Investigation of Micropunching Holes in Articular Cartilage for Applications in Tissue Engineering.
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 ... read moreaddress 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. Scaffolds also contain
artificial microtubules to enhance nutrient diffusion during early cellular development.
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 in this study 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 various die clearances and hydration levels, micro-scale holes were
mechanically punched in cartilage tissue. The maximum force required to punch a hole in
the cartilage sample was shown to have a relationship to both clearance of the die and
hydration level of the sample. As the die clearance increased, the maximum punching
force (MPF) decreased. In addition, as a sample dried out the MPF increased. However,
the failure mechanism changed for the different levels of hydration. Saturated samples
failed in tension, while dry samples failed in shear, producing larger resulting hole
sizes. Upon inspection, a 200 µm punch produced 50 µm holes in saturated and
hydrated samples, with fibers extending into the hole. In dry samples, a 200 µm
punch produced 200 µm holes with smooth hole walls. An analytical model was
developed to predict MPF based on the hydration level of the sample and the diameter of
the male punch. The Young's moduli of both porous (micropunched) and nonporous samples
were not found to be significantly different at 4% porosity. Yield occurred at strain
levels much higher than those encountered for in vivo cartilage. Therefore, results
indicate that porous samples maintain their mechanical properties for successful cell
culture and integration in vivo. A preliminary cell culture study indicated that seeded
cells can position themselves in the micro-holes and along the walls of the holes to
successfully take advantage of the three-dimensional architecture. The results of this
research indicate that a micropunching process is adequate to create holes in natural
cartilage tissue for the purpose of seeding progenitor cells and eventual fabrication of
a tissue implant.
Thesis (M.S.)--Tufts University, 2015.
Submitted to the Dept. of Mechanical Engineering.
Advisor: Thomas James.
Committee: Li Zeng, Lauren Black, and Jason Moore.
Keywords: Biomechanics, Mechanical engineering, and Biomedical engineering.read less
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