (PHWE) to extract bioactive compounds from OPW. Liquid chromatography with UV detection (LC/UV) and liquid chromatography mass spectrometry (LC/MS) were subsequently used to identify the bioactive compounds present. Through the optimization of the extraction temperature for PHWE, our results demonstrated that extraction temperatures of 60 °C and 80 °C yield distinct bioactive compounds and resulted in better antioxidant capacity compared to other extraction temperatures or organic solvent extraction. Despite having similar antioxidant capacity, their effects on endothelial cells were distinct. Specifically, treatment of endothelial cells with 60 °C OPW extracts inhibited TNFα-induced vascular inflammation and endothelial dysfunction in vitro, suggesting that OPW possess vasoprotective effects likely mediated by anti-inflammatory effects.

Application of Machine Learning in 3D Bioprinting: Focus on Development of Big Data and Digital Twin.

The application of machine learning (ML) in bioprinting has attracted considerable attention recently. Many have focused on the benefits and potential of ML, but a clear overview of how ML shapes the future of three-dimensional (3D) bioprinting is still lacking. Here, it is proposed that two missing links, Big Data and Digital Twin, are the key to articulate the vision of future 3D bioprinting. Creating training databases from Big Data curation and building digital twins of human organs with cellular resolution and properties are the most important and urgent challenges. With these missing links, it is envisioned that future 3D bioprinting will become more digital and in silico, and eventually strike a balance between virtual and physical experiments toward the most efficient utilization of bioprinting resources. Furthermore, the virtual component of bioprinting and biofabrication, namely, digital bioprinting, will become a new growth point for digital industry and information technology in future.

Fouling mitigation in reverse osmosis processes with 3D printed sinusoidal spacers.

Feed spacers are an essential part of spiral wound modules for reverse osmosis (RO). They create flow channels between membrane sheets and manipulate hydrodynamic conditions to control membrane fouling. In this work, additive manufacturing (Polyjet) was used to print novel sinusoidal spacers with wavy axial filaments connected by perpendicular (ST) or slanted (SL) transverse filaments. When tested with 2 g/L NaCl solution, conventional and SL spacers had similar flux while the ST spacer had about 5-7% lower flux. The pressure losses for ST and SL spacers increased by up to 3 folds depending on the flow condition. In the colloidal silica fouling and biofouling tests, the sinusoidal spacers showed lower membrane permeability decrease of 46% for ST, 41% for SL vs 56% for conventional and 26% for ST, 22% for SL vs 33% for conventional, respectively. Optical coherence tomography images from colloidal silica fouling and confocal images from biofouling tests revealed that fouling patterns were closely associated with the local hydrodynamic conditions. Overall, sinusoidal spacers showed promising results in controlling membrane fouling, but there is potential for further optimizations to reduce channel pressure loss.

A review on spacers and membranes: Conventional or hybrid additive manufacturing?

Over the past decade, 3D printing or additive manufacturing (AM) technology has seen great advancement in many aspects such as printing resolution, speed and cost. Membranes for water treatment experienced significant breakthroughs owing to the unique benefits of additive manufacturing. In particular, 3D printing's high degree of freedom in various aspects such as material and prototype design has helped to fabricate innovative spacers and membranes. However, there were conflicting reports on the feasibility of 3D printing, especially for membranes. Some research groups stated that technology limitations today made it impossible to 3D print membranes, but others showed that it was possible by successfully fabricating prototypes. This paper will provide a critical and comprehensive discussion on 3D printing specifically for spacers and membranes. Various 3D printing techniques will be introduced, and their suitability for membrane and spacer fabrication will be discussed. It will be followed by a review of past studies associated with 3D-printed spacers and membranes. A new category of additive manufacturing in the membrane water industry will be introduced here, known as hybrid additive manufacturing, to address the controversies of 3D printing for membrane. As AM technology continues to advance, its possibilities in the water treatment is limitless. Some insightful future trends will be provided at the end of the paper.

Three-Dimensional Printing of Food Foams Stabilized by Hydrocolloids for Hydration in Dysphagia.

Three-dimensional food printing offers the possibility of modifying the structural design, nutrition, and texture of food, which may be used for consumers with special dietary requirements such as dysphagic patients. One of the food matrices that can be used for liquid delivery to dysphagic patients is food foams. Foams are widely used in different food products to adjust food density, rheological properties, and texture. Foams allow the food to stay in the mouth for sufficient time to provide hydration while minimizing the danger of choking. Our work studies the foam properties and printability of both egg white foams and eggless foams with a strong focus on their foaming properties, rheological properties, printability, and suitability for dysphagic patients. Food hydrocolloid, xanthan gum (XG), is added to improve foam stability and rheological properties so that the inks are printable. Rheological and syneresis properties of the pre-printed foam inks are examined. The texture profile and microstructure properties are studied post-printing. International dysphagia diet standardization initiative tests are carried out to assess the inks' potential for dysphagic diets. Inks with XG performed better with minimal water seepage, better foam stability, and excellent printability. This suggests that hydrocolloids lead to more stable food foams that are suitable for 3DFP and safe for hydration delivery to dysphagic patients.

Fibroblast response to interstitial flow: A state-of-the-art review.

Interstitial flow (IF) modulates both the biochemical and biophysical cues surrounding cells. It represents a very important regulating mechanism for cell/tissue function and has been commonly utilized in tissue engineering (TE). This article discusses the possible regulating mechanisms of IF on fibroblasts, the various fibroblast responses to IF, the current challenges in understanding the IF-fibroblast relationship and the application of IF for fibroblast involved TE. In particular, IF can affect fibroblast growth at both intracellular (e.g., calcium signaling, protein/proteinase secretion) and cellular (e.g., autocrine/paracrine signaling, proliferation, differentiation, alignment, adhesion, migration) levels. One major challenge for understanding IF-fibroblast interaction has been the determination of the flow and cell growth condition at microlevel especially in a three-dimensional environment. To utilize IF and optimize the fluidic environment for TE, several influencing factors in the system including perfusate composition, flow profile, nutrient supply, signaling molecule effect, scaffold property, and fibroblast type should be considered.

Recent Advances on High-Entropy Alloys for 3D Printing.

Boosted by the success of high-entropy alloys (HEAs) manufactured by conventional processes in various applications, the development of HEAs for 3D printing has been advancing rapidly in recent years. 3D printing of HEAs gives rise to a great potential for manufacturing geometrically complex HEA products with desirable performances, thereby inspiring their increased appearance in industrial applications. Herein, a comprehensive review of the recent achievements of 3D printing of HEAs is provided, in the aspects of their powder development, printing processes, microstructures, properties, and potential applications. It begins with the introduction of the fundamentals of 3D printing and HEAs, as well as the unique properties of 3D-printed HEA products. The processes for the development of HEA powders, including atomization and mechanical alloying, and the powder properties, are then presented. Thereafter, typical processes for printing HEA products from powders, namely, directed energy deposition, selective laser melting, and electron beam melting, are discussed with regard to the phases, crystal features, mechanical properties, functionalities, and potential applications of these products (particularly in the aerospace, energy, molding, and tooling industries). Finally, perspectives are outlined to provide guidance for future research.

3D Printing of Polymeric Multi-Layer Micro-Perforated Panels for Tunable Wideband Sound Absorption.

The increasing concern about noise pollution has accelerated the development of acoustic absorption and damping devices. However, conventional subtractive manufacturing can only fabricate absorption devices with simple geometric shapes that are unable to achieve high absorption coefficients in wide frequency ranges. In this paper, novel multi-layer micro-perforated panels (MPPs) with tunable wideband absorption are designed and fabricated by 3D printing or additive manufacturing. Selective laser sintering (SLS), which is an advanced powder-based 3D printing technique, is newly introduced for MPP manufacturing with polyamide 12 as the feedstock. The acoustic performances of the MPPs are investigated by theoretical, numerical, and experimental methods. The results reveal that the absorption frequency bandwidths of the structures are wider than those of conventional single-layer MPPs, while the absorption coefficients remain comparable or even higher. The frequency ranges can be tuned by varying the air gap distances and the inter-layer distances. Furthermore, an optimization method is introduced for structural designs of MPPs with the most effective sound absorption performances in the target frequency ranges. This study reveals the potential of 3D printing to fabricate acoustic devices with effective tunable sound absorption behaviors and provides an optimization method for future structural design of the wideband sound absorption devices.

The global rise of 3D printing during the COVID-19 pandemic.

3D printing enables on-demand solutions for a wide spectrum of needs ranging from personal protection equipment to medical devices and isolation wards. This versatile technology is suited to address supply-demand imbalances caused by socio-economic trends and disruptions in supply chains.

Layer-by-layer ultraviolet assisted extrusion-based (UAE) bioprinting of hydrogel constructs with high aspect ratio for soft tissue engineering applications.

One of the major challenges in the field of soft tissue engineering using bioprinting is fabricating complex tissue constructs with desired structure integrity and mechanical property. To accomplish such requirements, most of the reported works incorporated reinforcement materials such as poly(ϵ-caprolactone) (PCL) polymer within the 3D bioprinted constructs. Although this approach has made some progress in constructing soft tissue-engineered scaffolds, the mechanical compliance mismatch and long degradation period are not ideal for soft tissue engineering. Herein, we present a facile bioprinting strategy that combines the rapid extrusion-based bioprinting technique with an in-built ultraviolet (UV) curing system to facilitate the layer-by-layer UV curing of bioprinted photo-curable GelMA-based hydrogels to achieve soft yet stable cell-laden constructs with high aspect ratio for soft tissue engineering. GelMA is supplemented with a viscosity enhancer (gellan gum) to improve the bio-ink printability and shape fidelity while maintaining the biocompatibility before crosslinking via a layer-by-layer UV curing process. This approach could eventually fabricate soft tissue constructs with high aspect ratio (length to diameter) of ≥ 5. The effects of UV source on printing resolution and cell viability were also studied. As a proof-of-concept, small building units (3D lattice and tubular constructs) with high aspect ratio are fabricated. Furthermore, we have also demonstrated the ability to perform multi-material printing of tissue constructs with high aspect ratio along both the longitudinal and transverse directions for potential applications in tissue engineering of soft tissues. This layer-by-layer ultraviolet assisted extrusion-based (UAE) Bioprinting may provide a novel strategy to develop soft tissue constructs with desirable structure integrity.

Development of a 95/5 poly(L-lactide-co-glycolide)/hydroxylapatite and beta-tricalcium phosphate scaffold as bone replacement material via selective laser sintering.

95/5 Poly(L-lactide-co-glycolide) was investigated for the role of a porous scaffold, using the selective laser sintering (SLS) fabrication process, with powder sizes of 50-125 and 125-250 microm. SLS parameters of laser power, laser scan speed, and part bed temperature were altered and the degree of sintering was assessed by scanning electron microscope. Composites of the 125-250 microm polymer with either hydroxylapatite or hydroxylapatite/beta-tricalcium phosphate (CAMCERAM II were sintered, and SLS settings using 40 wt % CAMCERAM II were optimized for further tests. Polymer thermal degradation during processing led to a reduction in number and weight averaged molecular weight of 9% and 12%, respectively. Compression tests using the optimized composite sintering parameters gave a Young's modulus, yield strength, and strain at 1% strain offset of 0.13 +/- 0.03 GPa, 12.06 +/- 2.53 MPa, and 11.39 +/- 2.60%, respectively. Porosity was found to be 46.5 +/- 1.39%. CT data was used to create an SLS model of a human fourth middle phalanx and a block with designed porosity was fabricated to illustrate the process capabilities. The results have shown that this composite and fabrication method has potential in the fabrication of porous scaffolds for bone tissue engineering.

Special Issue: NextGen Materials for 3D Printing.

Only a handful of materials are well-established in three-dimensional (3D) printing and well-accepted in industrial manufacturing applications. However, recent advances in 3D printable materials have shown potential for enabling numerous novel applications in the future. This special issue, consisting of 2 reviews and 10 research articles, intends to explore the possible materials that could define next-generation 3D printing.