Influence Of Tissue Plasminogen Activator And Its Inhibitors On Neurodegeneration Of Spinal Cord After Sciatic Nerve Transection

Background and Objective Impaired functional recovery after peripheral nerve injury is always a puzzle in neurosurgery. Neuronal degeneration and death induced by axon injury is considered to be one of the main causes. Tissue plasminogen activator (tPA), as the only U.S.FDA-approved thrombolytic medication for the treatment of patients with acute ischemic stroke since 1996, has recently been evidenced to potentiate excitotoxic neuronal death following toxic gultamatergic over-stimulation, as well as increase neuronal damage after focal cerebral ischemia. However, its role in neurodegeneraton of spinal cord after peripheral nerve injury remains to be further investigated. Furthermore, Type-1 plasminogen activator inhibitor (PAI-1) and neuroserpin (NSP) are known as the physiological inhibitors of tPA activity in peripheral tissue and in central nervous system (CNS) respectively. Whether both of them contribute to modulate tPA's role in this process need to be illuminated as well. Hence, we used a unilateral sciatic nerve transaction (SCNT) model in adult Sprague- Dawley rats to examine the dynamic expression of tPA, PAI-1 and NSP, as well as the morphologic change and survival decrease of motor neuron, in the injured side of spinal cord after axon transection, to estimate the role of tPA and its inhibitors in neurodegeneration following SCNT.Materials and Methods Fifty six male Sprague Dawley rats weighing 180～200g were randomly divided into normal group with 6 rats (as control) and SCNT group with 50. Rats were anesthetized with sodium pentobarbital (30mg/kg, i.p.). The right sciatic nerve was transected at 0.5cm away from piriformis muscle. A 1cm portion of the nerve was resected and the distal nerve was imbeded into biceps femoris to prevent regeneration. At several time points (1 day, 4 days, 7 days, 14 days and 21 days) following the SCNT, groups of rats (n=10 each group) were removed with the lumbar 4 to 6 of spinal cord. The segments were sectioned continuously into 5 μm paraffin slices (n=4 each time point) for hematoxylin-eosin (HE) staining, Nissl staining and immunohistochemical staining with antibody to tPA and PAI-1. Ultrastructure change was imaged by transmission electron microscope (TEM) (n=2 each time point). Total RNA was extracted from right side of spinal cord and the mRNA levels for tPA, NSP and PAI-1 were examined by semi-quantitative reverse transcription- polymerase chain reaction (RT-PCR) (n=4 each time point).