Abstract
Abstract
The International Conference on Bioprinting and Biofabrication in Bordeaux (3B'09) demonstrated that the field of bioprinting and biofabrication continues to evolve. The increasing number and broadening geography of participants, the emergence of new exciting bioprinting technologies, and the attraction of young investigators indicates the strong growth potential of this emerging field. Bioprinting can be defined as the use of computer-aided transfer processes for patterning and assembling living and non-living materials with a prescribed 2D or 3D organization in order to produce bio-engineered structures serving in regenerative medicine, pharmacokinetic and basic cell biology studies. The use of bioprinting technology for biofabrication of in vitro assay has been shown to be a realistic short-term application. At the same time, the principal feasibility of bioprinting vascularized human organs as well as in vivo bioprinting has been demonstrated. The bioprinting of complex 3D human tissues and constructs in vitro and especially in vivo are exciting, but long-term, applications.
It was decided that the 5th International Conference on Bioprinting and Biofabrication would be held in Philadelphia, USA in October 2010. The specially appointed 'Eploratory Committee' will consider the possibility of turning the growing bioprinting community into a more organized entity by creating a new bioprinting and biofabrication society. The new journal Biofabrication was also presented at 3B'09. This is an important milestone per se which provides additional objective evidence that the bioprinting and biofabrication field is consolidating and maturing. Thus, it is safe to state that bioprinting technology is coming of age.
1. Back to Europe
The International Conference on Bioprinting and Biofabrication in Bordeaux (3B'09), France, 6–8 July 2009, was held after several international meetings which had been organized previously. The First International Workshop on Bioprinting and Biopatterning [1] was held at the University of Manchester (UK) in September 2004 and was organized by Professor Brian Derby (University of Manchester), Douglas B Chrisey (Naval Research Laboratory, Washington, USA), Richard K Everett (ONR Global, London) and Nuno Reis (Universidade de Beira Interior, Covilha, Portugal). The Second International Workshop on Bioprinting, Biopatterning and Bioassembly was chaired by Vladimir Mironov in 2005 in Charleston (USA) [2]. The Third International Symposium on Bioprinting and Biofabrication was held in Kawasaki (Japan) in November 2006 and was organized by Professor Makoto Nakamura (University of Toyama, Japan).
After three years without an international meeting, Fabien Guillemot (INSERM, University of Bordeaux, France) and Professor Makoto Nakamura decided to organize the International Conference on Bioprinting and Biofabrication (3B'09) [3] in Bordeaux, the wine capital of the world. This 4th international meeting on bioprinting was endorsed by TERMIS (Tissue Engineering and Regenerative Medicine International Society) and sponsored by the clusters Advanced Materials in Aquitaine, Materials in Bordeaux and Route des Lasers, INSERM and the Aquitaine Regional Council. It took place at the Burdigala Hotel in a very pleasant and friendly atmosphere where new players, especially the young, were welcome and were given the opportunity to discuss their work. The balanced geographical representation was another important feature of the 3B'09 conference. Indeed, more than 65 scientists and engineers from 11 countries (Belgium, France, Germany, Italy, Japan, Poland, Portugal, Romania, The Netherlands, UK, USA) attended this conference, which included five oral presentation sessions, one poster session and social events organized at the Bordeaux City Hall and at the Château Giscours.
2. Scientific program
The scientific program was established to highlight the latest developments associated with bioprinting technologies and biofabrication approaches. Indeed, since the early pioneering works of Thomas Boland (Clemson University, USA) and Vladimir Mironov [4], the bioprinting scientific community has been evolving, bringing together physicists, biologists and physicians [5, 6]. Consequently, the oral presentation sessions covered: (1) the latest developments in bioprinting technologies; (2) the potential to combine the bioprinting process with other biofabrication and rapid prototyping methods; (3) matrices and biomaterials for bioprinting and biofabrication; (4) methods for designing, modeling and biomechanically evaluating 3D constructs; (5) developmental biology and tissue engineering.
This scientific schedule emphasized some of the main areas that have to be connected prior to envisaging real applications in regenerative medicine, pharmacokinetic and toxicological studies, where high throughput and high-resolution bioprinting technologies are required. Thus, it is now obvious that, in addition to core bioprinting technologies, biomaterial and bioink properties, rapid prototyping approaches and basic cell biology have to be taken into account within a single perspective. Consequently, while the initial definition of our burgeoning field was formulated at the First International Workshop on Bioprinting and Biopatterning in Manchester (UK) as 'the use of material transfer processes for patterning and assembling biologically relevant materials, molecules, cells, tissues, and biodegradable biomaterials with a prescribed organization to accomplish one or more biological functions', we propose to enlarge this definition to 'the use of computer-aided transfer processes for patterning and assembling living and non-living materials with a prescribed 2D or 3D organization in order to produce bio-engineered structures serving in regenerative medicine, pharmacokinetic and basic cell biology studies'.
Regarding the poster session, three young scientists were awarded a bottle of Château Giscours wine for their impressive work. Laureates were Virginie Kériquel in vivo high-throughput biological laser printing of nano-hydroxyapatite in mice calvaria critical sized defects: preliminary results) [7], Kayo Sakaue (Integration of 3D-micro tissue models for creation of tissue chip) [8] and Alberto Rainer (Regeneration of osteochondral segment via mesenchymal stem cells culturing on 3D rapid prototyped scaffolds) [9].
3. On bioprinting technologies
The 3B'09 conference was an excellent opportunity to perform an in-depth review of bioprinting technologies (jet-based and extrusion methods) through a number of exciting talks given by both academics and industrialists.
Recent advances in ink-jet bioprinting technologies were reviewed by Professor Thomas Boland and supplemented mainly by the presentations given by Professor Makoto Nakamura [10, 11] and Professor Brian Derby. Regarding laser-based technologies, a series of talks dealing with experimental and modeling approaches of laser-assisted bioprinting (LAB) [12] were given by members of Fabien Guillemot [13] and Boris Chichkov's groups [14] from The University of Bordeaux and Lazer Zentrum in Hannover (Germany), respectively. Utkan Demirci (Harvard University, USA) introduced a new ultrasonic wave-based bioprinting technology, the so-called layer-by-layer 3D tissue epitaxy by cell-laden hydrogel droplets [15] while Hedges and Wirth from Germany presented an aerosol jet bioprinting technology and its applications in surface biofunctionalization [16].
Besides the above-mentioned jet-based bioprinting techniques, many presentations were related to 3D plotting by extrusion processes. Giovani Vozzi's group (University of Pisa, Italy) presented several talks dealing with the micro-fabrication of two- and three-dimensional structures by pressure-assisted micro-syringe (PAM) [17]. Moreover, processing considerations for the 3D plotting of thermoplastic scaffolds were presented by Kim Ragaert (Ghent University, Belgium) [18] and Hendrik John (Sys-Eng, Germany) while Kentaro Iwami (Tokyo University, Japan) described for the first time rapid prototyping by extruding/aspirating/refilling thermoreversible hydrogel [19].
Finally, additional 3D biofabrication methods were presented. Matsusaki et al (Japan) introduced possible, layer-by-layer, short-term applications of bioprinting for biofabrication of tissue chips. Frasca et al (University of Paris-Diderot, France) demonstrated how nanotechnology (magnetic nanoparticles) can be employed in magnetic force-driven tissue engineering.
In summary, while each of these technologies displays specific properties such as high resolution, high throughput, low price, bio-safety, etc, it seems obvious that achieving more advanced applications in tissue engineering will require a combination of these tools, and thus a combination of their performances. Moreover, to print human organs, highly integrative approaches should be set up; these should include the development of new biomaterials, the improvement of reverse-engineering and rapid prototyping methods, intensification of scientific gateways between developmental biology and tissue engineering [20] (as introduced by Gabor Forgacs, University of Missouri, USA) [21], in particular with the help of numerical modeling. All these perspectives were discussed during the conference and are reported in the following sections.
4. Scaffold or not scaffold, that is the question!
One of the controversial topics of discussion during the conference was the definition of 'scaffold'. Synthetic biodegradable scaffold is considered to be a fundamental principle of the traditional scaffold-based or 'top down' tissue engineering approach [22]. The emerging modular or 'bottom-up' approach is sometimes called 'scaffold-free' [23] or 'scaffold-less'. Some participants insisted that scaffold according to definition is a temporal and removable support. Thus, using the term 'scaffold-less' when in reality a 'temporal supporting template' (for example, agarose hydrogel) has been used, is debatable. If in bioprinting technology, biodegradable, removable or sacrificial hydrogel is used, then the term 'scaffold-less' becomes even more inappropriate. Consequently, whether bioprinting is in essence a bottom-up approach, and thus differs from traditional scaffold-based tissue engineering, it does not imply necessarily that it is scaffold-less. Hence, 'solid scaffold-free' is probably a more justifiable term than 'scaffold-less'.
It does not however exclude the possibility of developing scaffold-less bioprinting based on using only cellular material without any temporal or supporting scaffold or hydrogels (biopapers). It has already been proven that an engineering vascular graft using synthetic scaffold-free cell sheet technology is feasible [24]. But even in this case, a so-called 'internal membrane' has been used which is nothing more that a decellularized extracellular matrix (or scaffold) synthesized by living cells in vitro.
It is debatable whether fast-evolving self-assembly technology and the generation of self-assembling natural-like or biomimetic extracellular matrices using methods of macromolecular chemistry and nanotechnology will make the difference between scaffold-less and scaffold based bioprinting technology even more fuzzy. Thus, it remains to be seen if 'solid scaffold-free' or 'scaffold-less' bioprinting technology is just semantic or a real innovation.
In addition to mechanical support and biological uses, it was shown during the conference that scaffolds could bring additional signals. Vozzi's group (University of Pisa, Italy) demonstrated such an approach in designing novel electroconductive scaffolds.
5. Living and non-living materials
In addition to the wider application of natural biomaterials, designing, synthesis and testing of novel biomaterials for bioprinting is critically important. Indeed, there is a need to develop both more effective and more biocompatible biofabrication processes. In this area, material features are some of the most important issues, since a material suited to a particular fabrication method can, on the one hand, enhance the fabrication ability and, on the other hand, influence cell fate. In this latter aim, biofunctionalization by bioactive molecules and humoral factors can help in controlling adhesion, migration, proliferation, and differentiation of living cells [25] as well as in promoting tissue development (including the formation of characteristic organ microstructures and induction of capillary vessels). This orientation was in particular discussed by Nakamura during the conference [26]. Design specification is then a first and critical step on the way to developing and testing tailored bioprocessible and biomimetic biomaterials for bioprinting.
Thus, broader participation of biomaterial scientists and chemical engineers in bioprinting and biofabrication research and conferences is highly desirable. Towards this aim, an interesting way for screening biomaterial and cell-material interactions, the so-called 'materiomics', was proposed by van Blitterswijk (Twente University, NL) [20]. Such study for exploring safe or biofunctional materials must contribute to the future development of biofabrication and tissue processing technologies.
In several talks given by Nakayama (Fukuoka University, Japan), van Blitterswijk, Mironov and Forgacs (University Missouri, USA), it was proposed that living tissues be used as building blocks for printing 3D functional living tissue constructs. This emerging modular [27] or bottom-up approach was discussed at the conference and was also recently presented in an invited Leading Opinion Paper in the journal Biomaterials. It is interesting that the Editor-in-Chief, Professor D F Williams, in the paper 'On the nature of biomaterials' [28] wrote: 'Mironov et al consider organ printing to be the layer-by-layer additive robotic fabrication of 3D functional living tissues and organ constructs [29]. Mironov argues ... that tissue engineering may be better served by a solid biodegradable scaffold-free process. This position is predicated on the assumption that tissues and organs are self-organizing systems and that they normally undergo biological self-assembly and organization without any external influence in the form of instructive, supporting and directing templates or solid scaffolds. There is obviously some way to go before such a paradigm could be translated into a practical reality, but many steps have been taken. It might seem at first sight that if tissue engineering can be achieved by self-assembly of tissue components without the need for conventional solid materials, biomaterials would have no further role. In line with many earlier statements in this essay, I would consider the self-assembled tissue to be an engineered construct and, therefore, a biomaterial in its own right'. Thus, the leading biomaterial expert suggested considering living tissue blocks as biomaterials. Whether this is a real paradigm shift in biomaterial science and tissue engineering associated with emerging bioprinting technology, or just a question of semantics, remains to be seen. Several conference participants strongly believe that it is a paradigm shift. However, the real issue, as Williams outlined, is a translation of this new paradigm into practical reality.
6. The value of mathematical modeling and computer simulation
Realization of rapid prototyping technology is impossible without computer science and the corresponding software. Acquisition of clinical images using modern imaging modalities and transforming them into computer-aided design (CAD) and STL files are important and essential steps in the bioprinting process as was well illustrated in Sun's (Drexel University, USA) state-of-the-art review [30]. However, existing commercial software is not flexible enough for biological applications especially for designing blueprints for soft 3D organs. For example, as was demonstrated in the talk about organ printing, bioprinted tissues undergo processes of tissue fusion and associated contraction and remodeling of printed constructs. The power of mathematical modeling in predicting post-printing tissue remodeling has also been illustrated in talks given by Forgacs' group (University of Missouri, USA). Mathematical modeling could be also used for prediction kinetics of cell seeding, tissue ingrowth, oxygenation and vascularization of bioprinted tissue constructs. New, more sophisticated and more flexible software, capable of incorporating experimentally estimated coefficients of tissue retraction, compaction and remodeling must be developed. This work is critical for advancing bioprinting technology and organizers of future bioprinting conferences must make a special effort to attract more specialists in mathematical modeling, computer simulation and computer science.
7. How to print a human organ
Most conference participants were very cautious about the potential of bioprinting technology to biofabricate functional human organs. The consensus was that it is a highly desirable, but long-term, goal. It was also agreed that without the engineering of an intraorgan branched vascular tree relatively thick 3D bioprinted tissue constructs will not survive. However, in at least one talk a comprehensive attempt was made to demonstrate how a complex 3D human organ such as a kidney could be bioprinted. The group from the Medical University of South Carolina (Charleston, USA) and Clemson University (Clemson, USA) presented a comprehensive step-by-step approach to bioprinting a functional kidney organ construct. It obviously still has a long way to go and a lot must be done for practical realization of this ambitious vision, although many essential elements of future organ printing technology are already in place. For example, the South Carolina group presentation as well as Forgacs' group presentations demonstrated that building a branching intraorgan vascular tree is a realistic and achievable goal. This issue was also addressed by Peter Wu (University of Oregon, USA) who presented applications of LAB in fabricating branch/stem structures with human endothelial cells [31] and T Boland who presented results on thermal inkjet printing of biomaterials and cells for capillary constructs [32].
8. A new prospect: bioprinting in vivo
One of the main novelties of this conference was the demonstration that in vivo bioprinting is feasible. Although many groups discussed the possibility of this idea, the view was that at best it is just science fiction. However, for certain applications (especially when there is no need for immediate vascularization) bioprinting in vivo can be a realistic option. The first attempt at bioprinting in vivo was presented by the French group [7].
One of the authors of this paper discussed the data presented at the conference with the CEO of one of the leading rapid prototyping companies and during this discussion the idea of a robotic clinical bioprinter was coined as a potential 'Michelangelo' project. Whether the clinical bioprinting system 'Michelangelo' will one day join the popular sophisticated 'Da Vinci' robotic system remains to be seen. As Chair of The Department of Surgery of Stanford University and a leading expert in surgical innovation Krummel recently wrote: 'There is no such things as a science fiction. There is only science eventuality'. If this is true, then one day we can compare printing a human ear in a laboratory using the 'Michelangelo' clinical bioprinting system with a similar procedure demonstrated in the science-fiction movie Face/off. How this in vivo bioprinting system will be designed and how it will work nobody yet knows, but it is safe to predict at least two things: that bioprinting tools will be used and that the future 'Michelangelo' system will have much more sophisticated capabilities than just the imitation and enhancement of a surgeon's hand movements as the modern 'Da Vinci' robotic system does.
9. What is next?
As previously mentioned, since the First International Workshop on Bioprinting and Biopatterning held at the University of Manchester (UK) in 2004, the bioprinting scientific community has been growing, now bringing together physicists, biologists and physicians. Consequently, and due to the practical impossibility of accommodating the growing demands for special bioprinting sessions at traditional tissue engineering conferences as well as the very interdisciplinary character of this emerging field, it was proposed to structure the community. After careful discussion and considering all the pros and cons, a dominant majority of 3B'09 attendees enthusiastically supported the idea to form the 'Exploratory Committee on Bioprinting' (made up of organizers of previous conferences) and to also create the 'Bioprinting, Biofabrication and Bioassembly Society' (3BS), which would endorse the new journal Biofabrication.
It was also decided that The 5th International Conference on Bioprinting, Biofabrication and Bioassembly (3B'10) will be organized by Professor Wei Sun (Drexel University, USA) and Professor Gabor Forgacs (University of Missouri, USA) in Philadelphia, USA; 3B'11 will be organized by Professor Makoto Nakamura in Toyama, Japan and 3B'12 will be again in Europe (potentially in Portugal or Italy, although the location is still open for proposals).
Finally, it seems that the bioprinting field is entering a phase of potentially explosive exponential growth and organizational consolidation or, simply speaking, bioprinting is indeed coming of age. It is essential that at future bioprinting and biofabrication conferences the established tradition of the emerging bioprinting community to provide a platform for newcomers for presenting novel innovative technologies and approaches, as well as open and constructive professional discussion of existing challenges and problems, is maintained. This will ensure that a vibrant, innovative and professional research community evolves.
Acknowledgements
Fabien Guillemot (FB) thanks the clusters Advanced Materials in Aquitaine, Materials in Bordeaux and Route des Lasers, INSERM and the Aquitaine Regional Council for supporting financially organization of the 3B'09 conference. Moreover, FG acknowledges tremendous work done by the members of the local organizing committee (Bertrand Guillotin, Sylvain Catros, Reine Bareille, Benjamin Pippenger and Jean-Christophe Fricain).
Makoto Nakamura (MN) thanks the Bordeaux local committee members for the success of the 3B'09 conference. He would also like to give his best regards to all of the participants who gathered from all over the world for this conference, hoping to ensure the succession of academic international activity for bioprinting and biofabrication research.
Vladimir Mironov acknowledges support from an NSF EPSCOR R-II grant.
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