Self-Assembling Peptide Nanofiber Scaffolds Accelerate Wound Healing Aurore Schneider 1,3 , Jonathan A. Garlick 1,2 , Christophe Egles 1,2 * 1 Division of Cancer Biology and Tissue Engineering, Department of Oral and Maxillofacial Pathology, Tufts University, School of Dental Medicine, Boston, Massachusetts, United States of America, 2 Department of Biomedical Engineering, Tufts University, Medford, Massachusetts, United States of America, 3 Center for Biomedical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America Cutaneous wound repair regenerates skin integrity, but a chronic failure to heal results in compromised tissue function and increased morbidity. To address this, we have used an integrated approach, using nanobiotechnology to augment the rate of wound reepithelialization by combining self-assembling peptide (SAP) nanofiber scaffold and Epidermal Growth Factor (EGF). This SAP bioscaffold was tested in a bioengineered Human Skin Equivalent (HSE) tissue model that enabled wound reepithelialization to be monitored in a tissue that recapitulates molecular and cellular mechanisms of repair known to occur in human skin. We found that SAP underwent molecular self-assembly to form unique 3D structures that stably covered the surface of the wound, suggesting that this scaffold may serve as a viable wound dressing. We measured the rates of release of EGF from the SAP scaffold and determined that EGF was only released when the scaffold was in direct contact with the HSE. By measuring the length of the epithelial tongue during wound reepithelialization, we found that SAP scaffolds containing EGF accelerated the rate of wound coverage by 5 fold when compared to controls without scaffolds and by 3.5 fold when compared to the scaffold without EGF. In conclusion, our experiments demonstrated that biomaterials composed of a biofunctionalized peptidic scaffold have many properties that are well-suited for the treatment of cutaneous wounds including wound coverage, functionalization with bioactive molecules, localized growth factor release and activation of wound repair. Citation: Schneider A, Garlick JA, Egles C (2008) Self-Assembling Peptide Nanofiber Scaffolds Accelerate Wound Healing. PLoS ONE 3(1): e1410. doi:10.1371/journal.pone.0001410 INTRODUCTION Skin functions to provide a physical and chemical interface that protects the host against invasion by toxins and microorganisms and prevents dehydration that can result from loss of barrier function. The loss of skin integrity and function due to wound injury has led to efforts designed to better comprehend the molecular and cellular mechanisms that can optimize wound repair [1,2,3]. The complex nature of wound healing requires the migration and proliferation of keratinocytes that are temporally-regulated by numerous growth factors and their receptors that are upregulated in the wound environment [4,5,6]. The complexity of the wound environment has been recreated in human, bioengineered in vitro 3D tissues known as human skin equivalents (HSE), that have many morphologic and phenotypic properties of human skin. Adaptating HSEs to study wound reepithelization has demonstrated several key responses including cell proliferation, migration, differentiation, growth factor responsiveness and protein expression that mimic the response to wounding seen in human skin [7]. In light of this, we used HSEs to study the effect of a new growth factor-releasing biomaterial on wound reepithelialization. As a mediator of wound repair, Epidermal Growth Factor (EGF), is involved in epidermal regeneration by stimulating the proliferation and migration of keratinocytes at the wound edge [8,9,10], through its interaction with high affinity receptors on both fibroblasts and keratinocytes [11,12]. EGF is thought to play a critical role in wound repair during the first few days after injury, until wound reepithelialization is complete. Due to its relatively short half-life of one hour [13] and its turnover results in loss of occupied receptors and decreased activity [12], it is necessary to apply EGF frequently to a wound to maintain an effective local concentration during initiation of wound healing [13]. Therefore, topical EGF application that would result in the sustained flux of EGF into the wound environment would be a very effective way to locally deliver EGF. To achieve this therapeutic goal, it would be optimal to deliver biologically-meaningful doses of EGF in a wound dressing. Such a bioactive dressing should fulfill several criteria for its optimal function: 1) biocompatibility in the absence of cytotoxicity, 2) easily applied to the wound, able to conform to the wound surface, and easily removable after healing, 3) provides a moist environment and protects the wound against dehydration, 4) allows gas exchange between the wounded tissue and the external environment, 5) biofunctionalized to allow the slow release of bioactive agents. In consideration of these criteria, we have studied if a novel, self- assembling peptide (SAP) nanofiber scaffold combined with EGF, could serve as a bio-active wound dressing. SAP nanofibers have already been shown to serve as excellent materials for a variety of controlled, molecular-release applications [14,15]. The individual nanofiber consists of ionic, self-complementary peptides with 16 amino-acids (RADA16-I, Ac-RADARADARADARADA-CONH2) that undergo self-assembly into hydrogels containing 99.5% w/v water when exposed to physiological media or salt solution. These scaffolds closely mimic the structure and porosity of Academic Editor: Shuguang Zhang, Massachusetts Institute of Technology, United States of America Received September 13, 2007; Accepted December 7, 2007; Published January 9, 2008 Copyright: ß 2008 Schneider et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This study was supported by the Center for Integrated Tissue Engineering at School of Dental Medicine at Tufts University and NIH/NIBIB funds via the P41 Tissue Engineering Resource Center. AS is indebted to the Fondation pour la Recherche Me ´dicale for financial support. Competing Interests: The authors have declared that no competing interests exist. * To whom correspondence should be addressed. E-mail: Christophe.egles@ tufts.edu PLoS ONE | www.plosone.org 1 January 2008 | Issue 1 | e1410