Table of contents

BF2010, the 2010 International Conference on Biofabrication was held in Philadelphia, Pennsylvania, USA (4–6 October 2010). The objective of BF2010 was to provide a broad communication venue for multidisciplinary scientists, researchers and industrial participants to exchange and disseminate recent scientific discoveries, research, development and emerging applications in the field of biofabrication, to promote international collaboration, and to explore new directions in research on biofabrication. The conference's major topics included, but were not limited to, the following thrust areas:

• Bioprinting of cells, proteins, and biologics, including inkjet printing, bioplotting, biological laser printing and other novel bioprinting technologies;

• Biofabrication of biological models, tissue models, disease pathogeneses models, drug toxicological and discovery models, and cell/tissue-on-a-chip systems;

• Biofabrication of tissue scaffolds and tissue engineered substitutes for regenerative medicine;

• Integrated bio-micro (micro-bio) and bio-nano (nano-bio) fabrication, bio-additive manufacturing, and the state-of-the-art and future of biomaterials suitable for bioprinting and biofabrication;

• Design, model, and evaluation of the biofabrication process; computer-aided biofabrication; modeling on biofabricated structures, cell aggregates and tissue ingrowth;

• Design and fabrication of various drug delivery vehicles;

• Biofabrication industry, trends and future directions.

BF2010 embraced over 130 attendees representing the following countries: Portugal, France, The Republic of Korea, China, the USA, the UK, Australia, Japan, Italy, Poland, Germany, Canada, The Netherlands, Greece, and Brazil. This was the largest gathering of the international biofabrication community. The BF2010 technical program consisted of three full-day 16 parallel sessions with six keynote presentations, 88 oral presentations and 26 poster presentations. The poster presentations were reviewed by selected judges from the BF2010 scientific committee along with participants of the conference. The top three posters were awarded at the closing ceremony. At the closing ceremony Professor Makoto Nakamura announced `BF2011 in Toyama' (www.lni.co.jp/biofabrication2011/).

One milestone event of BF2010 was the launch of the International Society of Biofabrication (ISBF), a scientific and professional non-profit society, which promotes advances in biofabrication research, development, education, training, and medical and clinical applications. ISBF's core purpose is to foster scientific and technological innovation and excellence for the benefit of humanity. ISBF promotes interaction between the different disciplines of the field of biofabrication as well as between basic research and applied practice. An important objective of ISBF is cooperation with other scientific organizations and communities.

This special issue is a selection of 15 papers from the BF2010 conference that are representative of the recent developments in the field of biofabrication. Several selected papers introduce various enabling techniques for the fabrication of three-dimensional (3D) tissue scaffolds. For example, in the paper `Development of a hybrid scaffold with synthetic biomaterials and hydrogel using solid freeform fabrication technology', Shim et al at POSTECH, Korea, report the development of fabricating a 3D hybrid scaffold that consists of synthetic biomaterials and a natural hydrogel using the multi-head deposition system (MHDS) based on additive manufacturing technology. Zhu et al from the University of Saskatchewan, Canada, describe the `Development of novel hybrid poly(l-lactide)/chitosan scaffolds using the rapid freeze prototyping technique'. In the reported work, 3D scaffolds are fabricated from a mixture of chitosan microspheres (CMs) and poly(l-lactide) by means of a rapid freeze prototyping (RFP) technique. Hamid et al at Drexel University, USA, present `Fabrication of three-dimensional scaffolds using precision extrusion deposition with an assisted cooling device'. They apply a precision extrusion deposition (PED) technique to fabricate micro-scaled scaffolds with high printing resolution, precision, and control for a special set of biopolymers. In `Laser sintering fabrication of three-dimensional tissue engineering scaffolds with a flow channel network' Niino et al at Tokyo University, Japan, introduce a process in which a biodegradable plastic powder is mixed with fine salt grains and processed by laser sintering additive manufacturing technology, to develop a scaffold with a fine flow channel network.

Various cell printing/deposition techniques are also reported in this special issue. Work on `Three-dimensional inkjet biofabrication based on designed images' is reported by Arai et al from Toyama University, Japan. They introduce a custom-made inkjet printer, `Inkjet 3D bioprinter', which is based on inkjet technology for fabricating 3D structures composed of living cells and biomaterials with which semi-solid hydrogel structures can be constructed in liquid medium. Hamon et al from Tokyo University, Japan, report a new methodology for engineering thick liver tissues `Avidin–biotin-based approach to forming heterogenic cell clusters and cell sheets on a gas-permeable membrane'. They describe a new methodology for the formation of a functional thick hepatic tissue usable for cell sheet technology.

A `Laser-guidance-based cell deposition microscope for heterotypic single-cell micropatterning' by Ma et al at Clemson University, USA, is reported, which is capable of producing high resolution micropatterns of different cell types on a substratum and allows for on-stage incubation for long-term cell culturing. Tejavibulya et al at Brown University, USA, report `Directed self-assembly of large scaffold-free multi-cellular honeycomb structures'. The authors use a direct self-assembly approach to create large multi-cellular honeycomb building blocks, whereby cell-to-cell adhesion drives the formation of a 3D structure.

Work on `Bioprinting cell-laden matrigel for radioprotection study of liver by pro-drug conversion in dual-tissue microfluidic chip' is reported by Snyder et al from Drexel University, USA. The paper introduces a combined cell-printing and microfabrication technique to develop a microfluidic system to study drug conversion and radiation protection of living liver tissue analogs. In `Microengineering methods for cell-based microarrays and high-throughput drug-screening applications' Xu et al from Harvard University, USA, report the use of microengineering technology to produce 3D cell-based drug-screening assays.

Work on `Biofabrication of chitosan–silver composite SERS substrates enabling quantification of adenine by spectroscopic shift' is reported by Luo et al from the University of Maryland, USA. They present a new biofabrication strategy using surface-enhanced Raman scattering (SERS) to fabricate substrates that enable the quantification through a newly discovered spectroscopic shift due to chitosan–analyte interactions.

In `Design and fabrication of a novel porous implant with pre-set channels based on ceramic stereolithography for vascular implantation', Bian et al at Xi'an Jiaotong University, China, report a novel biomimetic design approach for blood vessel implants with pre-set channels to treat early femoral head necrosis. Khoda et al from the University at Buffalo, USA, introduce a novel continuous tool path planning methodology to design `A functionally gradient variational porosity architecture for hollowed scaffolds fabrication'. In `CAD/CAM-assisted breast reconstruction' Melchels et al from Queensland University of Technology, Australia, report on the development of a generic algorithm for the design and control of porosity patterns within a scaffold.

We hope that the selected papers will be of interest to the reader, and can encourage and stimulate further research in the field of biofabrication. We also would like to take this opportunity to thank all contributors and reviewers for their support, and thank IOP Publishing for publishing this special issue.

The SC Project is an alliance of 10 colleges and universities working together to achieve the goal of engineering a functional, 3D, bioengineered construct. Scientific progress includes computational modeling of vascular trees and experimental testing of natural and engineered constructs. Future directions of the science focus on overcoming challenges such as scalability, sustainability of biofabricated constructs, and identification of chemical or physiological factors that can accelerate the differentiation and maturation of biofabricated vascular tissues (maturogens). Studies include those of hemodynamic forces or growth factors that can promote expression and assembly of collagen and elastin fibers.

Screening for effective therapeutic agents from millions of drug candidates is costly, time consuming, and often faces concerns due to the extensive use of animals. To improve cost effectiveness, and to minimize animal testing in pharmaceutical research, in vitro monolayer cell microarrays with multiwell plate assays have been developed. Integration of cell microarrays with microfluidic systems has facilitated automated and controlled component loading, significantly reducing the consumption of the candidate compounds and the target cells. Even though these methods significantly increased the throughput compared to conventional in vitro testing systems and in vivo animal models, the cost associated with these platforms remains prohibitively high. Besides, there is a need for three-dimensional (3D) cell-based drug-screening models which can mimic the in vivo microenvironment and the functionality of the native tissues. Here, we present the state-of-the-art microengineering approaches that can be used to develop 3D cell-based drug-screening assays. We highlight the 3D in vitro cell culture systems with live cell-based arrays, microfluidic cell culture systems, and their application to high-throughput drug screening. We conclude that among the emerging microengineering approaches, bioprinting holds great potential to provide repeatable 3D cell-based constructs with high temporal, spatial control and versatility.

Natural biomaterials such as hyaluronic acid, gelatin and collagen provide excellent environments for tissue regeneration. Furthermore, gel-state natural biomaterials are advantageous for encapsulating cells and growth factors. In cell printing technology, hydrogel which contains cells was printed directly to form three-dimensional (3D) structures for tissue or organ regeneration using various types of printers. However, maintaining the 3D shape of the printed structure, which is made only of the hydrogel, is very difficult due to its weak mechanical properties. In this study, we developed a hybrid scaffold consisting of synthetic biomaterials and natural hydrogel using a multi-head deposition system, which is useful in solid freeform fabrication technology. The hydrogel was intentionally infused into the space between the lines of a synthetic biomaterial-based scaffold. The cellular efficacy of the hybrid scaffold was validated using rat primary hepatocytes and a mouse pre-osteoblast MC3T3-E1 cell line. In addition, the collagen hydrogel, which encapsulates cells, was dispensed and the viability of the cells observed. We demonstrated superior effects of the hybrid scaffold on cell adhesion and proliferation and showed the high viability of dispensed cells.

Being a multi-etiological factors disease, osteonecrosis of the femoral head affects many young people, leading to the collapse of the femur head; eventually the hip arthroplasty is needed if not treated in time. Unfortunately, as yet, no satisfactory therapy to repair necrotic bone at an early stage is present. Novel implants with pre-set channels were designed for the treatment of early femoral head necrosis. Ceramic stereolithography was applied to fabricate the green part from β-TCP powder. Other processes, such as dehydration, rinsing, drying and sintering, were processed successively. The final ceramic part remains the same as the engineered part in both shape and internal structure. No significant deformation or crack occurred. X-ray diffraction showed that no facies changed or chemical reaction occurred during the fabrication process. The chemical composition remains the same as that of the original β-TCP powder. The compressive strength is 23.54 MPa, close to that of natural cancellous bone. Novel implants with a pre-set channel were designed and fabricated for blood vessel implantation. Bioceramic stereolithography technology based directly on the CAD model in this research shows advantages in accurate design, optimization of 3D scaffold and critical control of the fabrication process. This proposed implant shows promising clinical application in the restoration of early femoral head necrosis.

The fabrication of tissue engineering scaffolds for the reconstruction of highly oxygen-dependent inner organs is discussed. An additive manufacturing technology known as selective laser sintering was employed to fabricate a highly porous scaffold with an embedded flow channel network. A porogen leaching system was used to obtain high porosity. A prototype was developed using the biodegradable plastic polycaprolactone and sodium chloride as the porogen. A high porosity of 90% was successfully obtained. Micro x-ray CT observation was carried out to confirm that channels with a diameter of approximately 1 mm were generated without clogging. The amount of residual salt was 930 µg while the overall volume of the scaffold was 13 cm³, and it was confirmed that the toxicity of the salt was negligible. The hydrophilization of the scaffold to improve cell adhesion on the scaffold is also discussed. Oxygen plasma ashing and hydrolysis with sodium hydroxide, typically employed to improve the hydrophilicity of plastic surfaces, were tested. The improvement of hydrophilicity was confirmed by an increase in water retention by the porous scaffold from 180% to 500%.

Engineered scaffolds have been shown to be critical to various tissue engineering applications. This paper presents the development of a novel three-dimensional scaffold made from a mixture of chitosan microspheres (CMs) and poly(l-lactide) by means of the rapid freeze prototyping (RFP) technique. The CMs were used to encapsulate bovine serum albumin (BSA) and improve the scaffold mechanical properties. Experiments to examine the BSA release were carried out; the BSA release could be controlled by adjusting the crosslink degree of the CMs and prolonged after the CMs were embedded into the PLLA scaffolds, while the examination of the mechanical properties of the scaffolds illustrates that they depend on the ratio of CMs to PLLA in the scaffolds as well as the cryogenic temperature used in the RFP fabrication process. The chemical characteristics of the PLLA/chitosan scaffolds were evaluated by Fourier transform infrared (FTIR) spectroscopy. The morphological and pore structure of the scaffolds were also examined by scanning electron microscopy and micro-tomography. The results obtained show that the scaffolds have higher porosity and enhanced pore size distribution compared to those fabricated by the dispensing-based rapid prototyping technique. This study demonstrates that the novel scaffolds have not only enhanced porous structure and mechanical properties but also showed the potential to preserve the bioactivities of the biomolecules and to control the biomolecule distribution and release rate.

This paper presents a novel continuous tool-path planning methodology for hollowed scaffold fabrication in tissue engineering. A new functionally gradient porous architecture is proposed with a continuous material deposition planning scheme. A controllable variational pore size and hence the porosity have been achieved with a combination of two geometrically oriented consecutive layers. The desired porosity has been achieved with consecutive layers by geometrically partitioning each layer into sub-regions based on the area and the tissue scaffold design constraints. A continuous, interconnected and optimized tool-path for layers has been generated for a three-dimensional biomaterial deposition/printing process. A zigzag pattern tool-path has been proposed for an accumulated sub-region layer, and a concentric spiral-like optimal tool-path pattern has been generated for the successive layer to ensure continuity along the structure. Three-dimensional layers, formed by the proposed tool-path plan, vary the pore size and the porosity based on the biological and mechanical requirements. Several examples demonstrate the proposed methodology along with illustrative results. Also a comparative study between the proposed design and conventional Cartesian coordinate scaffolds has been performed. The results demonstrate a significant reduction in design error with the proposed method. Moreover, sample examples have been fabricated using a micro-nozzle biomaterial deposition system, and characterized for validation.

Cell patterning methods enable researchers to control specific homotypic and heterotypic contact-mediated cell–cell and cell–ECM interactions and to impose defined cell and tissue geometries. To micropattern individual cells to specific points on a substrate with high spatial resolution, we have developed a cell deposition microscope based on the laser-guidance technique. We discuss the theory of optical forces for generating laser guidance and the optimization of the optical configuration (NA ≈ 0.1) to manipulate cells with high speed in three dimensions. Our cell deposition microscope is capable of patterning different cell types onto and within standard cell research devices and providing on-stage incubation for long-term cell culturing. Using this cell deposition microscope, rat mesenchymal stem cells from bone marrow were micropatterned with cardiomyocytes into a substrate microfabricated with polydimethylsiloxane on a 22 mm × 22 mm coverglass to form a single-cell coculturing microenvironment, and their electrophysiological property changes were investigated during the coculturing days.

Surface-enhanced Raman scattering (SERS) has grown dramatically as an analytical tool for the sensitive and selective detection of molecules adsorbed on nano-roughened noble metal structures. Quantification with SERS based on signal intensity remains challenging due to the complicated fabrication process to obtain well-dispersed nanoparticles and well-ordered substrates. We report a new biofabrication strategy of SERS substrates that enable quantification through a newly discovered spectroscopic shift resulting from the chitosan–analyte interactions in solution. We demonstrate this phenomenon by the quantification of adenine, which is an essential part of the nucleic acid structure and a key component in pathways which generate signal molecules for bacterial communications. The SERS substrates were fabricated simply by sequential electrodeposition of chitosan on patterned gold electrodes and electroplating of a silver nitrate solution through the chitosan scaffold to form a chitosan–silver nanoparticle composite. Active SERS signals of adenine solutions were obtained in real time from the chitosan–silver composite substrates with a significant concentration-dependent spectroscopic shift. The Lorentzian curve fitting of the dominant peaks suggests the presence of two separate peaks with a concentration-dependent area percentage of the separated peaks. The chitosan-mediated composite SERS substrates can be easily biofabricated on predefined electrodes within microfluidic channels for real-time detection in microsystems.

In the field of biofabrication, tissue engineering and regenerative medicine, there are many methodologies to fabricate a building block (scaffold) which is unique to the target tissue or organ that facilitates cell growth, attachment, proliferation and/or differentiation. Currently, there are many techniques that fabricate three-dimensional scaffolds; however, there are advantages, limitations and specific tissue focuses of each fabrication technique. The focus of this initiative is to utilize an existing technique and expand the library of biomaterials which can be utilized to fabricate three-dimensional scaffolds rather than focusing on a new fabrication technique. An expanded library of biomaterials will enable the precision extrusion deposition (PED) device to construct three-dimensional scaffolds with enhanced biological, chemical and mechanical cues that will benefit tissue generation. Computer-aided motion and extrusion drive the PED to precisely fabricate micro-scaled scaffolds with biologically inspired, porosity, interconnectivity and internal and external architectures. The high printing resolution, precision and controllability of the PED allow for closer mimicry of tissues and organs. The PED expands its library of biopolymers by introducing an assisting cooling (AC) device which increases the working extrusion temperature from 120 to 250 °C. This paper investigates the PED with the integrated AC's capabilities to fabricate three-dimensional scaffolds that support cell growth, attachment and proliferation. Studies carried out in this paper utilized a biopolymer whose melting point is established to be 200 °C. This polymer was selected to illustrate the newly developed device's ability to fabricate three-dimensional scaffolds from a new library of biopolymers. Three-dimensional scaffolds fabricated with the integrated AC device should illustrate structural integrity and ability to support cell attachment and proliferation.

A significant challenge to the field of biofabrication is the rapid construction of large three-dimensional (3D) living tissues and organs. Multi-cellular spheroids have been used as building blocks. In this paper, we create large multi-cellular honeycomb building blocks using directed self-assembly, whereby cell-to-cell adhesion, in the context of the shape and obstacles of a micro-mold, drives the formation of a 3D structure. Computer-aided design, rapid prototyping and replica molding were used to fabricate honeycomb-shaped micro-molds. Nonadhesive hydrogels cast from these micro-molds were equilibrated in the cell culture medium and seeded with two types of mammalian cells. The cells settled into the honeycomb recess were unable to attach to the nonadhesive hydrogel and so cell-to-cell adhesion drove the self-assembly of a large multi-cellular honeycomb within 24 h. Distinct morphological changes occurred to the honeycomb and its cells indicating the presence of significant cell-mediated tension. Unlike the spheroid, whose size is constrained by a critical diffusion distance needed to maintain cell viability, the overall size of the honeycomb is not limited. The rapid production of the honeycomb building unit, with its multiple rings of high-density cells and open lumen spaces, offers interesting new possibilities for biofabrication strategies.

Implantation of sheet-like liver tissues is a promising method in hepatocyte-based therapies, because angiogenesis is expected to occur upon implantation from the surrounding tissues. In this context, we introduce here a new methodology for the formation of a functional thick hepatic tissue usable for cell sheet technology. First, we report the formation of composite tissue elements in suspension culture. Composite elements were composed of human hepatoma Hep G2 cells and mouse NIH/3T3 fibroblasts which are important modulators for thick-tissue formation. To overcome the very low attachment and organization capability between different cells in suspension, we synthesized a new cell-to-cell binding molecule based on the avidin–biotin binding system that we previously applied to attach hepatocytes on artificial substrata. This newly synthesized biotin-conjugated biocompatible anchoring molecule was inserted in the plasma membrane of both cell types. NIH/3T3 cells were further conjugated with avidin and incubated with biotin-presenting Hep G2 cells to form highly composite tissue elements. Then, we seeded those elements on highly gas-permeable membranes at their closest packing density to induce the formation of a thick, composite, functional hepatic tissue without any perfusion. This methodology could open a new way to engineer implantable thick liver tissue sheets where different cell types are spatially organized and well supplied with oxygen.

The objective of this paper is to introduce a novel cell printing and microfluidic system to serve as a portable ground model for the study of drug conversion and radiation protection of living liver tissue analogs. The system is applied to study behavior in ground models of space stress, particularly radiation. A microfluidic environment is engineered by two cell types to prepare an improved higher fidelity in vitro micro-liver tissue analog. Cell-laden Matrigel printing and microfluidic chips were used to test radiation shielding to liver cells by the pro-drug amifostine. In this work, the sealed microfluidic chip regulates three variables of interest: radiation exposure, anti-radiation drug treatment and single- or dual-tissue culture environments. This application is intended to obtain a scientific understanding of the response of the multi-cellular biological system for long-term manned space exploration, disease models and biosensors.

Tissue engineering has been developed with the ultimate aim of manufacturing human organs, but success has been limited to only thin tissues and tissues with no significant structures. In order to construct more complicated tissues, we have developed a three-dimensional (3D) fabrication technology in which 3D structures are directly built up by layer-by-layer printing with living cells and several tissue components. We developed a custom-made inkjet printer specially designed for this purpose. Recently, this printer was improved, and the on-demand printing mode was developed and installed to fabricate further complicated structures. As a result of this version, 3D layer-by-layer printing based on complicated image data has become possible, and several 2D and 3D structures with more complexity than before were successfully fabricated. The effectiveness of the on-demand printing mode in the fabrication of complicated 3D tissue structures was confirmed. As complicated 3D structures are essential for biofunctional tissues, inkjet 3D biofabrication has great potential for engineering complicated bio-functional tissues.

The application of computer-aided design and manufacturing (CAD/CAM) techniques in the clinic is growing slowly but steadily. The ability to build patient-specific models based on medical imaging data offers major potential. In this work we report on the feasibility of employing laser scanning with CAD/CAM techniques to aid in breast reconstruction. A patient was imaged with laser scanning, an economical and facile method for creating an accurate digital representation of the breasts and surrounding tissues. The obtained model was used to fabricate a customized mould that was employed as an intra-operative aid for the surgeon performing autologous tissue reconstruction of the breast removed due to cancer. Furthermore, a solid breast model was derived from the imaged data and digitally processed for the fabrication of customized scaffolds for breast tissue engineering. To this end, a novel generic algorithm for creating porosity within a solid model was developed, using a finite element model as intermediate.