Application of Microforming to Create Chondrocyte Home Sites in a Natural Cartilage Matrix

Abstract

Articular cartilage degeneration is a central pathological feature of osteoarthritis. Cartilage in the adult does not regenerate in vivo and, as a result, cartilage damage in osteoarthritis is irreversible. With our ever-aging population, osteoarthritis has become a leading cause of disability and unfortunately, no optimal treatments for osteoarthritis are currently available. To address this problem, a research community is focused on the development of both natural and synthetic biodegradable tissue scaffolds. The scaffolds must contain depressions or holes for the purpose of chondrocyte seeding and growth in order to create an implantable construct. In addition to chondrocytes, cartilage tissue consists of the extracellular matrix (ECM). Studies of many tissue types have established that ECM plays an important role in regulating cell behavior and controlling processes such as tissue differentiation and tumor progression. Unlike most natural tissues, adult cartilage ECM is exceptionally dense and lacking in vascularity, which makes it difficult for chondrocytes to be transplanted directly into the matrix. Current methods of creating cell home sites through chemical decellularization of the ECM degrade the mechanical integrity of the cartilage tissue. The research conducted here used a mechanical, rather than chemical, method to create cell home sites. A novel micropunching machine was developed to fabricate 200 μm diameter holes in cartilage, thereby creating a porous natural scaffold while maintaining a healthy ECM. Equine articular cartilage slices were harvested from the cadaver’s back knee joint and cryo-sectioned into 100 μm thick slices. Using die clearances of 3.7%, 6.8%, and 8.9%, the results indicate that micro-scale holes can be mechanically punched in cartilage tissue. The maximum punching force showed a slight trend of decreasing as die clearance increased, but there was no statistical significance. Punching force, as well as hole size, was highly dependent on sample hydration. Upon inspection, the resulting hole sizes were approximately 50 μm to 150 um, indicating 25% to 75% shrinkage in reference to the male punch diameter. Finally, the resulting hole shape was observed to be slightly non-circular and the edges of the hole exhibited a raggedness, which was indicative of the cartilage tearing during hole punching.