IMPACT OF AMNIOTIC FLUID STEM CELL CONDITIONED MEDIA ON PERIPHERAL NERVE AND MUSCLE RECOVERY AFTER PERIPHERAL NERVE INJURY

Abstract

Introduction:Peripheral nerve injuries (PNI) are devastating, leading to significant morbidity and functional loss. In the United States alone, approximately 20 million people suffer from PNI annually, with associated healthcare costs reaching $150 billion. Sunderland’s classification describes injury severity, with Class V (neurotmesis) being the most severe, involving complete nerve transection. Autografts remain the gold standard for Class V injuries but are limited by donor site morbidity and availability. Allografts offer an alternative but are constrained by immunogenicity. Tissue-engineered nerve grafts (TENGs) have emerged as a promising option; however, their efficacy in large PNIs remains limited, particularly due to the need for autologous cell seeding. Amniotic fluid stem cell-conditioned medium (AFS-CM) provides a cell-free alternative that leverages the paracrine regenerative properties of stem cells while avoiding complications associated with cell therapies. Research Problem: Our study investigates the effects of AFS-CM on peripheral nerve recovery. Specifically, we examine: (1) Schwann cell proliferation and survival under normative and oxidative stress, (2) functional recovery after PNI through gait analysis and EMG studies, and (3) AFS-CM’s role in nerve remyelination, neuromuscular junction stabilization, and mitigation of muscle tissue injury.

Methods: In vitro analysis was conducted using primary rat Schwann cells to assess the effects of AFS-CM on proliferation, survival, oxidative stress (ROS formation), and antioxidant levels. In vivo experiments involved sciatic nerve transection in CD1 mice, followed by three interventions: (1) direct nerve apposition, (2) hydrogel + AFS-CM-coated silicone-based apposition, and (3) hydrogel-coated silicone-based apposition (control). Recovery was evaluated at two weeks, one month, and two months post-injury using gait analysis, EMG, nerve conduction studies, and immunohistochemistry of the peripheral nerve, motor endplate, and skeletal muscle. Conclusion: AFS-CM promotes Schwann cell proliferation and survival, reducing oxidative stress and protecting against senescence. In in vivo models, AFS-CM improves gait function, enhances nerve remyelination, and reduces oxidative stress markers. Future Directions: Future studies should address the translational applicability of AFS-CM by standardizing treatment delivery, optimizing timing, and identifying key bioactive components critical for peripheral nerve regeneration.