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What Happens After Nerve Injury: Understanding Human Motor Endplate Degeneration after Peripheral Nerve Injury
Luigi Porciuncula Gonzales, BS1; Vivian Chen, BS1; Tyler R Johnston, MD1; Oswald Steward, PhD1; Ranjan Gupta, MD2
1University of California, Irvine, Irvine, CA; 2Orthopedic Surgery, University of California, Irvine, Orange, CA

Introduction: There is often limited functional recovery after peripheral nerve injury (PNI) despite optimal surgical intervention. One of the reasons for these poor outcomes is a time-dependent degeneration of denervated motor endplates (MEPs) at neuromuscular junctions (NMJs). Prior studies have demonstrated that human MEPs have different morphology, size, and density than murine MEPs. Moreover, our recent studies indicate that unlike mouse and rat models, some MEPs in humans persist even >6 months post-PNI. These results suggest that degeneration of denervated human MEP may not be analogous to murine MEP degradation. Here, we use a new methodology for NMJ visualization to assess human NMJ degeneration in PNI patients.
Materials and Methods:Denervated and innervated muscle samples were obtained in patients undergoing standard of care surgery 3-6 months post-PNI. Samples were prepared either (1) by immunostaining sections for ?-BTX, neurofilament (NF) and synaptophysin (SYN) or (2) clearing the whole muscle sample with CUBIC R1 solution and immunostaining with NF, SYN and acetylcholine receptor (AChR)-?. Muscle samples were imaged with a Keyence BZ-X810 microscope.
Results: MEPs in denervated tissue exhibited morphological degenerative changes when compared to innervated muscle including a mix of abnormal morphologies ranging from pretzel to fragmented. At similar time points post-injury, some patients had greater NMJ degeneration with a higher proportion of plaque MEPs compared to others. MEP morphology was better defined in samples immunostained for AChR-a than with a-BTX. CUBIC-clearing prior to immunostaining allowed for excellent antibody penetration, enabling 3D reconstruction and morphometric quantification of MEP integrity in biopsied muscle.
Conclusions: Our data indicate that (1) human NMJ degeneration is species specific and (2) preserved MEPs in humans predict greater functional recovery with nerve repair surgery. Because human NMJ degeneration shows tremendous variation, some patients have the potential for recovery with surgery even if performed >6 months post-injury. These data suggest a role for pre-operative muscle biopsy prior to reconstructive surgery after a significant nerve injury.


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