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Technology and medicine are ever-advancing. On the one hand, 3D-printing is revolutionizing our creation process. On the other hand, cryopreservation through cryogenic temperatures could possibly allow for incredible developments in the medical field. But the real magic happens when you put technology and medicine together to create something entirely new. Let me introduce to you: Cryogenic 3D Printing. 

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Research

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Cryogenic 3D printing aims to create frozen hydrogels using cryopreservation to stabilize the printed object in an ideal state. Then, when they’re needed, they can be rewarmed. This procedure was first published by the Department of Mechanical Engineering at the University of California in 2015. It’s since been used as a foundation for other publications and further research studies.

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The final goal is the successful creation of fully functional 3D-printed human organs and human tissue for transplantation. However, to understand how cryogenic 3D printing works, we first have to make ourselves familiar with the individual parts it’s made up of.

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3D Printing

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Surely you know what “printing” means. In 1455 the earliest known printer was made by Johann Gutenberg. He kickstarted the education renaissance by mass-producing books for the first time. Prior to this invention, every book had to be written by hand. If you wanted a second copy, you had to write the same book again - Inconceivable by today's standards. 

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3D printing aims to extend this revolution beyond the 2-dimensional plane by making it possible to print any 3D object you can think up. This has its limitations though, which keep 3D printers from being used in mass-production. Instead, they are currently utilized to print specific, hard to manufacture objects like design-prototypes. They can also print final products such as jewelry, tools, toys, novelty items or even prosthetics. 

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3D printing is an additive process. This means that, unlike wood-carving or most metal-work, the desired object is built from the ground up and not carved out of an existing object. Computer programs are used to tell the printer what to print. With these, designers can create digital 3D models, or CAD models, which then get sent to the 3D printer to handle production. Autodesk Fusion 360, Autodesk AutoCAD, Ultimaker Cura, TinkerCAD and SketchUp are amongst the most-used programs at the moment. There are different types of 3D-printers out there, using different methods (and materials) to create the 3D object. If you want a compact overview of what’s on the market, take a look at this video. 

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Bioprinting

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Bioprinting is a subcategory of 3D printing that uses biological materials like cells or biomaterials to fabricate biomedical parts. Unlike the usual material used for 3D printing, plastic or resin, biological components require extra steps to retain their shape (and function):

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• Pre-Bioprinting - Unlike normal 3D-printing, building “whatever we want” isn’t the goal of bioprinting. Instead, scientists are trying to replicate human biology as faithfully as possible. To enable that vision, they have to obtain a diagnostic test, also called biopsy, first. This is done via computed tomography (CT) and magnetic resonance imaging (MRI). Because 3D-printers are limited to layer-by-layer printing, a tomographic reconstruction of the resulting images has to be made as well. When the images are ready to be sent to the printer, specific cells are isolated, multiplied and later mixed with a special liquefied material that provides vital nutrients (f.e. oxygen) to these cells.

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• Bioprinting - In the next step, the biological cocktail that was mixed together gets stored in a printer cartridge and deposited together with the patient's medical scans. This bioprinted pre-tissue gets put into an incubator and slowly matures into a regular tissue. 
  
  Bioprinting usually involves the use of biocompatible scaffolds, designed to hold the individual layers in place. These are of critical importance for tissue engineering as they provide a suitable mechanical and chemical environment that enables further growth into fully functional tissue.
  
  While the production of tissues and smaller organs has proven successful, bigger organs like livers and lungs made by 3D bioprinting lack some crucial elements. This is primarily due to the limitations of current 3D-printers. Some intricate vital functions like working blood vessels don’t possess the stability or viability necessary to be used in applied medicine at this point in time.

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• Post-Bioprinting - To ensure a stable, functional structure for the future, a well-maintained post-bioprinting process is necessary. Scientists apply both mechanical and chemical stimulation to the bioprinted objects. These send out signals to the cells, aiming to control the growth process of the tissues. The type and amount of signals sent depends on the individual tissue. These so-called Bioreactors can range from transporting additional nutrients, creating specialized environments, adjusting air pressure and many more.

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Cryogenic 3D Printing

Let’s take everything we’ve learned so far and take a look at what works, and what doesn’t.

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What Works:

• Print 3D objects out of regular construction materials
• Print 3D objects out of biological materials
• Print basic human tissue
• Print fully functional small organs

What Doesnt:

• Print fully functional large organs
• Guarantee stability while printing intricate structures
• Guarantee functionality while printing intricate structures

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Looking at this checklist, we’ve already come quite far. Still, the last steps turn out to be a great hurdle to overcome. Current 3D bioprinting technologies lack the functionality to ensure stability of complex bioprinted formations. 

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But wait, isn’t there a scientific field out there that’s all about stability and preservation (or rather cryopreservation) of structures? Boris Rubinsky, researcher and professor at UC Berkeley, and Michal Adamkiewicz thought the exact same thing, and started experimenting. Their result: The so-called Cryogenic 3D Printing.

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In Cryogenic 3D Printing the bioprinting process is conducted with the 3D-printed object constantly immersed in a liquid coolant (liquid nitrogen). This coolant is adjusted so it’s always level with the highest layer of the print, allowing for a very precise freezing process. The temperatures during the procedure generally range from -20°C to -25°C. CO2 gas gets injected into the chamber rapidly, enabling it to achieve high cooling rates at the start of the process. Vapor Compression Refrigeration (VCR) then ensures sustained heat removal for the rest of the procedure.

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The end-results are fully stabilized (through freezing) bioprinted structures that exhibit a significantly higher likelihood of maintaining a workable shape. This approach has proven especially effective when it comes to creating scaffolds, demonstrating the potential of cryopreservation.

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Conclusion

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Science still needs more time to solve all the problems associated with recreating human biological functions. Nevertheless, we are making steady progress in many different fields of research. Scientists have already managed to transplant a fully functional bladder (many years ago) that’s still working today. One day, we might be able to print ourselves new lungs, a new pancreas or even a new heart when needed. Research in cryogenics and human cryopreservation might potentially help us develop the technology and reach this goal. Vise versa, advancements in the field of bioprinting might allow for treatment of Biostasis patient’s organs before they get revived in the future.

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The two fields of science hope to complement each other with these medical advancements. If you want to know more about the Biostasis side of things, feel free to schedule a call with us.

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Source: M. Adamkiewicz, B. Rubinsky, Cryogenic 3D printing for tissue engineering, Cryobiology (2015), http://dx.doi.org/10.1016/j.cryobiol.2015.10.152

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