Toward enhancing the durability and integration of engineered articular cartilage
- Author(s): Link, Jarrett Marshall
- Advisor(s): Athanasiou, Kyriacos A
- et al.
Articular cartilage, a connective tissue that provides a lubricious surface for the movement of diarthrodial joints, does not naturally heal and can degenerate to osteoarthritis due to age, disease, or trauma. Although tissue-engineering has the potential for joint restoration, deploying engineered implants in the clinical setting depends on factors such as 1) whether implanted neocartilage constructs can integrate with adjacent, native tissue; and 2) whether the implant can withstand the proinflammatory environment that can result from surgery, trauma, or disease, such as osteoarthritis. Using self-assembled articular cartilage as a model, the global objectives of this work were: 1) to enhance cartilage integration; 2) to investigate how bioactive factors may protect neocartilage against macrophage challenges; and 3) to validate in vivo the preclinical safety profile of our tissue-engineering strategies using a preclinical model.
Using chondroitinase ABC for integration (C-ABCint), the following cartilage integration hindrances were targeted: 1) lack of cells at the interface; 2) repulsive negative charges induced by cartilage glycosaminoglycans (GAGs); 3) extracellular matrix density; and 4) a limited number of stabilizing collagen crosslinks at the interface. It was hypothesized that a combination of C- ABCint and bioactive factors (i.e., TGF-β1, C-ABC, and lysyl oxidase like 2) would enhance integration between native and engineered articular cartilage. Also, it was hypothesized that effective C-ABCint dose would depend on construct maturity, but would not affect construct mechanical properties. It was found that C-ABCint, whose dose depended on construct maturity, and TCL enhanced interface Young’s modulus synergistically and led to increases in interface Young’s modulus up to 11.4-fold. Importantly, construct mechanical properties were not affected. The process of administering a GAG-removal agent such as C-ABCint at the periphery of engineered cartilage could be readily adapted to a clinical setting, due to its simplicity, efficacy, and dose control.
Using a novel, direct co-culture system designed to probe cartilage mechano-immunology, the interplay between differentially stimulated macrophages and engineered neocartilage was also examined. It was hypothesized that stiffer engineered neocartilage would elicit an enhanced inflammatory response, but that the addition of bioactive factors would mitigate tissue damage. Although, confirming our hypothesis, stiff constructs caused a 2.5-fold increase in tumor necrosis factor alpha (TNF-α) secretion compared to softer groups, these stiff tissues also demonstrated an improved ability to withstand macrophage challenge. Regardless of stimulation, macrophages did not decrease TCL-treated construct aggregate modulus when compared to baseline values. The unexpected robustness of stiff constructs against inflammatory factors bodes well for the implantation of these constructs in vivo.
Finally, to validate the preclinical safety of the approaches developed to this point, this work concluded with a large animal study examining the effects of tissue-engineered constructs in an orthotopic defect. Allogeneic, self-assembled constructs were implanted into full-thickness chondral defects in the minipig distal femur. As hypothesized, the allogeneic implants did not lead to adverse local or systemic reaction and, thus, were deemed safe for investigation of the efficacy of repair, moving our approach one step closer to clinical translation.