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Introduction
Originally, 3D printing technologies were designed for nonbiological materials, such as metals, ceramics and thermoplastic polymers (Murphy and Atala 6). However, recent advances have enabled 3D printing of biocompatible materials, cells, and supporting components into complex 3D functional living tissues, giving rise to a new technology referred to as ‘3D Bioprinting.’ However, unlike non-biological 3D printing, it involves important complexities, such as choice of materials, cell types, growth and differentiation factors, and technical challenges related to the sensitivities of living cells and the construction of tissues (Murphy and Atala 1). Overall, 3D Bioprinting is primarily based on three methodologies: biomimicry (reproduction of specific cellular components), autonomous self-assembly (intervention-free self-organization and self-patterning), and mini-tissue building blocks (expansive fabrication and assembly).
However, three main technologies had been used in depositing and patterning biological materials. These are inkjet, microextrusion, and laser-assisted bioprinting (Murphy and Atala 3). Inkjet Bioprinting uses modified inkjet printers that use thermal or acoustic forces to eject drops of liquid onto a substrate, which can support or form part of the final construct (Murphy and Atala 3-4). Microextrusion Bioprinting is the most common and affordable non-biological 3D printers. It uses robotically controlled extrusion of a biomaterial onto a substrate, yielding continuous beads of material rather than liquid droplets and resulting to accurately fabricate complex structures with multiple cell types (Murphy and Atala 5). Laser-assisted Bioprinting uses focused laser pulses to generate a high-pressure bubble that propels cell-containing materials towards the collector substrate (Murphy and Atala 5-6).
The paper will provide a concise background on the developmental history of the concept of Bioprinting, discuss its general technological process and specific methodological variations, and a brief run through of its current and potential applications in the field of medicine and allied fields handling biomaterials.
Background
The concept of the development of 3D Bioprinting follows three stages: image data preparation; bioprinter development, and; experimental studies on biofabrication (Henmi et al. 38). The first stage involves the transformation of the histological structure into 3D data. The second stage consists of the development of the bioprinter. The third stage demands experimental biofabrication of the 2D and 3D cell patterning into 3D structures that are useful in tissue and organ engineering. Moreover, the concept and technology of 3D Bioprinting can be traced far back in history, to the invention of woodblock printing and the later development of the industrial-scale printing press in the 15th century. The latter development resulted to the rapid reproduction of text and images and the dissemination of information.
The following appearance of the subtractive methods of manufacturing, which commonly involved a controlled material-removal process as had been classic in machining, opened a new dimension in printing technology that improved fabrication methodologies. Machining generate exact shapes of desired objects using high precision sculpturing, filing and turning parts through the process of milling and grinding.
Less than a century ago, printing technology has advanced from two-dimensional (2D) printing into three-dimensional (3D) printing. Charles Hull first described 3D printing in 1986 when he introduced a new technology called “stereolithography,” which used thin layers of a material treated with ultraviolet light to create a solid 3D structure (Murphy and Atala 1). The advent of 3D printing or additive methodology in manufacturing changed the landscape. Rapid prototyping of tools, for instance, reduced lead time and cost of developing prototypes of new parts and devices through the additive approach (3D Printing n. p.). Prototyping under the subtractive method was renowned for its slow and expensive procedure. Not long later, its potential in fabricating biological structures led into a new field in 3D printing called “3D Bioprinting.” One important challenge though was to adapt 3D printing technologies designed to print molten plastics and metals to the printing of sensitive, living biomaterials.
The crucial test, however, is to replicate the intricate micro-architecture of extracellular matrix components and multiple cell types in sufficient resolutions to review biological functions (Murphy and Atala 1). Eventually, the need for more precise and dependable bio-replication technology led to the functional modifications of available 3D printers to support the use of biological raw materials in the fabrication of living structures with wide-ranging applications in reconstructive medicine. The technology, for instance, of the laser-assisted Bioprinting came from industrial laser-assisted printed for transferring metals (Murphy and Atala 5).
Process
The 3D Bioprinting technology roughly follows the typical four-step 3D printing operational process: digital designing, printing preparation, design printing, and object creation.
Step 1: Designing the object. The object to be fabricated is designed virtually in a Computer Aided Design (CAD) file using either a 3D modeling program (for a totally new object) or a 3D scanner (to copy an existing object or its design). The scanner can place the copied object into a 3D modeling program.
Step 2: Preparation for printing. The 3D modeling program slices the final model into hundreds or thousands of horizontal layers. Each layer is seen as a 2D image.
Step 3: Printing the design. The prepared file is uploaded into the 3D printer, which creates the objects digitally layer by layer. The 3D printer reads every slice (layer) and proceeds to recreate the object by blending each layer together with no visible sign of the layering. A printed 3D structure comes off as the output.
Step 4: Creating the object. Different technologies, such as SLS (selective laser sintering), FDM (fused deposition modeling) and SLA (stereolithography), can create the object.
Selective laser sintering uses a high-power laser to fuse small particles of plastic, metal, ceramic or glass powders into a mass of three-dimensional structure. The laser scans the layers on the surface of a powder bed, selectively fusing the powdered material. Then a new layer of material is applied on top, followed by another layer until the object is completed. Fused deposition modeling uses a plastic filament or metal wire, which is unwound from a coil and supplies material to an extrusion nozzle that can turn the flow on and off. The nozzle is heated to melt the material and can be moved both horizontally and vertically by a numerically controlled mechanism that CAM software directly controls. The melted materials are extruded to form layers. The material hardens immediately after extrusion from the nozzle, creating the object. Stereolithography mainly uses photopolymerization to produce a solid part from a liquid. Using a vat of liquid ultraviolet (UV) curable photopolymer resin, a UV laser builds layers of the object one at a time. For each layer, the laser beam traces a cross-section (layer) of the part pattern on the surface of the liquid resin. Exposure to the ultraviolet laser light cures and solidifies the pattern traces on the resin and joins it to the previous layer. Then, the elevator platform descends a layer thick (typically 0.5 mm to 0.15 mm). A resin-filled blade sweeps across the layer, recoating it with fresh material. On this new liquid surface, the subsequent layer pattern is traced, joining the previous layer. Thus, the complete 3D object is created.
Meanwhile, Murphy and Atala (3-6) proposed a three-step 3D Bioprinting process, consisting of imaging and design, choice of materials and cells, and printing of the tissue construct. Step 1 (imaging and digital design) consists of two sub-steps, namely: imaging and design approach. Imaging involves the acquisition of detailed and contrasted structural 2D images, using any of the contemporary medical imaging technologies, such as computed tomography (CT) imaging (the most common modality) and magnetic resonance imaging (MRI), which provides information on the 3D structure and functions at the micro (cellular) and macro (organism) levels (Murphy and Atala 3). CT imaging produces closely spaced axial layers of tissue architecture fully describing the tissue volume after surface rendering and stereolithographic editing. Conversely, MRI can also provide high spatial resolution in soft tissue with increased contrast resolution, making it more effective for imaging soft tissues in close proximity to each other without exposure to the ionizing radiation in CT. Contrasts in biological structures greatly increase with the use of barium or iodine for CT scans and iron oxide, gadolinium or metalloproteins for MRI. Digital design involves the use of computer-aided design and computer-aided manufacturing (CAD-CAM) tools and mathematical modeling to collect and digitize the intricate tomographic and architectural information into 3D images viewable in multiple ways. Then the completed structural model gets interfaced with numerically controlled Bioprinting systems for the prototyping and manufacturing of multi-layered 3D structured design. Transformation of the 2D images may use any of the 3D Bioprinting methodologies of biomimicry, self-assembly, mini-tissues, or a combination thereof.
Step 2 (materials and cells selection) involves two sub-steps: material selection and cell selection. Material selection consists of screening biomaterials based on five ideal properties for 3D Bioprinting in such categories as printability, biocompatibility, degradation kinetics and byproducts, structural and mechanical properties, and material biomimicry (Murphy and Atala 6). Biomaterials must be easy for the bioprinter to handle and deposit, do not induce undesirable responses from the host (local or systemic), degradation rates must be less as fast as the cells’ ability to produce their own ECM, must be compatible with the required mechanical properties of the construct, and has desirable endogenous material composition. Cells selections depend largely upon desirable sources as it is crucial for correct functioning of the fabricated structure (Murphy and Atala 8). Specific and essential biological functions must be replicated in the transplanted structure. Selected cells must closely mimic the physiological state of cells in vivo and maintain their in vivo functions under optimized conditions. Moreover, they must be expandable into sufficient numbers for bioprinting.
Step 3 (structure construct printing) uses any of the three main bioprinting technologies: inkjet, microextrusion, and laser-assisted bioprinting. Inkjet Bioprinting consists of two modalities: inkjet and thermal (Murphy and Atala 3). Thermal inkjet bioprinters use a print head that heats between 200 OC and 300 OC with high print speed and low cost. Acoustic inkjet bioprinters contain a piezoelectric crystal that creates an acoustic wave inside the print head to break the liquid into droplets at regular intervals, resulting into uniform droplet size (Murphy and Atala 4). Microextrusion Bioprinting involves robotically controlled extrusion of biomaterials and deposition onto a substrate by a microextrusion head (Murphy and Atala 5). It yields continuous beads of biomaterials instead of liquid droplets. The CAD-CAM software directs small beads of biomaterials for deposition into 2D, each deposited layer serving as the foundation of the next layer. It has been used to fabricate multiple types of tissues, such as aortic valves, branched vascular trees, and in vitro pharmacokinetics. Laser-Assisted Bioprinting follows the principle of laser-induced forward transfer (Murphy and Atala 5). It is increasingly used in tissue and organ engineering, and is particularly successful in peptides, DNA and cells. The focused laser pulses hit the absorbing layer of the ribbon to generate a high-pressure bubble that propels cell-containing materials toward the collector substrate. It is nozzle-free; thus free from the common clogging problem. It can deposit cells at a density of up to 108 cells per ml with resolution of a single cell per drop.
Applications
3D Bioprinting has primary applications in medicine, particularly tissue engineering (Henmi et al. 36; Murphy and Atala 1) wherein so much research can be done on any biomaterial. Current applications include the generation and transplantation of several tissues, including multi-layered skin, soft tissues, bone, vascular grafts, stents, tracheal splints (for localized tracheobronchomalacia), heart tissue, and cartilage structures (Murphy and Atala 1).
Potential applications of 3D Bioprinting include the development of high-throughput of 3D-bioprinted tissue models for research, drug discovery, and toxicology (Murphy and Atala 1).
Biomimicry approach had been applied in many technological problems such as flight, materials research, cell-culture methods, and nanotechnology (Murphy and Atala 1). Its application to 3D Bioprinting involves identical reproductions of the cellular and extracellular components of a tissue or organ. In addition, mini-tissues building blocks had been used to form branched vascular networks and, using 3D Bioprinting, accurately reproduce functional tissue units to create ‘organs-on-a-chip,’ which is maintained and connected by a microfluidic network (Murphy and Atala 1). It has been used in the screening of drugs and vaccines or as in in vitro models of disease. Furthermore, 3D Bioprinting has current and potential applications in various fields of study such as paleontology (reconstruction of fossils), archaeology (replicating ancient and priceless artifacts), and forensic pathology (reconstruction of bones; and body and reconstruction of heavily damaged evidence acquired from crime scene investigation).
Conclusion
For only a few decades, the additive manufacturing technology, or 3D printing, has gone a long way in its usefulness. From its exclusive industrial applications, the subtractive 2D technologies helped explode the fabrication technologies into its additive counterpart that provide of much employ in the medical and other sciences that involves the examination and intervention of biomaterials either for research or treatment purposes. The advent of 3D Bioprinting has spawned technological developments in adapting current printing technologies that use inks, non-biological liquids and lasers into printing machines that hand human biomaterials. As a result, a growing understanding of biomaterial properties is driving bioprinting technologies into far-ranging applications, which are expected to grow wider and more specialized in the future. That makes the future of 3D Bioprinting highly exciting and full of anticipations.
Works Cited
“What is 3D Printing?” 3dprinting.com. Web
Henmi, C. et al. “New Approaches for Tissue Engineering: Three Dimensional Cell Patterning
Using Inkjet Technology.” Inflammation and Regeneration January 2008 28(1), 36-40. PDF file
Murphy, S.V. and A. Atala. “3D Bioprinting of Tissues and Organs.” Nature Biotechnology
2014, 1-13. PDF file
