DESCRIPTION The objective of this project is to develop a tissue-engineered construct including an engineered nucleus pulposus and annulus fibrosus to treat defects of the annulus fibrosus (AF) and degenerative changes of the intervertebral disc (IVD). Three Specific Aims are pursued: " Specific Aim 1 will consist of establishing in vitro tissue engineered NP fabrication from rabbit and human NP and MSC cells in hyaluronic acid (HA)-based hydrogel constructs, and evaluation of cell viability, gene regulation, ECM production, and biomechanical properties as a function of gel biophysical properties, cell seeding density, and extended culture in a disc-like environment. " Specific Aim 2 will entail establishing and evaluating cellular infiltration and functional maturation engineered AF-like structures starting from rabbit and human AF and MSC seeded nanofibrous scaffolds in vitro. " Specific Aim 3 will combine the tissue-engineered NP and AF developed in Aims 1 and 2 to form a composite disc-like construct, and to then apply this construct to an in vivo rabbit disc model of total discectomy. Efficacy will be assesse via MRI T2 mapping and subsequent ex vivo analysis of motion segment mechanical, biochemical, and histologic properties. Research Design: This study will utilize in vitro tissue culture for Specific Aims 1-2; Specific Aim 3 will utilize a rabbit animal model and magnetic resonance imaging (MRI). Methodology: Specific Aims 1 and 2 are laboratory-based studies that will utilize in vitro tissue culture. Specific Aim 3 will be performed in vivo with a rabbit mdel for discectomy and tissue-engineered disc implantation with subsequent ex vivo MRI of the spine and tissue analysis. The primary study outcome measures for Specific Aims 1-3 are biochemical analysis of tissue culture and biomechanical analysis of tissue-engineered constructs. Additionally, Specific Aim 3 will utilize these same outcomes, as well as implement quantitative MRI T2 maps of the implanted tissue-engineered constructs with comparisons made to control (non-treated discectomy) and adjacent normal discs. Findings: n/a at this time. Clinical Relationships: The goal of this project is to develop tissue-engineered constructs to trea degenerative disc disease as well as to validate radiologic methods to noninvasively assess potential therapeutic treatments of degenerative disc disease. Impact/Significance: It is anticipated that the proposed study will offer an increased understanding of degenerative disc disease and potential novel therapeutic treatment to both reverse degenerative changes as well as decrease risk of recurrent disc herniation. Further, it will provide critical support for the applicant as he develops a research-intensive clinical practice in spine surgery at the Philadelphia VA Medical Center and the University of Pennsylvania.

PUBLIC HEALTH RELEVANCE: This Career Development Award proposal builds on the applicant's prior experience in intervertebral disc biology research, including his experience with tissue culture, small animal surgery, and advanced imaging techniques. In this proposed project, the applicant will form a core mentorship team to develop a translational research program focused on treatment strategies for degenerative disc disease. This strategy will entail the application of tissue-engineered constructs to a rabbit model of intervertebral discectomy. In addition to the research, a core component of this project is the mentoring team supporting the applicant in developing a research foundation that will lead to independent funding and independence as a VA investigator and surgeon-scientist.