Rapamycin insensitive regulation of engineered ligament structure and function by IGF-1.

Following rupture, the anterior cruciate ligament (ACL) will not heal and therefore more than 400,000 surgical repairs are performed annually. Ligament engineering is one way to meet the increasing need for donor tissue to replace the native ligament; however, currently these tissues are too weak for this purpose. Treating engineered human ligaments with insulin-like growth factor-1 (IGF-1) improves the structure and function of these grafts. Since the anabolic effects of IGF-1 are largely mediated by rapamycin complex I (mTORC1), we used rapamycin to determine whether mTORC1 was necessary for the improvement in collagen content and mechanics of engineered ligaments. The effect of IGF-1 and rapamycin was determined independently and interactions between the two treatments were tested. Grafts were treated for 6 days before mechanical testing and analysis of collagen content. Following 8 days of treatment, mechanical properties increased 34% with IGF-1 and decreased 24.5% with rapamycin. Similarly, collagen content increased 63% with IGF-1 and decreased 36% with rapamycin. Interestingly, there was no interaction between IGF-1 and rapamycin, suggesting that IGF-1 was working in a largely mTORC1-independent manner. Acute treatment with IGF-1 did not alter procollagen synthesis in growth media, even though rapamycin decreased procollagen 55%. IGF-1 decreased collagen degradation 15%, whereas rapamycin increased collagen degradation 10%. Once again, there was no interaction between IGF-1 and rapamycin on collagen degradation. Together, these data suggest that growth factor-dependent increases in collagen synthesis are dependent on mTORC1 activity; however, IGF-1 improves human-engineered ligament mechanics and collagen content by decreasing collagen degradation in a rapamycin-independent manner. How the anticatabolic effects of IGF-1 are regulated have yet to be determined. IGF-1 increases and rapamycin decreases mechanical and material properties of engineered human ligaments by regulating collagen content and concentration. There was no interaction between IGF-1 and rapamycin, suggesting that IGF-1 and rapamycin work independently. We found that IGF-1 improves collagen content by decreasing collagen degradation in a rapamycin-independent manner, whereas growth factor-dependent increases in collagen synthesis are blocked by rapamycin. These data may explain why interventions to increase IGF-1 have not helped rehabilitation.