Imagine the day when organs are produced in the lab and are available for transplant, a scenario often repeated by doctors, patients, and medical reporters alike. What seems like a scene from a science-fiction movie may one day become reality—and sooner than we may imagine. Our understanding of biological systems and cellular mechanisms is rapidly expanding, accompanied and supported by advances in bioengineering. 3D printing, now a household word, has been hailed among the most exciting inventions of this decade with hardly a week passing by without new “firsts” using 3D printing technology. This novel technology has vast potential for multiscale innovations in almost every discipline: health-care, industry, academia, and the arts. In health-care, 3D printing is often called a game changer. It has already customized prosthetic and implant design and impacted the pharmaceutical industry and drug delivery systems, medical education, and most of all—tissue engineering and regenerative medicine. Here we briefly describe the basic concepts of this technology for the busy clinician and how it can be applied to tissue engineering with a special focus on airway regeneration.
3D printing process
3D printing, also known as rapid prototyping or additive manufacturing, was introduced in the 80s for industrial purposes. Not until a miniaturized “desktop” version of the printer was developed did its role in medicine begin to expand. All 3D printers, regardless of their types, follow similar principles. Products of 3D printers are objects made by sequential addition (z-stacking) of 2D layers creating three-dimensional structures. The objects of interest can be designed using computer-aided design (CAD) software or taken directly from 3D-reconstructed images of CT scans and MRIs. The image files are saved as a (.STL) file and processed by “slicer” software to generate G-code files that relay the control instructions to the printer. Depending on the size of the printer, the “ink” used, and the size and geometrical complexity of the desired object, it can take from a few minutes to several hours or days to print. Unlike the monochromatic older generations of 3D printers, newer devices are emerging on the market with either multiple printing heads or “ink” chambers. The latter provides greater freedom in materials choice and is of particular interest in tissue regeneration for its ability to print and compartmentalize different cellular components.
Types of 3D printers
There are multiple types of 3D printers available commercially, benefiting from an open-source platform that allows customization and improvement of the current devices. Similarly, different types of “inks” are utilized depending on the printer’s design but also on the desired end-product. We mention some of the common types of printers and direct our focus to the last two, given their particular application in regenerative medicine:
• Selective laser sintering
• Electron beam melting
• Direct laser metal sintering
• Selective heat sintering
• Electron beam freeform fabrication
• Fused deposition modeling
• Photopolymerized extrusion stereolithography
Fused deposition modeling printers
Fused deposition modeling (FDM) printers utilize a heated extrusion head and are closely similar to a desktop inkjet printer. FDM printers benefit from their wide commercial availability and relatively low cost, making them among the most popular 3D printers currently in use. The “ink” utilized is a thermoplastic material typically prepared as a thread spool fed to the heated head and sequentially deposited as droplets with an approximate 100-micron resolution. The thermoplastics harden within few seconds, allowing fast and precise 3D object production. Several thermoplastics are used, some of which are also biocompatible and have been traditionally used in implant manufacturing and have shown promise for tissue engineering, such as polylactic acid (PLA), polylactic-glycolic acid (PLGA), and polycaprolactone (PCL). PCL has attracted substantial interests within the medical community given its low inflammatory profile, slow rate of hydrolysis, and ability to promote cellular attachment and growth (Shimao. Curr Opin Biotechnol. 2001;12[3]:242). In fact, PCL has been successfully printed as a splint for bronchial malacia in a baby suffering from repeated bouts of pneumonia and difficulty breathing (Zopf et al. N Engl J Med. 2013;368[21]:2043). FDM printers are thus most suitable for manufacturing prosthetic devices and possibly tissue scaffolds for cellular growth.
Photopolymerized extrusion stereolithography printers
In this form of 3D printing, the extrusion head consists of a motorized syringe-plunger containing the liquid ink. This is polymerized into a solid shape after extrusion and upon exposure to UV light (or another light source depending on the chemical content of the mixture). Polymerizable liquid inks are grouped under the term hydrogels, formerly defined by the International Union of Pure and Applied Chemistry (IUPAC) as “nonfluid colloidal network or polymer network that is expanded throughout its whole volume by a fluid.” Several formulations of hydrogels exist and we have used varying combinations of polyethylene glycol diacrylate and alginate to create hydrogels with different mechanical properties. Photopolymerization is slower than FDM; however, it has the added benefit of incorporating the cells into the prepolymerized liquid mix, which allows cellular inclusion into the final product. In essence, this type of printing will likely represent the future of “bioprinting” with its ability to compartmentalize cellular components within the scaffold. 3D printed ears (Manoor et al. Nano Lett. 2013;13[6]:2634) and aortic valves (Hockaday et al. Biofabrication. 2012;4[3]:035005) were generated using this approach. 3D printing was successful in replicating the shape of the desired organs with high fidelity while permitting maintained cellular growth, ushering a new era for regenerative medicine. However, the current mechanical properties of these structures do not permit organ transplantation and lack the necessary vascularized network for maintained in vivo growth.