| AIM:Epilepsy is a chronic neurological disorder with extremely disturbing therapeutic consequences. It is characterized by spontaneous recurrent seizures, which also occur in demyelinating diseases of the central nervous system (CNS) with a higher prevalence, such as multiple sclerosis (MS), indicating an association between demyelination and seizures by an unknown mechanism. Furthermore, demyelination occurings observed in CNS of epilepsy patients have been occasionally reported, although no confirmative evidence has yet been given. Thus, by establishing a rat pentylenetetrazol model of epilepsy seizures, and using electroencephalogram (EEG), Western blotting, Enzyme-Linked Immunosorbent Assay (ELISA) and immunohistochemistry, the present study aimed to provide direct evidence for possible myelin sheath damage in the CNS induced by epilepsy seizures of model rats, and to further demonstrate its mechanism and influence on epilepsy seizures. METHODS:1. Study of myelin sheath damage in Penicillin-induced model of acute epilepsy: acute epilepsy model in Sprague-Dawley rats was established following induction by single intracortical injection of Penicillin G, and confirmed by evaluation on rats'behavior and EEG recording; Western Blot was used to analyze the expression of myelin basic protein (MBP) of the hippocampus and cerebral cortex of rats, reflecting the myelin situation in the central nervous system (CNS).2. Study of myelin sheath damage in pentylenetetrazole-induced model of chronic epilepsy: chronic epilepsy model in Sprague-Dawley rats was established following induction by systematic intraperitoneal injection of pentylenetetrazole (PTZ), and confirmed by evaluation on rats'behavior and EEG recording; Western Blot was used to analyze the expression of myelin basic protein (MBP) of the hippocampus and cerebral cortex of rats at time intervals of 2, 4, and 6 weeks, reflecting the myelin situation in the central nervous system (CNS).3. Mechanism study of myelin sheath damage in PTZ-induced model of chronic epilepsy: to determine the possible relationship between myelin sheath damage and oligodendrocyte, dynamics of oligodendrocyte cell generation during PTZ treatment was recorded by observing the expression of NG2 in rat's hippocampus and cerebral cortex; To determine whether immunoreactivity was a possible cause of myelin sheath damage, anti-MBP antibody concentration in the serum of PTZ-induced epilepsy rats and control rats was assessed by ELISA using MBP as antigen; to assess the possible damage of blood-brain barrier (BBB) represented by increased leakage, EB was intravenously injected to PTZ-treated rats (2, 4, and 6 week) and control rats as a tracer, and EB stained coronal sections of the hippocampus and cerebral cortex were recorded by digital photomicrographing; mechanism of myelin sheath damage in PTZ-induced rat model of chronic epilepsy was illustrated by analysis of combined tests above,4. study of protective effect of preventative pharmacotherapy for demyelinating disease in PTZ-induced chronic epilepsy rats: Application of drugs to prevent demyelination of seizures: therapeutic dose of Graham acetate (immune regulator in demyelinating disease treatment) was administered as a preventive treatment for PTZ-treated rats; and evaluation on rats'behavior and EEG recording were used to observe its effect on myelin sheath damage in PTZ-induced chronic epilepsy rats.RESULTS:1. Myelin sheath was not affected in Penicillin-induced acute epilepsy rats: Behavioral and EEG records showed successful establishment of acute epilepsy in rats by penicillin induction; whereas the Western Blot results showed no changes of MBP expression in the hippocampus and cerebral cortex of Penicillin-induced acute epilepsy rats was observed, indicating that myelin sheath was not affected in Penicillin-induced acute epilepsy rats.2. Myelin sheath damage in PTZ-induced chronic epilepsy rats: Behavioral and EEG records showed that PTZ successfully induced chronic epileptic seizures in rats with aggravation of seizures and increased frequency observed during PTZ administration; Western Blot results showed that MBP expression (at time interval of 2, 4, and 6 weeks, respectively) in the hippocampus and cerebral cortex of PTZ -induced chronic epilepsy rats was significantly decreased compared with that in the control group, whereas MBP expression in model group between the three time points was not statistically significant, indicating that myelin sheath damage occurred in the early phase of PTZ-induced chronic epileptic seizures. However, no further damage was observed along with the aggravation of PTZ-induced epileptic seizures in rats.3. Immune mechanism of myelin sheath damage in PTZ-induced chronic epileptic seizures: The immunohistochemical staining method confirmed that myelin sheath damage in PTZ-induced chronic epilepsy rats was not the result of oligodendrocyte destruction, but the autoantibodies against myelin basic protein (MBP) produced in peripheral circulation accompanied by increased permeability of blood-brain barrier (BBB) formed in the development of epileptic seizures. Thus, it is highly indicated that autoimmunoreaction might be the possible cause of myelin sheath damage in PTZ-induced chronic epileptic seizures.4. preventative therapeutic effect of GA on PTZ-induced chronic epileptic seizure in rats: Behavioral record (at time interval of 2, 4, and 6 weeks, respectively) showed that frequency of epileptic seizure in GA treated rats were lower than that in control group, and no significant differences were observed in treatment group at the three time points, indicating GA treatment was capable at seizure control at a lower level. However, the EEG record (at time interval of 2, 4, and 6 weeks, respectively) showed that occurrence of spontaneous epileptic form discharge in GA treated rats was not statistically significant compared with that in control group, indicating that GA treatment had no effect on spontaneous epileptic discharges in PTZ-induced chronic epileptic seizure in rats.CONCLUSION:1. By observing MBP expression in acute and chronic epilepsy rats, we firstly provided experimental evidence for myelin sheath damage in chronic epilepsy seizures induced by PTZ but not acute epilepsy seizures induced by penicilin, and further demonstrated that its possible cause was autoimmunoreaction but not oligodendrocyte destruction in chronic epilepsy.2. By administration of immune regulator in demyelinating disease treatment on PTZ-induced chronic epilepsy rats, we firstly provided experimental evidence for preventative therapeutic effect of GA in way of significant ameliorating behavioral index. These results are important in terms of providing new possible medication for clinical treatment of chronic epilepsy.
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