The main extracellular matrix components of articular cartilage, proteoglycans (PG) and collagens (COL), and their interaction with each other provide the unique biomechanical properties that vary with development, aging, and depth from the articular surface. Negatively-charged aggrecan (AGC), composing ~90% of PG, is mainly responsible for the compressive resistance, the fixed charge density (FCD), and the high osmotic pressure (π_PG) within the tissue. The COL network (CN) provides the restraint that counterbalances π_PG at rest or in compression. This dissertation analyzes the engineering of mechanically functional aggrecan-laden cartilaginous grafts by elucidating the role of PG and its interaction with COL in the compressive properties of articular cartilage and by developing rapid novel methods for shaping, assembling, and concentrating matrix-laden constructs.

The application of a refined FCD–π_PG relationship to native cartilage demonstrated that extrafibrillar FCD and π_PG change with growth, age and depth of the tissue. Mature cartilage from bovine calf, adult and human young sources had higher FCD_EF and π_PG than immature (bovine fetal) or aged (human old) tissue due to COL content variations. Depth-related variations in the strain, FCD_EF, π_PG, and σ_CN profiles for human cartilage revealed the loss of a functional superficial layer in aged cartilage. These findings in native tissue provided guidance for engineered constructs and novel methods for the assembly cells and matrix components in engineered constructs to modulate shape, AGC retention, and matrix content. Molding of chondrocyte-based constructs resulted in shaping on one, two, or neither construct surfaces in combination with biomimetic layering of the chondrocyte subpopulations. Addition of PG aggregates, consisting of AGC with hyaluronan and link proteins, to hydrogel constructs resulted in rapid assembly, enhanced AGC retention, and increased the construct compressive stiffness. Further, the addition of COL to PG-hydrogel constructs increased the compressive properties, highlighting the importance of PGCOL interaction in mechanical function. Finally, the mechanical compaction of these constructs rapidly increased the matrix concentrations and material properties.

These results may be useful in rapidly engineering mechanically functional cartilaginous grafts and facilitate for more rapid application of the grafts into the mechanical demanding environment of an in vivo joint.