Proteomic Study Of Hippocampus And Cerebral Cortex In The Rats Model Of Temporal Lobe Epilepsy

Objective:Epilepsy is a clinical syndrom with over-synchronous abnormal discharge of brain neurons caused by various reasons, and it is the common disease of nervous system. Temporal lobe epilepsy (TLE) is the main form of intractable epilepsy, and its patho-genesis remains poorly understood up tp now. At present, hippocampal sclerosis is a notable pathologic feature of TLE including hippocampal atrophy, neuronal loss, gliosis, disperse of dentate granular cells and mossy fiber sprouting.Abnormal discharge of neurons, which is associated with abnormaity of ion chan-nel preoteins and neurotransmitters, is the basis of electrophysiology in epileptic sei-zure. The abnormal discharge causes continuous spread of abnormal potential through repeating synaptic connections and is widely projected to the cerebral cortex, so that epileptic seizures of different forms appear. Therefore, the pathological alterations of hippocampus and cerebral cortex relate to TLE.Recently, the occurrence and application of proteomics provide us a powerful fool to find changes of proteins in tissues and cells and explore pathogenesis of diseases. The experimental subject of proteomics is proteins. Proteomics is a technology of studying pathogesis of diseases from the level of proteins and it has become the key point of life science research.This experiment is to study the differentially expressed proteins of rats hippo-campus and cerebral cortex in the lithium-pilocarpine model of TLE using compared proteomics. The 2D-DIGE spectrums of rats hippocampus and cerebral cortex with TLE were established. The expressing changes of proteins in hippocampus and cerebral cortex were analysed. The differentially expressed proteins were identified finally. The results may offer a clue to explore pathogenesis of TLE and seek for new therapeutic targets.Experimental methods:1. Set up the rats model of TLE,14 grown-up healthy male Wistar rats, provided by Shandong University Laboratory Animal Center, weight:200～250g. Rats were randomly divided into epileptic group (n=8) and control group (n=6). The epileptic rats were injected with lithium chloride (127mg/kg, intraperitoneally (i.p.)) 24h before pilocarpine treatment (30mg/kg, i.p.). Methylscopolamine (1mg/kg, i.p.) was injected 30min before administration of pilocarpine to counteract the peripheral cholinergic effects. Diazepam (10mg/kg, i.p.) was injected 90min after the onset of status epi-lepticus to terminate the seizure.Control rats were injected with the same volume of saline instead of pilocarpine.2. The hippocampus and cerebral cortex of rats were separated using 2D-DIGE. The gels were scanned on the Typhoon multifuctional laser scanning imaging system and obtained pictures through ImageMaster software. Then the differentially express-ed spots of proteins were screened out.3. The differential spots were analysed through MALDI-TOF-MS and PMFs were obtained. At last, the differential proteins were identified by researching protein databases with Mascot software.Results:1. The rats model of TLE:the seizure degrees of rats were judged by Racine grading standard of epileptic seizure. The epileptic rats achieving 3～5 degree were successful induced.They showed head bobbing, facial muscle twitching, limbs clonus and falling.Two rats of Epileptic group were dead and rejected in the experimental group. There was no death in control group.2. There were 48 differentially expressed spots in rats hippocampus and 19 proteins were finally identified through researching the protein databases.13 proteins were up-regulated and 6 proteins were down-regulated in the hippocampus. There were 43 differentially expressed spots in rats cerebral cortex and 15 proteins were finally identified through researching the protein databases.8 proteins were up-regulated and 7 proteins were down-regulated in the cerebral cortex.3. The proteins that were identified in the experiment related to nervous injury, remodeling of cytoskeleton, cellular metabolism, oxidative stress, etc. It can infer that these proteins play an important role in the pathogenesis of TLE.Conclusion:The 2D-DIGE sprctrums of rats hippocampus and cerebral cortex with TLE were successfully established. The differential spots of proteins were found and identified through compare and analysis.They may be connected with pathogenesis of TLE.