本文選題：神經(jīng)元損傷　切入點：鋁鹽　出處：《重慶醫(yī)科大學(xué)》2010年碩士論文

【摘要】： 目的:建立氯化鋁致原代培養(yǎng)大鼠海馬神經(jīng)元損傷模型,探討5-LO的表達與鋁鹽致海馬神經(jīng)元損傷的關(guān)系。 方法: (1)新生24h內(nèi)SD大鼠海馬神經(jīng)元原代培養(yǎng),7天后用免疫組化鑒定純度;給予氯化鋁(Al3+ 200μmol·L-1)建立原代培養(yǎng)大鼠海馬神經(jīng)元損傷模型。 (2)以HE染色及熒光顯微鏡觀察海馬神經(jīng)元病理形態(tài)變化,采用生化方法檢測MTT值、LDH漏出率、SOD活性和MDA含量,分別用RT-PCR和Western Blotting方法檢測5-LO mRNA和蛋白表達。 (3)5-LO過表達腺病毒轉(zhuǎn)染原代培養(yǎng)海馬神經(jīng)元,觀察5-LO過表達神經(jīng)元的變化,以及5-LO過表達對鋁鹽致海馬神經(jīng)元損傷的易感性。 (4)用咖啡酸抑制5-LO活性,RNA干擾抑制5-LO表達,觀察其對鋁鹽致海馬神經(jīng)元損傷的作用。 結(jié)果: (1)神經(jīng)元純度超過95%。鋁負荷致原代培養(yǎng)海馬神經(jīng)元數(shù)目明顯減少,神經(jīng)元突起減少變短,活力降低,LDH漏出率明顯增加,SOD活性降低,MDA含量升高,5-LO mRNA和蛋白表達增加。 (2)與空載腺病毒轉(zhuǎn)染組相比,單純5-LO過表達神經(jīng)元的數(shù)目、形態(tài),細胞活力,LDH漏出率,SOD活性,MDA含量無明顯變化,5-LO mRNA和5-LO蛋白表達增加。與鋁鹽加空載腺病毒轉(zhuǎn)染組相比,5-LO過表達神經(jīng)元給予鋁鹽負荷,神經(jīng)元數(shù)目減少更為明顯,細胞碎片增加,細胞活力和SOD活性顯著降低,LDH漏出率和MDA含量顯著增加,5-LO mRNA和5-LO蛋白表達也明顯增加。 (3)咖啡酸給予能劑量依賴性地減輕鋁負荷/和5-LO腺病毒過表達海馬神經(jīng)元損傷,提高神經(jīng)元存活力,降低LDH漏出率,明顯阻遏鋁負荷神經(jīng)元SOD活性的降低和MDA含量的升高,能下調(diào)5-LO mRNA和蛋白的表達。 (4)5-LO RNA干擾能明顯減輕鋁負荷所致的海馬神經(jīng)元損傷,提高神經(jīng)元的存活力,降低LDH的漏出率,明顯阻遏鋁負荷神經(jīng)元SOD活性的降低和MDA含量的升高,明顯降低5-LO mRNA和蛋白的表達。 結(jié)論: (1)神經(jīng)元單純過表達5-LO并不引起明顯的細胞損傷,但能增加神經(jīng)元對鋁負荷損傷的易感性。 (2)抑制5-LO活性、下調(diào)5-LO表達對鋁負荷致海馬神經(jīng)元損傷有明顯的保護作用。
[Abstract]:Aim: to establish the model of primary cultured rat hippocampal neuron injury induced by aluminum chloride, and to investigate the relationship between the expression of 5-LO and the damage of hippocampal neurons induced by aluminum salt.Methods:(1) the primary cultured hippocampal neurons of SD rats within 24 hours were cultured for 7 days, the purity of hippocampal neurons was identified by immunohistochemistry, and the injury model of hippocampal neurons in primary cultured rats was established by Al 3 200 渭 mol L -1).(2) the histopathological changes of hippocampal neurons were observed by HE staining and fluorescence microscope. Sod activity and MDA content were detected by biochemical method and 5-LO mRNA and protein expression were detected by RT-PCR and Western Blotting respectively.The changes of 5-LO overexpression neurons and the susceptibility of 5-LO overexpression to the damage of hippocampal neurons induced by aluminum salt were observed in primary cultured hippocampal neurons transfected with adenovirus.(4) the inhibitory effect of caffeic acid on the expression of 5-LO in hippocampal neurons induced by aluminum salt was observed.Results:The purity of neurons was more than 95%.The number of primary cultured hippocampal neurons was significantly decreased, the neuronal process was decreased, the leakage rate of LDH was decreased, the activity of SOD was increased, the content of malondialdehyde (MDA) was increased and the expression of 5-LO mRNA and protein was increased in primary cultured hippocampal neurons.(2) compared with the control group, the number and morphology of 5-LO overexpression neurons, the leakage rate of 5-LO activity and the content of 5-LO protein were not significantly increased in the control group compared with those in the non-loaded adenovirus transfection group, and the expression of 5-LO mRNA and 5-LO protein were not significantly increased.Compared with the aluminum-salt + no-load adenovirus transfection group, the number of neurons decreased and the cell fragments increased when the overexpression neurons of 5-LO were loaded with aluminum salt.Cell viability and SOD activity significantly decreased the leakage rate and the content of MDA significantly increased the expression of 5-LO mRNA and 5-LO protein.(3) caffeic acid could attenuate the damage of hippocampal neurons under aluminum load / 5-LO overexpression in a dose-dependent manner, increase the viability of neurons, decrease the leakage rate of LDH, and significantly inhibit the decrease of SOD activity and the increase of MDA content in aluminum-loaded neurons.It can down-regulate the expression of 5-LO mRNA and protein.The interference of 5-LO RNA could significantly reduce the damage of hippocampal neurons induced by aluminum load, increase the viability of neurons, decrease the leakage rate of LDH, and inhibit the decrease of SOD activity and the increase of MDA content in the neurons under aluminum loading.The expression of 5-LO mRNA and protein was significantly decreased.Conclusion:1) single overexpression of 5-LO in neurons did not cause obvious cell damage, but increased the susceptibility of neurons to aluminum load damage.2) inhibition of 5-LO activity and down-regulation of 5-LO expression have obvious protective effects on hippocampal neuron injury induced by aluminum load.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2010
【分類號】：R363

【參考文獻】

相關(guān)期刊論文 前5條

1 田雪菲,蒲小平;咖啡酸對神經(jīng)毒素MPP~+誘發(fā)大鼠小腦顆粒細胞凋亡的保護作用[J];北京大學(xué)學(xué)報(醫(yī)學(xué)版);2004年01期

2 江千里,王健民,溫麗敏,江汕,周虹;批量快速測定法測定標(biāo)志基因為GFP的重組病毒滴度[J];第二軍醫(yī)大學(xué)學(xué)報;2002年09期

3 張鵬;楊俊卿;蘇強;;咖啡酸對慢性鋁過負荷致大鼠腦損傷的保護作用[J];中國藥理學(xué)通報;2009年02期

4 徐峰,宋丹青,甄永蘇;咖啡酸鈉與絲裂霉素抗腫瘤的協(xié)同作用[J];藥學(xué)學(xué)報;2002年06期

5 ;Caffeic acid ameliorates early and delayed brain injuries after focal cerebral ischemia in rats[J];Acta Pharmacologica Sinica;2006年09期

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本文編號：1714680