Identifying Mechanobiological Deficits in Allograft Anterior Cruciate Ligament Reconstructions

FY20 PRMRP Topic Area: Musculoskeletal Health.

Objective: The goal of this project is to understand why a certain type of anterior cruciate ligament (ACL) reconstruction (i.e., allografts) have high failure rates in active military members. Specifically, we will test whether cells in allografts have a detrimental response to mechanical forces and investigate what might cause this negative response. Ultimately, this work will help understand why allografts perform poorly in active individuals.

Background: The ACL is a ligament within the knee that is essential for stable joint motion. Tears of the ACL result in pain and knee dysfunction and also lead to the onset of post-traumatic osteoarthritis. To restore proper knee function and prevent further injury, patients undergo ACL reconstruction surgery, which has become one of the most common procedures performed in orthopaedic surgery. ACL tears are an especially big problem for the military. Active service members are 10 times more likely to tear their ACL compared to the general population. They are also significantly more likely to rupture their reconstruction post-surgery. This is particularly true for reconstructions that replace the ACL with a cadaveric tendon (allograft) compared to a tendon from the patient (autograft). Approximately 25% of allograft reconstructions fail in active service members, which is over twice the rate of autograft failure. Therefore, allografts are not recommended for active military members. However, harvesting an autograft tendon from patients leads to increased postoperative pain, decreased range-of-motion, and muscle weakness. Additionally, if an autograft reconstruction fails, the revision surgery generally requires an allograft since there is not another tendon available from the same patient. Therefore, there is significant clinical interest in improving allograft ACL reconstruction outcomes to match that of autografts.

Data in the literature suggest that the poor clinical performance of allografts is due to biological events post-surgery. Prior to implantation, both graft types have the same mechanical properties, and while the cells in allografts are dead prior to surgery, all the cells in autografts also die immediately after implantation. Following implantation, both grafts types are infiltrated and repopulated by new host cells. Additionally, the grafts undergo significant structural remodeling and an initial drop in mechanical properties (e.g., stiffness). Over time, the graft mechanics slowly recover; however, allografts do not make the same gains in mechanical properties that are typically seen in autografts during the remodeling process.

Our novel overall hypothesis is that allografts exhibit impaired remodeling because cells in allografts have a deficient response to mechanical stimuli. Increased patient activity (and presumably greater graft forces) is associated with the greatest differences in outcomes between allografts and autografts. Additionally, allograft reconstructions contain fewer myofibroblasts, which are highly active cells that are responsible for remodeling fibrous tissues and are specifically activated by mechanical stimuli. Together, this suggests that there may be a difference in how cells in allografts and autografts respond to mechanical forces. Furthermore, allografts have a more disorganized structure and contain more immune cells, both of which influence the production of myofibroblasts and their sensitivity to mechanical forces. Therefore, the objective of this project is to compare the response of cells within allograft and autograft ACL reconstructions to mechanical stimuli and determine whether differences in their response are due to differences in tissue structure and immune cell composition.

Innovation: This proposal is innovative because no study has investigated how cells respond to mechanical forces during ACL graft remodeling. Additionally, while data suggest that immune cell activity affects the success of ACL reconstructions and myofibroblast behavior, differences in the immune cell composition between graft types are unclear. To address these knowledge gaps, we will harvest allograft and autograft reconstructions from rabbits 8 weeks post-surgery, keep them alive in culture, and apply forces to the grafts using a mechanical device mounted on a microscope. This technique enables us to measure how cells respond to mechanical stimuli directly in the grafts themselves and preserves the important interactions that occur between the cells and the graft tissue. Additionally, we will isolate the cells from each graft type and investigate their response to mechanical forces on engineered substrates with varying organization and with different types of immune cells. This will enable us to identify whether these factors contribute to poor allograft remodeling in response to increased mechanical forces.

Ultimate Applicability/Impact: The findings of this work will provide novel insight into the cause of increased allograft failure. Our data will establish the foundation for future opportunities to improve allograft remodeling by manipulating the ways cells sense and respond to mechanical stimuli. Additionally, we can evaluate the efficacy of these treatments by applying them to live grafts in culture exposed to different levels of mechanical forces. Ultimately, this work may lead to future studies aimed at enhancing allograft remodeling and outcomes.