Simulated Joint Loading Enhances the Expression of of Superficial Zone Markers in Tissue Engineered Cartilage Constructs
Kaupp, James Arthur
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Tissue engineering is a promising approach for repairing focal defects in articular cartilage. However, using current technologies tissue engineered cartilage displays insufficient biochemical, mechanical and structural properties which compromise the efficacy of implantable material. Researchers have utilized mechanical stimulation as a means to enhance these shortcomings, but few studies have applied mechanical stimulation in a complex manner similar to forces experienced by the tissue in vitro. It is hypothesized that application of simulated joint loading (SJL), a small moving contact area over the surface of tissue engineered constructs, will affect the expression and accumulation of superficial zone specific constituents, leading to improvements in construct functionality. Optimal factors of SJL (i.e. compressive load, frequency and duration) were determined via reverse transcription polymerase chain reaction (RT-PCR) and in situ hybridization (ISH). The optimal combination of factors, chosen via peak levels of superficial zone genes, was discovered to be 9.81 mN, 1 Hz and 15 minutes. A study to determine spatial expression of select superficial zone genes stimulated at optimal factors was investigated via ISH, and contrasted with a finite element model (FEM) of constructs stimulated at optimal parameters displayed a correlation between gene expression and surface and sub-surface stress and strain. SJL at optimal factors (9.81 mN, 1 Hz, 15 minutes) was applied to chondrocyte-agarose hydrogel cultures in long-term studies over a period of four weeks and protein accumulation, expression and mechanical properties was investigated. Intermittent long-term application of SJL enhanced the expression and accumulation of structural proteins and enhanced the compressive and shear properties of tissue engineered constructs over a period of three weeks. Combined, the results illustrate the effectiveness of SJL as a method to affect the short-term expression, long-term accumulation and mechanical properties of superficial zone constituents. Both short and long term experiments illustrated a strain dependent behavior. The mechanism behind these results is unclear, but transduction of strain events by mechanoreceptive elements of integrins, ion channels and the pericellular matrix are potential mediators. This study demonstrated the effectiveness of SJL as a stimulation method, and demonstrated the potential to affect regional expression through alterations of SJL contact mechanics.