Emerging Themes and Technologies in Tissue Engineering and Bioprinting Agenda

Conference Registration, Coffee, and Breakfast Pastries

Chairman's Opening Remarks
Jon Rowley, Chief Executive And Technology Officer, Rooster Bio Inc, United States of America

Coffee Break, Networking with Exhibitors, and Poster Viewing

3D Bioprinting of Vascularized Living Tissue
Jennifer Lewis, Professor, Harvard University School of Engineering and Applied Sciences, United States of America

The ability to pattern biomaterials in planar and three-dimensional forms is of critical importance for several applications, including 3D cell culture, tissue engineering, and organ mimics. 3D printing enables one to rapidly design and fabricate soft materials in arbitrary patterns without the need for expensive tooling, dies, or lithographic masks.  In this talk, the design of novel cell-laden, hydrogel (extracellular matrix) and fugitive (vascular) inks with tailored rheological properties for 3D bioprinting will be described.  We will also present recent advances in 3D bioprinting of vascularized, heterogeneous cell-laden tissue constructs with as well as ongoing efforts to characterize these 3D living architectures.

3D Biofabrication of Complex Living Cardiovascular Tissues: Past, Present, Future
Jonathan Butcher, Associate Professor, Biomedical Engineering, Cornell University, United States of America

Cardiovascular disease remains a leading cause of death worldwide. Despite their potential, achieving functional living tissue replacements has been an elusive “Holy Grail” for the last two decades.  Among the challenges is that natural tissues are highly complex, hierarchical structures that are difficult to replicate. Tissue biofabrication technology, in particular 3D tissue printing, has made significant strides over the past 5 years to close this gap. 3D tissue printing has the capacity to prescribe both macro and microstructural environmental cues that are essential for coordinating whole tissue function. We discuss recent findings on two prominent applications: trileaflet heart valves and vascularized tissue flaps.

3D BioFabrication for Biological Machines and Tissue Engineering
Rashid Bashir, Professor And Head, University Of Illinois, United States of America

The integration of living cells with soft scaffolds can enable the fabrication of biological machines and soft robotics. These cell-based biological machines can be defined as a set of sub-components consisting of living cells and cell-instructive micro-environments that could eventually perform a range of prescribed tasks. The realization of biological machines and their sub-components will require a number of suitable cell sources, biomaterials, and enabling technologies. Here, we review our group’s recent efforts towards this goal and of developing cell based biological machines. We have fabricated locomotive ‘‘bio-bots’’ from hydrogels and cardiomyocytes using a 3D printer. The multi-material bio-bot consisted of a ‘biological bimorph’ cantilever structure as the actuator to power the bio-bot, and a base structure to define the asymmetric shape for locomotion. The cantilever structure was seeded with a sheet of contractile cardiomyocytes. We will also describe the development of a 3D-printed electrically paced skeletal muscle based ‘bio-bot’ devices where skeletal myoblasts embedded in ECM proteins compacted around a hydrogel structure were used to create the power source of the biological walking machine. While the specific applications are yet to be defined, these devices could have potential applications in drug delivery, power generation, and other biomimetic systems.

Networking Lunch and Exhibit, Poster Viewing

Multi-Arm Bioprinting: From Perfusable Tissues to in-situ Bioprinted Tissues
Ibrahim Ozbolat, Hartz Family Associate Professor of Engineering Science and Mechanics, The Huck Institutes of the Life Sciences, Penn State University, United States of America

This talk highlights the recent advances in extrusion-based advanced bioprinting technologies using multi-arm bioprinting. The Multi-Arm BioPrinter (MABP) created at The University of Iowa enables synchronized motion of multiple arms in tandem to bioprint multiple bioink at the same time. The MABP is flexible to be equipped with different nozzle apparatus for various purposes including vascular tissue printing, hybrid tissue scaffold printing, scaffold-free vascularized perfusable tissue printing and in-situ multi-arm bioprinting. In this talk, we will emphasize recent significant capabilities including vascularized perfusable tissue printing for drug testing and in-situ bioprinting of bone tissue for translation of advanced robotics technologies from bench to bedside.

3D Scaffolds with Tailorable Topology and Topography through Biofabrication Technologies
Lorenzo Moroni, Professor, Biofabrication for Regenerative Medicine, Maastricht University and Founder MERLN Institute for Technology-Inspired Regenerative Medicine, Netherlands

A key factor in scaffold-based tissue and organ regeneration relies on enhancing (stem) cell-material interactions to obtain the same original functionality. Different approaches include delivery of biological factors and surface topography modifications. Although both strategies have proved to augment cell activity on biomaterials, they are still characterized by limited control in space and time, which hampers the proper regeneration of complex tissues. Here, we present a few examples where the integration of biofabrication technology platforms allowed the generation of a new library of 3D scaffolds with tailored biological, physical, and chemical cues at the macro, micro, and nano scale. By engineering their topological properties, these porous biomaterials influence the activity of seeded cells, thereby initiating the regeneration of skeletal, vascular, and neural tissues. Future efforts should aim at further improving our understanding of scaffold topological properties to achieve a fine control on cell fate at multiple scales. This will enable the regeneration of complex tissues including vasculature and innervation, which will result in enhanced in vivo integration with surrounding tissues. By doing so, the gap from tissue to organ regeneration will be reduced, bringing regenerative medicine technologies closer to the clinics.

Extrusion Printing of Hydrogels with Embedded Cells
Paul Calvert, Professor of Bioengineering, UMass Dartmouth, United States of America

A 3D printer has been used to make porous open gel “logpiles” with 200 micron bars containing embedded cells including yeast, fibroblasts and algae.  The activity of the cells has been studied as a function of gel composition.  As expected, cells can metabolize and multiply within a 100 microns of the gel surface.  Following the activity of a yeast membrane-bound invertase and comparing this with known diffusion coefficients allows us to quantitatively model metabolism. The gel formulations need to give good strength coupled with easy diffusion of nutrients and products will be discussed.  Diffusion coefficients have also been measured for a number of macromolecules in different types of printed gels in order to determine the possibility of using encapsulated cells in 3D gel scaffolds as small bioreactors to produce proteins.

Coffee Break, Networking, Exhibit and Poster Viewing

3D Bioprinting of Cartilage Tissue with Novel CELLINK
Paul Gatenholm, Professor, Director of 3D Bioprinting Center, Chalmers University of Technology, Sweden; CEO, CELLHEAL AS, Norway, Sweden

The development of high resolution 3D Bioprinters enables positioning of several human cell types with high accuracy and reproducibility and thus reconstruction of complex tissue and organs. Rapid advances in stem cell isolation from patient own tissue such as adipose make it possible to have access to sufficient amount of autologous cells for tissue repair in one step surgery. The bioinks needs however to be developed and commercialized to secure supply of printable and cell friendly scaffolds. We have developed a new generation of water borne biomimetic printable scaffolds with unique printability into 3D shapes and ability to support cartilage growth. This lecture will review our work with this novel CELLINK for 3D Bioprinting of ear, meniscus, trachea and articular cartilage.

Development of Bioinks for 3D-Bioprinting
Jos Malda, Professor of Biofabrication in Translational Regenerative Medicine, University Medical Centre Utrecht, Netherlands

Hydrogels are particularly attractive as “bioinks” for biofabrication as they recapitulate several features of the natural extracellular matrix and allow cell encapsulation in a highly hydrated mechanically supportive 3D environment. Additionally, they allow for efficient and homogeneous cell seeding, can provide biologically-relevant chemical and physical signals and can be formed in various shapes and biomechanical characteristics. Nevertheless, there exists a significant challenge in biofabrication: the optimization of – intrinsically weak – hydrogels to address the physico-chemical demands of the biofabrication process and the right conditions for cell survival on the one hand, and to address the harsh in vivo mechanical environment on the other. We have developed novel hydrogel-based bioink formulations that allow for the construction of intricate 3D structures, whilst providing the cells with a biologically suitable environment.

Bioprinting and Bioassembly of Bioficial Organs
Stuart Williams, Director, Bioficial Organs, University of Louisville, United States of America

The assembly of functional organs from a patient’s own tissue and cells remains a major goal of regenerative medicine.  With the development of computer assisted design of tissues and organs and the integration of additive manufacturing technologies the era of 3D Bioprinting has emerged as a new technology toward the manufacturing of custom organs.  This presentation will describe the next generation of bioprinting systems that include new multi-axis robotic systems.   Advancements in biomaterials have created new “bioinks” a critical component of tissue and organ 3D bioprinting.  The field of 3D Bioprinting is positioned to become a major clinical tool to address numerous diseases as well as aid in the development of new diagnostic tests and in drug development.

Tools for the Development of Bioprinting Technology
Jeffrey Lipton, Founder & Chief Technology Officer, SERAPH ROBOTICS, United States of America

3D bio printing is a field with unique technical needs and requirements in the 3D printing community. Bio printing, being in the early stages of process development, needs flexible platforms to allow rapid innovation. Current 3D printer architectures are ether locked down and inaccessible to researchers, or have a high cost and technical barrier. We present the XDFL and Fab@Home printer frameworks as an alternative to these barriers to innovation.

Networking Reception: Enjoy Premium Beers, California Wines, and Appetizers with Your Colleagues with a Beautiful View of Boston and The Charles River

Close of Day 1 of the Conference

Morning Coffee and Breakfast Pastries

High Performance Additive Manufacturing for Long-Term Implantable Devices
Adam Hacking, Chief Scientific Officer, Oxford Performance Materials (OPM), United States of America

High performance additive manufacturing (HPAM) describes the direct fabrication of fully functional, mission critical devices. HPAM of medical devices requires an understanding of material properties, manufacturing techniques, device design and biological performance. Oxford performance materials has developed a unique laser sintering process with a high performance thermoplastic, poly-ether-ketone-ketone (PEKK) that enables the generation of load-bearing additively manufactured devices. OPM is the first company to have obtained FDA clearance for additively manufactured polymeric devices for long-term implantation. This talk will introduce OPM's technology, describe the development and commercialization process, provide an overview of the biological response to PEKK devices and outline current development.

4D Bioprinting: A New Paradigm for Engineering Complex Tissues
Fabien Guillemot, CEO, Poietis, France

Dealing with tissue complexity and reproducing the functional anisotropy of human tissues remain a puzzling challenge for tissue engineers. Emergence of the biological functions results from dynamic interactions between cells, and with extracellular matrix. The important literature showing that cell fate (migration, polarization, proliferation…) is triggered by biochemical and mechanical signals arising from cell microenvironment suggests that tissue formation obeys to short range orders without reference to a global pattern. In that context, the winning tissue engineering strategy might rely on controlling tissue organization at the cell level. Emerging during the last decade, Bioprinting has been 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”. From a technological point of view, the Laser-Assisted Bioprinting (LAB) technology has been developped as an alternative method to inkjet and bioextrusion methods, thereby overcoming some of their limitations (namely clogging of print heads or capillaries) to pattern living cells and biomaterials with a micron-scale resolution. By harnessing this high printing resolution, we observe that tissue self-organization over time depends on the cell patterns initially printed by LAB, as well as cell types. To engineer complex tissues, we then emphasize the need to consider the spatio-temporal dynamics of tissue self-organization when designing blueprints.

Coffee Break, Networking, Exhibit and Poster Viewing

Bioprinting – The Patent Landscape
Robert Esmond, Director, Sterne, Kessler, Goldstein & Fox P.L.L.C, United States of America

Ways to protect bioprinting innovations and patent filings to date will be discussed.  A patent landscape search has revealed that there have been many patent filings on bioprinted organs and tissues and methods of production over the last 10+ years.  Avoiding patent infringement while in the clinic will be discussed as well as the patent expiration dates.

The Bio-Pick, Place and Perfuse: A New Instrument for Layer-by-Layer Building of Tissues and Organs
Jeffrey Morgan, Professor of Medical Science and Engineering, Brown University, United States of America
Anubhav Tripathi, Professor of Engineering, Co-Director, Center for Biomedical Engineering, Brown University, United States of America

The grand challenge of the field of tissue engineering is the fabrication of large living structures with high cell density akin to native organs (liver, kidney) and to sustain the viability and functionality of these structures in vitro prior to transplantation. Despite twenty years of engineering different types of scaffolds for cell attachment, this problem remains unsolved and even new efforts to form functional organs by seeding cells into decellularized organs have significant limitations. Rather than pursue a monolithic approach where cells are seeded into a scaffold or decellularized tissue, we are pursuing a bottom up, layer-by-layer and scaffold-free strategy to address some of these engineering problems. When cells are seeded into micro-molds of a nonadhesive hydrogel (agarose) they will aggregate and self-assemble a 3D multi-cellular microtissue. The shape of this microtissue is directed by our design of the micro-mold. We’ve self-assembled large honeycomb shaped tissues. When harvested from the micro-molds, these microtissues will fuse with each other to become one tissue. To build with these microtissue parts, we’ve devised a novel device, the Bio-Pick, Place, and Perfuse (Bio-P3) inspired by the pick & place machines used in the high-speed assembly of multi-component electronics. A significant advance over bio-printing, our first generation device, picks up microtissue building parts, transports them to a build area, and places them at the desired location while maintaining perfusion. In this way, piece by piece, a larger tissue construct can be assembled layer-by-layer, while building and perfusing a vascular-like network that maintains the viability needed for microtissue parts to fuse. This talk will review our progress to date and discuss how this approach may be able to address some of the challenges of organ fabrication.

Technology Spotlight:
Novel Technology Platform of Bio 3D-Printing
Koji Kuchiishi, CEO, Cyfuse Biomedical

Our Bio 3D Printer Regenova® assembles cellular aggregates (spheroids) into any desired 3D shape by skewering spheroids on a fine needle array. After removing the needles and maturing in a bioreactor, functional tissues are created.  This platform technology is applicable to various types of cells for both scaffold-free and collagen-based approaches. Examples of printed tissues include blood vessels, liver, and 3D constructs of MSCs for regeneration of osteochondral defects.  Further applications with neural cells and cardiomyocytes are currently being explored by Japanese academia.  The system is commercially available in Japan and will become so in the US in 2015. We are seeking potential users in the US market.

Networking Lunch, Visit the Exhibitors and Poster Viewing

Technology Spotlight:
Build with Life
Danny Cabrera, CEO, Biobots Inc

Bioprinting is challenging. There are numerous barriers to entry – biomaterials expertise, costs, unreliable printing platforms, and difficult to use software, to name a few. BioBots is addressing each of these constraints with a new line of easy to use, flexible, high resolution, low cost 3D bioprinters. Our devices are making 3D bioprinting accessible to researchers around the world and contributing widely to tissue engineering.

Computer-Aided Design and Evaluation of Bio-fabricated Scaffolds
Wojciech Swieszkowski, Professor in Materials Design Division, Faculty of Materials Science and Engineering, Warsaw University of Technology, Poland

This study shows how to design and preclinically evaluate the biofabricated scaffolds for othopeadic applications using experimental and numerical methods. Biodegradable scaffolds, with honeycomb-like pattern, fully interconnected channel network, and controllable porosity, were fabricated in the form of layers of directionally aligned microfibres deposited via computer-controlled extrusion process. The influence of the applied biomaterials and internal architecture of the scaffolds on their mechanical and biological properties was evaluated based on experimental results. Moreover, the novel numerical methods were developed to predict mechanical and biological performances of the tissue engineering constructs taking into account their degradation conditions and cell growth. The obtained results have confirmed the usefulness of the presented experimental and numerical approaches in development of new constructs for tissue engineering.

Coffee Break, Networking, Exhibit and Poster Viewing

Vascular Patches for Pediatric Vascular Tissue Engineering
Joyce Wong, Professor of Biomedical Engineering and Materials Science & Engineering, Boston University, United States of America

This talk will focus on methods to generate vascular patches for pediatric vascular tissue engineering applications. We have developed methods involving cell sheet engineering to control the structural organization and functional properties of vascular patches. We will also discuss how computational methods can be integrated with experimental techniques for iterative tissue engineering design and validation.

Three Dimensional Printing of Biodegradable Cardiovascular Biomaterials
John Fisher, Fischell Family Distinguished Professor & Department Chair; Director, NIH Center for Engineering Complex Tissues, University of Maryland, United States of America
Anthony Melchiorri, , Fischell Department of Bioengineering, United States of America

Rapid advances in three dimensional (3D) printing, whether based on extrusion or lithography, are bringing the technology closer to clinical use in tissue engineering.  3D printing allows for precise bulk geometry and interior architecture control.  Based upon material selection, the resolution of the interior and bulk geometry can typically range from 25 µm to 100 µm.  Using 3D printing, we have developed biodegradable scaffolds with custom pore sizes.  These scaffolds have been non-destructively analyzed with microcomputed tomography to calculate changes in scaffold pore size, porosity, and wall thickness during a 16 week degradation study.  One potential clinical application of 3D printing is for the treatment of cardiovascular diseases, through which outcomes may be drastically improved with custom fabrication of patient-specific grafts.  Current clinical strategies rely upon surgeons constructing tailor-made implants during surgery with generic grafts.  Advancements in imaging technologies, such as MRI and CT, allow for the production of high-quality 3D images from which patient-specific grafts and implants can be generated prior to surgery.  To this end, we have formulated a poly(propylene fumarate) (PPF) based resin to 3D print vascular features with mechanical properties similar to native blood vessels.  The grafts support the growth of vascular cells in vitro and support good neotissue formation after one and three months in vivo.  Further, these grafts may be modified with biomolecules, such as VEGF, so as to recruit and mobilize endothelial cell populations and thus quicken graft endothelialization.

Open Source 3D Printing for Tissue Engineering: From Melt Extrusion to Laser Sintering
Jordan Miller, Assistant Professor of Engineering and Founder of the Advanced Manufacturing Research Program, Rice University, United States of America

The ethos of the open-source software movement—making designs and code, like the Linux computer operating system, freely and legally available to anyone—has now bled into the software toolchain and hardware designs for consumer-level 3D printers. The diversity of interests in 3D printing, coupled with the wide distribution of printers themselves, mean a plethora of opportunities exist for applying 3D printing technologies to biology and medicine. Here, we describe efforts to open-source 3D printing hardware and software for melt-extrusion, inkjet printing, and selective laser sintering and describe applications of these technologies to tissue engineering. Opportunities to formally engage with the “Maker” community will be highlighted which are infusing new ideas and increased attention to tissue engineering research, while simultaneously dropping costs and removing the technical and educational barriers to advanced 3D printing technologies.

Engineering Synthetic Tissues for Treatment of Chronic Liver Disease
Kelly Stevens, Research Scientist, Massachusetts Institute of Technology, United States of America

Cell-based therapies for organ regeneration have recently emerged as a potential alternative to whole-organ transplantation. Unfortunately, orthotopic cell-based therapy may not be feasible or effective in all diseased organs. For example, in end-stage liver disease, the inhospitable fibrotic microenvironment in cirrhotic liver is likely to limit cellular engraftment. Our goal is to build functionally stabilized engineered tissues that can be implanted ectopically and ultimately used to contribute to host liver functions. To create functionally stable engineered liver tissue, we have developed versatile microtissue molding and bioprinting-based methods that enables rapid, scalable, and multicompartmental cellular placement in various patterns and material systems across tissue sizes relevant for in vitro, pre-clinical, and clinical biologic studies. We have used these methods to identify multicellular architectural tissue configurations that best support parenchymal primary human or induced pluripotent stem cell (iPS)-derived hepatocyte survival and function in vitro and in vivo. Ongoing work seeks to extend these findings for application liver repair and regeneration using canonical liver injury model systems.

Close of Day 2 of the Conference