Three-dimensional photopatterning of hydrogels using stereolithography for long-term cell encapsulation† Vincent Chan, ad Pinar Zorlutuna, ad Jae Hyun Jeong, b Hyunjoon Kong b and Rashid Bashir * acd Received 16th March 2010, Accepted 11th June 2010 DOI: 10.1039/c004285d Cell-encapsulated hydrogels with complex three-dimensional (3D) structures were fabricated from photopolymerizable poly(ethylene glycol) diacrylate (PEGDA) using modified ‘top-down’ and ‘bottoms-up’ versions of a commercially available stereolithography apparatus (SLA). Swelling and mechanical properties were measured for PEGDA hydrogels with molecular weights (M w ) ranging from 700 to 10 000 Daltons (Da). Long-term viability of encapsulated NIH/3T3 cells was quantitatively evaluated using an MTS assay and shown to improve over 14 days by increasing the M w of the hydrogels. Addition of adhesive RGDS peptide sequences resulted in increased cell viability, proliferation, and spreading compared to pristine PEG hydrogels of the same M w . Spatial 3D layer-by- layer cell patterning was successfully demonstrated, and the feasibility of depositing multiple cell types and material compositions into distinct layers was established. Introduction The need for in vitro 3D model systems that can substitute for specific tissues is becoming increasingly prevalent in applications ranging from fundamental scientific studies, cancer metastases, stem cell biology, drug discovery, and the replacement of organs. 1 Native tissues are composed of heterogeneous mixtures of cell types and extracellular matrix (ECM) molecules that are arranged in complex 3D hierarchies and supported by an intricate network of blood vessels. Mimicking the spatial organization of these cells and ECM molecules is one of the major challenges toward developing tissue equivalents. Hydrogels have been of particular interest as biomaterial scaffolds in these systems because of their close resemblance to native tissues. They are crosslinked polymer networks that are highly hydrated and possess tissue-like elasticity. 2,3 In particular, poly(ethylene glycol) (PEG) is a synthetic hydrogel that has been widely used because of its hydrophilicity, biocompatibility, and ability to be chemically tailored. 4 Cell adhesion domains, 5,6 growth factors, 7 and hydrolytic 8 and proteolytic 9,10 sequences have also been incorporated into PEG hydrogels to guide cellular processes such as differentiation, proliferation, and migration. By modifying the ends with either acrylates or methacrylates, PEG hydrogels can be photocrosslinked in the presence of appropriate initiating agents. 11,12 This type of curing offers spatial control over polymerization that, while increasingly popular, 13–15 has not been fully exploited. Computer-aided design (CAD)-based rapid prototyping tech- nologies have recently been applied as enabling tools that provide excellent spatial control over scaffold architecture. 16,17 Rapid prototyping is the process of creating complex 3D structures by repetitive deposition and processing of individual layers using computer-controlled devices. Usually, a blueprint is developed first and translated into a 3D design in a format that can be used by the rapid prototyping system. The design is sliced into a collection of 2D cross-sectional layers that is then processed into a real 3D structure using layer-by-layer deposition. By controlling the micro- and macro-architecture of these scaffolds, rapid pro- totyping technologies can potentially be used to create artificial vasculature to facilitate the flow of oxygen and nutrients into the construct, thereby increasing the potential size of the tissue. 18,19 Additionally, angiogenic factors can be added to the vasculature to induce the formation of new blood vessels. 20 Because of their excellent spatial control, it is possible to create 3D structures with multi-cellular components that are required for complex tissue function. 21–23 Furthermore, rapid prototyping technologies provide a means for large-scale production of reproducible tissue constructs that can be used in a variety of applications. A common issue in the vast majority of rapid prototyping technologies is the acellular environment in which the scaffolds are fabricated in. Because cells cannot survive these processing conditions, they are normally seeded on top of the scaffolds and induced to migrate and populate into the inner regions. In this approach, however, it is often difficult to obtain scaffolds that are evenly seeded with cells. 24 An approach where it might be possible to entrap cells in the scaffolds during the fabrication process would be very advantageous because of their homoge- neous distribution. One of the technologies that may be mild enough to encapsulate living cells during the fabrication process is stereolithography. The conventional stereolithography apparatus (SLA) uses ultraviolet (UV) light to selectively solidify photosensitive a Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, Illinois, 61801, USA b Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, Urbana, Illinois, 61801, USA c Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, Urbana, Illinois, 61801, USA d 2000 Micro and Nanotechnology Laboratory, University of Illinois at Urbana-Champaign, MC-249, 208 North Wright Street, Urbana, Illinois, 61801, USA. E-mail: rbashir@illinois.edu; Fax: +1 (217) 244-6375; Tel: +1 (217) 333-3097 † Electronic supplementary information (ESI) available: Fig. S1–S5 and other experimental details. See DOI: 10.1039/c004285d 2062 | Lab Chip, 2010, 10, 2062–2070 This journal is ª The Royal Society of Chemistry 2010 PAPER www.rsc.org/loc | Lab on a Chip Downloaded by Massachusetts Institute of Technology on 11 July 2011 Published on 05 July 2010 on http://pubs.rsc.org | doi:10.1039/C004285D View Online