Cartilage molecular engineering using biomimetic aggrecan shows infiltration and distribution of biomimetic aggrecan throughout the cartilage extracellular matrix

Abstract

Event Abstract Back to Event Cartilage molecular engineering using biomimetic aggrecan shows infiltration and distribution of biomimetic aggrecan throughout the cartilage extracellular matrix Evan Phillips1, Maria Lefchak1, Mary Mulcahey2, Katsiaryna Prudnikova1 and Michele Marcolongo1 1 Drexel University, Materials Science, United States 2 Drexel University, College of Medicine, United States Introduction: Aggrecan is a vital component to the hydration and mechanical properties of articular cartilage and is lost during the early stages of osteoarthritis (OA). We propose to molecularly engineer the damaged cartilage with a novel biomimetic aggrecan (BA) to restore hydration and mechanical function. BA mimics the 3-D bottle brush structure and properties of naturally occurring aggrecan and consists of a poly(acrylic acid) (PAA) core with chondroitin sulfate (CS) bristles[1]. As a step towards this objective, we report the effect of BA concentration, BA molecular size, and media ionic strength on BA infiltration into bovine osteochondral plugs and examine BA distribution through the cartilage extracellular matrix (ECM). Materials and Methods: Osteochondral plugs were cored from the femoral condyles of mature bovine knee joints. All BA solutions were synthesized using previously described methods[1], fluorescently labeled with DCCH (7-Diethylaminocoumarin-3-Carboxylic acid, Hydrazide)[2], and reconstituted in PBS. The osteochondral plugs were equilibrated in 1X PBS, then immersed in fluorescently labeled BA solutions for 24 hours with care taken only to expose only the articular surface to the solution. Plugs soaked in 1X PBS 10mg/mL BA10 (180kDa) solution were used to compare all the BA infiltration effects in this experiment. The remaining groups (n=5) were BA250 (MW 2,000kDa), 5mg/mL BA10 concentration, 20mg/mL BA10 concentration, 0.1X PBS solvent, 10X PBS solvent, and a control group (no fluorescence). The samples were then cryosectioned and observed under a confocal microscope through a DAPI filter. The fluorescence was quantified from the images (area percent, Matlab) and significance (α=0.05) was determined using a one-way ANOVA with a Tukey post hoc test. Results: BA molecules passively diffused throughout the cartilage ECM. The BA molecules aggregate around the chondrocytes and were able to diffuse to the tidemark. As the BA concentration increased, the amount of BA which diffused into the cartilage increased. Both size BA molecules infiltrated the cartilage ECM; however, the smaller (BA10, 180kDa) molecule had a higher area percentage than the larger (BA250kDa, 2,000kDa) molecule. There was less infiltration of BA at the lowest ionic strength (0.1XPBS) when compared to 1 and 10X PBS media, but there was little difference between 1 and 10X PBS media. Discussion: We have demonstrated that BA can infiltrate normal articular cartilage, distribute throughout the ECM, and localize around the chondrocytes. This is a first demonstration of cartilage molecular engineering and has been shown in the absence of cellular activity (ex vivo). The potential is to introduce regenerating molecules like BA, even as drug delivery vehicles, into the cartilage ECM in a minimally invasive manner via intra-articular injections. Infiltration through the cartilage surface may serve as method to introduce BA into arthritic cartilage to molecularly engineer the tissue and restore its hydration and mechanical properties. Conclusion: Pain reduction and joint mobility for patients with OA is a major challenge. Restoration of the hydration and mechanical stability of OA cartilage may be achieved by intra-articular injection of BA solutions, allowing for passive diffusion of BA into cartilage ECM. We would like to thank the Clinical and Translational Research Institute at Drexel for funding this project.