Since the development of in-depth stem cell research and particularly the ability to induce pluripotency – the ability to differentiate cells into many or all cell-types – the promise of generating replacement tissues and organs for patients has been a virtual “holy grail” for the field of regenerative medicine. Many advances have been made in transforming somatic body cells into pluripotent stem cells, and pathways have been uncovered to differentiate those cells into almost any known cell type in a controlled manner. However, these cells need to be organized into a defined structure for the engineered tissue to be of any use for transplantation. Two conflicting camps have advocated for distinctly different methods, one using a framework of biodegradable polymer treated with drugs to encourage cell growth within the matrix, and the other using direct cell “printing,” which deposits individual cells to create complex structures. Both of these methods have their merits, and the argument essentially boils down to building top-down versus bottom-up – there is, however, a lot of common ground between the two.
As early as the late 1990’s, the most popular idea for organizing cells into an organ was the top-down scaffolding concept. This approach involves developing some biocompatible structure on which seeded cells could grow and differentiate in a controlled way. This idea was in development in multiple forms before the big stem cell boom — when it became possible to reprogram somatic cells to stem cells instead of finding a donor — when it became possible to reprogram somatic cells to stem cells instead of finding a donor – and subsequently became massively popular as the potential to combine the potency of stem cells with the organization of a scaffold electrified the regenerative medicine field. Numerous methods of producing a scaffold have since been developed. The most organic, though perhaps least clinically relevant, is a method that involves de-cellularizing a cadaveric organ using detergents delivered via the organ’s vasculature.4 After this, the remaining connective tissue structure is re-populated with cells. This method has demonstrated a degree of potential: one group reported success using neonatal rat cardiac cells to repopulate an adult heart to create a functional organ, and another group used a similar procedure on an adult mouse kidney and repopulated it with murine embryonic stem cells, with similar success.4 While these experiments present the promise of scaffolds in tissue engineering, they also come with severe drawbacks, namely the need for an existing cadaveric organ (which in the clinical case would have to come from a different individual than the patient, introducing complications to the transplantation procedure).
To address these issues, scientists have been pursuing multiple avenues for the generation of artificial, biodegradable polymer scaffolds, using techniques ranging from photolithography to syringe-based gel deposition to freeform solid fabrication. However, most of these methods lack important features, such as internal architecture resolution, which allows for more complex internal structures to develop (such as, very importantly, vasculature), flexibility or simply the capacity for “high-throughput” production, the ability to produce tissue quickly and efficiently.1 It has proven difficult to find a balance between the ability to carefully define intricate internal structures, defined by multiple cell types, and the fabrication time necessary to produce a prototype which can then be seeded with organs. Furthermore, many cell-types (typically mesoderm-derived types, like muscle cells) often require a biomechanical force to be applied to promote cell growth and organization, which is something that current polymer scaffolds cannot address.8 As a result of these drawbacks, scaffolding methods have had the most success in tissue generation that involves relatively simple layouts, such as the engineered cartilaginous tissue developed by Feng-Huei Lin’s group for the potential treatment of osteoporosis.5
An alternative camp developed sometime around 2003, when Roger Markwald and Thomas Boland modified a commercial inkjet printer to print protein solutions and individual cells on glass and gel surfaces.2,6 The software they developed to accompany their custom cell-printer allowed them to deposit cells on a gel in pre-defined, high resolution patterns with very high cell viability. Further modifications to the system by subsequent researchers allowed for printing inside of a three-dimensional block of gel to create intricate cell structures, internal and external, of varying cell-types, with very high viability and cell-cell fusion to create solid surfaces of tissues. Using this method, very intricate organ structures can be designed using the computer software and then printed using “bioinks” made from partially or fully differentiated cells and proteins that promote cell adhesion.2 The drawback, however, is the time necessary to produce the organ itself, since the process involves building the organ cell by cell, layer by layer, which is a lengthy process for something as complex and large as an adult heart, for instance. Other groups have been experimenting with alternate printing methods to improve printing speed and resolution further, such as Fabien Guillemot’s group, which used laser assisted printing to print slightly faster with increased cell density and resolution depending on the laser’s scanning frequency (how rapidly it passes over the surface to be printed).3 The scaling and throughput issue intrinsic to this approach to organ building is a continuous source of new work in the field, with new innovations coming in by the droves.
Finally, Ulrich Schubert’s group has arrived at an unconventional middle-ground between these two methods: organ weaving. Their method involves using suturing threads that are coated in an alginate gel with suspended cells, and then taking multiple threads, coated with either the same or different cell types, and literally weaving them around a base structure.7 They experiment using gradients of cell types based on the pattern used in the weave, and in all cases achieve very high cell viability. Their method combines the high-throughput ease and speed of artificial scaffolding with the ability to better control the resolution and cell types used like with cell-printing. This is a unique and very promising approach to tissue engineering.
As stem cell technology advances further, making patient-specific stem cells easier to make, maintain, and differentiate into target cell types, tissue engineering is evolving to keep in step with the demands of the clinical promise of custom organs. Though some critics are concerned that in vitro organogenesis is the first step on the “slippery slope” of human cloning, the promise for highly beneficial clinical application under careful and insightful oversight is enough to calm most opponents’ fears. As the technology develops, we see less conflict between the top-down and bottom-up approaches, and instead see a harmonious combination of the two approaches, melding the benefits of both to create new avenues for successful organ growth in the future.
- Boland, Thomas, et al. “Advances in Tissue Engineering: Cell Printing.” Journal of Thoracic and Cardiovascular Surgery 129, no. 2, (2005): 470-472.
- Boland, Thomas, et al. “Cell and Organ Printing 1: Protein and Cell Printers.” The Anatomical Record, Part A 272A, (2003): 491-496.
- Guillemot, Fabien, et al. “Laser Assisted Bioprinting of Engineered Tissue with High Cell Density and Microscale Organization.” Biomaterials 31, (2010): 7250-7256.
- Kobayashi, Eiji, et al. “Kidney Organogenesis and Regeneration: A New Era in the Treatment of Chronic Renal Failure?” Clinical Experimental Nephrology 12, (2008): 326-331.
- Lin, Feng-Huei, et al. “A Highly Organized Three-Dimensional Alginate Scaffold for Cartilage Tissue Engineered Prepared by Microfluidic Technology.” Biomaterials 32, (2011): 7118-7126.
- Markwald, Roger, et al. “Organ Printing: Computer-aided Jet-based 3D Tissue Engineering.” Trends in Biotechnology 21, no. 4, (2003): 157-161.
- Schubert, Ulrich, et al. “Organ Weaving: Woven Threads and Sheets As a Step Towards a New Strategy for Artificial Organ Development.” Macromolecular Bioscience 11, (2011): 1491-1498.
- Stoica, Adrian. “Robotic Scaffolds for Tissue Engineering and Organ Growth.” NASA Tech Briefs. Jan 23, 2012. http://www.techbriefs.com/component/content/article/10747.
- HIA. “Herzklappe.” Wikimedia Commons. Dec. 2, 2010. http://en.wikipedia.org/wiki/File:Herzklappe.JPG
Aleks Penev is a third year student at the University of Chicago majoring in Biology with a minor in Computer Science. He is also currently doing research with induced pluripotent stem cells and hopes to extend his research focus into the translational aspects of patient-specific organogenesis and transplantation. Follow The Triple Helix Online on Twitter and join us on Facebook.