Dissertation > Medicine, health > Neurology and psychiatry > Neurology > Brain diseases > Epilepsy

The Effects of Chloroquine on Cell Cycle Expression of GFAP in the Brain of Rat with Seizures Induced by Pentylenetrazole

Author LiNingNing
Tutor WuShuHua
School Binzhou Medical College,
Course Pathology and Pathophysiology
Keywords Astrocytes Proliferation cycle GFAP c-Fos CyclinD1 Pentylenetetrazol Chloroquine
CLC R742.1
Type Master's thesis
Year 2010
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Objective: To observe the the different interventions dose of chloroquine pentylenetetrazol (Pentylenetrazole PTZ) induced chronic epileptic rat brain astrocytes (astrocyte, Ast) proliferation cycle, glial fibrillary acidic protein (GFAP), c-Fos and cycle expression of hormone D1 (CyclinD1). Study chronic epileptic state chloroquine intervention rat brain Ast proliferation, activation status, and to explore the mechanism of action and the best use of chloroquine antiepileptic dose. Methods: The experimental production of an experimental animal models, grouping, behavioral observations, and cell cycle analysis of 60 healthy male SD rats were randomly divided into a control group (10 and 40mg/kg injection of saline for 4 weeks), 50 with to the production of chronic epilepsy animal model (40 mg / kg injection of PTZ). Divided into five according to the Racine score, where at least five times in a row above II hair of modeling success. Maintenance dose is given after successful modeling. 50 have successfully induced epileptic rats were randomly divided into epilepsy group (set to the the simulation intervention group, the injection of 40 mg / kg of normal saline) and chloroquine intervention group, the intervention group was further divided into the intervention group (injection of 20 mg / kg chloroquine) intervention group (injection of 30 mg / kg chloroquine), the intervention group (injection of 40 mg / kg chloroquine), the intervention group (chloroquine 50 mg / kg injection) four subgroups (n = 10). PTZ injection, respectively, to observe and record the behavior of six groups of rats and EEG changes within 2 hours after the chloroquine treatment. Intervention after two weeks of treatment, 3% chloral hydrate (1 ml/100g) anesthesia, rapid brains were removed, and the separation of the cerebral cortex and hippocampus. The DNA ploidy kit glial cell chromosome staining, flow cytometry cell cycle, the cell cycle was observed. The second experiment Immunohistochemical detection of GFAP, c-Fos, CyclinD1 expression press packet preparation of animal models, methods, ibid. Extract bilateral cerebral cortex and hippocampus, more than 4% paraformaldehyde, conventional tissue processing, producer. GFAP protein expression in different tissues using immunohistochemical methods to detect and observe each group of GFAP positive for Ast morphology, quantity, and GFAP expression level changes; synchronous detection of c-Fos, CyclinD1 protein expression in the Ast. Using HPLAS-1000 high-definition color pathological the graphic analysis systems for image analysis, each group a total of seven rats 14 slices, each slice under 10 × 20 magnification randomly selected 10 fields, counting the number of positive cells, whichever average; integrated optical density of positive cells was measured at the same time, take the mean value of (OD). Packet to extract the brain tissue of laboratory animals, step-by-step, CyclinD1 expression Westernblot detected. Results: 1, behavioral observation shows that most of the animals injected with 9 days (d) after the the grade Ⅱ attack, all animals injected 14d after all a grade Ⅱ attack. The control group had no epileptic seizures, chloroquine intervention group compared with the epilepsy group was no significant difference (P gt; 0.05); remaining three intervention groups compared with the epilepsy group there were significant differences (P lt; 0.05). Obvious 3 groups of adverse drug reactions, including intervention and the emergence of a rats died intervention four groups of three deaths. EEG (electroencephalogram EEG) records, the control group had no epileptiform discharges and epilepsy group showed frequent high amplitude sharp waves, spikes, spike and wave or spike slow wave complexes, the intervention group showed slow-wave and small spines wave, with increasing doses of intervention, epilepsy waves weakened. 3, flow cytometric detection, PTZ induced the epilepsy group cortex and hippocampus Ast proliferative activity the strongest S period the proportion of an average of 47%; proportion of the S phase of the control group average of 0.5%. The intervention group with chloroquine treatment dose increased gradually decreased S phase proportion, all indices tended to be normal. 4, immunohistochemistry epilepsy group than in the control group Ast quantity significantly increased (P lt; 0.001), and uneven distribution of the significant changes in cell morphology, cell volume increased significantly, the cell body protrusions thickening variable length. And intervention groups Ast Quantity in turn gradually reduce cell morphology has been restored, the difference between the group and the epilepsy group was significant (P lt; 0.01). GFAP expression in epileptogenic group, the hippocampus, and the intervention groups GFAP expression were decreased compared with the epilepsy group difference was statistically significant (P lt; 0.05); c-Fos in the the epileptogenic group expression strong intervention group comparing differences was significant, and with the intervention dose the increase differences more obvious (P lt; 0.05); of CyclinD1, induced epilepsy group was highly expressed, the intervention group comparison there is a significant difference (P lt; 0.05) . Various indicators show that the intervention increased dose, gradually weakening trend Ast number of activation state, the latter tends to be more stable level. 5, Westernblot CyclinD1 content in the most epileptogenic group, four intervention groups in descending order, each group with the control group, there were significant differences (P lt; 0.05). Conclusion: 1, 0.5% PTZ can successfully copy rat model of chronic epilepsy, status epilepticus Ast excessive proliferation and activation is closely related. Chloroquine chronic seizures induced epileptic rats state has a certain extent, and with the the intervention dose increase inhibition was first increased and then tends to a steady state. Chloroquine by inhibiting Ast proliferation and GFAP, C-Fos, CyclinD1 high expression, thereby inhibiting the proliferation and activation of chronic epilepsy rat brain Ast, and thus play a role in anti-epileptic. The chloroquine antiepileptic effect with intervention dose increase was enhanced when the The chloroquine treatment dose was increased to a certain value, adverse drug reactions enhancements, and intervention effects do not significantly increase, the the best intervention dose range is 30mg-40mg/kg. 2,3,4 conclusions of this experiment has not been reported

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