本文選題：癲癇　切入點：托吡酯　出處：《山東醫(yī)藥》2017年27期 　論文類型：期刊論文

【摘要】：目的觀察托吡酯(TPM)腹腔注射后癲癇大鼠海馬組織損傷改變,并探討其可能機制。方法將40只大鼠依據(jù)隨機數(shù)字表法分為正常組、模型組、TPM低劑量組、TPM高劑量組。模型組、TPM低劑量組、TPM高劑量組采用腹腔注射氯化鋰180 mg/kg、匹羅卡品30 mg/kg制作癲癇模型。模型組、TPM低劑量組、TPM高劑量組大鼠分別于癲癇發(fā)作后5 h經(jīng)腹腔注射生理鹽水10 m L/(kg·d)、TPM 40 mg/(kg·d)、TPM 80 mg/(kg·d)。各組連續(xù)用藥4周后處死大鼠取海馬組織,HE染色觀察大鼠海馬組織病理變化;應(yīng)用基因芯片技術(shù)篩選各組差異表達的miRNA,并用實時熒光定量PCR法進行驗證;采用TUNEL法檢測海馬組織凋亡細胞,并計算凋亡細胞數(shù);分別采用Western blotting法、免疫熒光法和免疫組化法檢測各組大鼠海馬組織中的Caspase-8。結(jié)果與正常組相比,模型組海馬組織損傷明顯,TPM低劑量組和TPM高劑量組海馬組織損傷較模型組減輕,且TPM低劑量組損傷最輕�；蛐酒瑱z測發(fā)現(xiàn)模型組出現(xiàn)12個表達上調(diào)的miRNA和14個表達下調(diào)的miRNA,經(jīng)實時熒光定量PCR驗證,只有miR-146a在各組表達的差異有統(tǒng)計學(xué)意義。模型組、TPM高劑量組、TPM低劑量組、正常組海馬組織中miR-146a相對表達量依次降低(P均0.05)。細胞凋亡檢測發(fā)現(xiàn),與正常組相比,模型組大鼠CA1區(qū)、CA3區(qū)和DG區(qū)凋亡細胞細胞數(shù)高于正常組(P均0.05),而TPM高劑量組和TPM低劑量組凋亡細胞數(shù)低于模型組(P均0.05)。Western blotting結(jié)果顯示,模型組Caspase-8蛋白相對表達量高于正常組,模型組、TPM高劑量組、TPM低劑量組Caspase-8蛋白相對表達量依次降低(P均0.01)。免疫熒光結(jié)果顯示,與正常組相比,模型組大鼠CA1區(qū)、CA3區(qū)和DG區(qū)Caspase-8陽性細胞數(shù)高于正常組(P均0.05),TPM高劑量組和TPM低劑量組Caspase-8陽性細胞數(shù)低于模型組(P均0.05)。免疫組化結(jié)果顯示,模型組IOD值高于正常組,模型組、TPM高劑量組、TPM低劑量組IOD值依次降低(P均0.05)。結(jié)論 TPM腹腔注射可減輕癲癇大鼠海馬組織病理改變,減少海馬組織細胞凋亡,下調(diào)海馬組織中miR-146a和Caspase-8的表達,其中40 mg/(kg·d)的TPM較80 mg/(kg·d)效果更加明顯。TPM的抗癲癇作用機制可能與調(diào)節(jié)miRNA表達、減少細胞凋亡、下調(diào)Caspase-8表達有關(guān)。
[Abstract]:Objective to observe the changes of hippocampal injury in epileptic rats after intraperitoneal injection of topiramate (TPM) and to explore its possible mechanism. Model group TPM low dose group TPM high dose group, model group TPM low dose group TPM high dose group by intraperitoneal injection of lithium chloride 180 mg / kg, pilocarpine 30 mg/kg to make epilepsy model. Five hours after seizure, the rats were injected intraperitoneally with normal saline (10 mL / kg 路d ~ (-1)) for 40 mg/(kg 路d ~ (-1) TPM 80 mg/(kg 路d ~ (-1). After 4 weeks of continuous administration, the rats were sacrificed to observe the pathological changes of hippocampus tissue by HE staining. The differentially expressed miRNAs were screened by gene chip technique and verified by real-time fluorescence quantitative PCR. The apoptotic cells in hippocampus were detected by TUNEL method and the number of apoptotic cells was calculated. Western blotting method was used respectively. Caspase-8 in hippocampal tissue of rats in each group was detected by immunofluorescence and immunohistochemistry. Results compared with the normal group, the hippocampal tissue damage in the model group was significantly reduced compared with the model group in the low dose group and the high dose group of TPM. Gene chip analysis showed that 12 up-regulated miRNA and 14 down-regulated miRNAs were found in the model group, which were verified by real-time fluorescence quantitative PCR. Only the difference of miR-146a expression in each group was statistically significant. The relative expression of miR-146a in hippocampal tissues of the model group and the normal group decreased in turn, and the relative expression of miR-146a in the hippocampus of the normal group was decreased in turn. The results of apoptosis detection showed that compared with the normal group, the relative expression of miR-146a in the model group was significantly lower than that in the control group. The number of apoptotic cells in the CA1 CA3 and DG regions of the model group was higher than that in the normal group (P 0.05), while the number of apoptotic cells in the high dose TPM group and the low dose TPM group was lower than that in the model group. The results showed that the relative expression of Caspase-8 protein in the model group was higher than that in the normal group. The relative expression of Caspase-8 protein decreased in turn in the model group with high dose of TPM and low dose group (P < 0.01). The immunofluorescence results showed that, compared with the normal group, the relative expression of Caspase-8 protein in the model group was significantly lower than that in the normal group. The number of Caspase-8 positive cells in the CA1 CA3 and DG regions in the model group was higher than that in the normal group (P 0.05) and the number of Caspase-8 positive cells in the low-dose TPM group was lower than that in the model group (P 0.05). The immunohistochemical results showed that the IOD value in the model group was higher than that in the normal group. Conclusion Intraperitoneal injection of TPM can attenuate pathological changes of hippocampus, decrease apoptosis of hippocampal cells and down-regulate the expression of miR-146a and Caspase-8 in hippocampus of epileptic rats. The antiepileptic mechanism of 40 mg/(kg 路d TPM is more obvious than that of 80 mg/(kg 路d). The mechanism may be related to the regulation of miRNA expression, the reduction of apoptosis and the down-regulation of Caspase-8 expression.
【作者單位】： 延邊大學(xué)附屬醫(yī)院;延邊大學(xué);
【基金】：吉林省醫(yī)藥衛(wèi)生科研計劃項目(C2017103)
【分類號】：R742.1

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