The Effect of the Duration of Mechanical Stimulation and Post-Stimulation Culture on the Structure and Properties of Dynamically Compressed Tissue-Engineered Menisci

This study investigated the hypothesis that timing and duration of dynamic compression are integral to regulating extracellular matrix (ECM) assembly of tissue-engineered (TE) menisci. The goal of this study was to examine the effects of varying load and static culture duration on structure, composition, and mechanical properties of TE menisci. We accomplished this by varying the duration of dynamic loading over 4 weeks of culture, and by examining increasing periods of static culture after 2 weeks of dynamic loading. Bovine meniscal fibrochondrocytes were seeded into 2% w/v alginate, crosslinked with CaSO4, injected into anatomical micro-computed tomography-based molds, and post-crosslinked with CaCl2. Meniscal constructs were dynamically compressed three times a week via a custom bioreactor for a total of 2 h, with an hour of rest between loading cycles, for 1, 2, or 4 weeks. They were then placed in static culture. After 4 weeks of culture, increased load duration was found to be beneficial to matrix formation and mechanical properties, with superior mechanical and biochemical properties in samples loaded for 2 or 4 weeks. Further, the mechanical properties of these constructs were similar, suggesting that the additional 2 weeks of loading may not be necessary. Samples loaded for 2 weeks followed by a 4-week static culture period yielded the most mature matrix with significant improvements in collagen bundle formation, 2.8-fold increase in the glycosaminoglycan content, 2-fold increase in the collagen content, and 4.3-fold increase in the compressive equilibrium modulus. Overall, this study demonstrated the importance of timing and duration of loading. By switching to prolonged static culture after 2 weeks of loading, we decreased the amount of ECM lost to the media, while significantly increasing biochemical and mechanical properties of TE menisci.