本文選題：錳 + PC12細(xì)胞�。� 參考：《遵義醫(yī)學(xué)院》2013年碩士論文

【摘要】：目的：研究錳在不同劑量和不同時(shí)間下對鼠嗜鉻神經(jīng)瘤細(xì)胞(PC12)功能和PARK2(Parkin基因)表達(dá)的影響,并進(jìn)一步探討錳致PARK2表達(dá)與PC12細(xì)胞功能損害的關(guān)系。 方法：鼠嗜鉻神經(jīng)瘤細(xì)胞(PC12)中分別加入①0、100、300、500μmol/L濃度的Mnc12分別染毒24h；②500μmol/L Mncl2染毒6、24、48h。采用MTT法檢測線粒體功能損傷；化學(xué)比色法測定細(xì)胞內(nèi)丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性和谷胱甘肽過氧化物酶(GSH-PX)活性；反相高效液相(RP-HPLC)-熒光法測定細(xì)胞內(nèi)多巴胺(DA)含量；實(shí)時(shí)熒光定量PCR (real time quantitative PCR)檢測PARK2mRNA水平；蛋白免疫印跡法(Western blot)檢測Parkin蛋白含量。 結(jié)果-MTT實(shí)驗(yàn)結(jié)果顯示,錳可誘導(dǎo)PC12細(xì)胞線粒體損傷,與對照組相比,300-500μmol/L MnCl2作用6、24、48h對線粒體有明顯損傷(P0.01)。比色法結(jié)果顯示：隨著染錳濃度的增高,MDA的含量逐漸升高,SOD、GSH-PX活性逐漸降低。與對照組相比,300-500μmol/L MnCl2作用24h, PC12細(xì)胞內(nèi)MDA、SOD及GSH-PX水平差異有顯著性(P0.01);500μmol/L MnCl2作用6、24、48h,MDA含量、SOD及GSH-PX活性差異有顯著性(P0.01)。RP-HPLC-熒光法檢測結(jié)果顯示：隨著染錳濃度的增高,多巴胺含量呈下降趨勢,與對照組比較,300μmol/L或以上錳濃度所致多巴胺含量降低有統(tǒng)計(jì)學(xué)意義(P0.01)。500μmol/L錳暴露6h,24h,48h,多巴胺含量明顯降低(P0.01)。實(shí)時(shí)熒光定量PCR及Western blot結(jié)果顯示：與對照組比較,300μmol/L或以上錳濃度作用下,PARK2mRNA及蛋白表達(dá)下調(diào)(P0.05)。500μmol/L錳暴露6h,24h,48h, PARK2mRNA及蛋白表達(dá)下調(diào)(P0.05)。 結(jié)論：錳可誘導(dǎo)PC12細(xì)胞功能損害,該作用過程可能與錳致PARK2的表達(dá)下調(diào)有關(guān)。
[Abstract]:Aim: to study the effects of manganese on the function of rat pheochromocytoma cell line (PC12) and the expression of PARK2(Parkin gene at different dose and time, and to explore the relationship between the expression of PARK2 and the damage of PC12 cell function induced by manganese. Methods: rat pheochromocytoma cells (PC12) were exposed to 10100300500 渭 mol/L Mnc12 for 24 h and 2 500 渭 mol/L Mncl2 for 48 h. The contents of malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX) in the cells were determined by MTT assay, and the contents of dopamine butadiene (DAA) in the cells by RP-HPLCI-fluorescence method were measured by reversed-phase high performance liquid phase (RP-HPLCI-fluorescence) method, and the activity of glutathione peroxidase (GSH-PX) was determined by chemical colorimetry. The level of PARK2mRNA was detected by real-time fluorescence quantitative PCR real time quantitative PCR, and the content of Parkin protein was detected by Western blotting. Results the results of -MTT assay showed that manganese could induce mitochondrial damage in PC12 cells. Compared with the control group, 300-500 渭 mol/L MnCl2 could significantly damage the mitochondria for 48 h. The results of colorimetry showed that the activity of GSH-PX decreased with the increase of manganese concentration. Compared with the control group treated with 300-500 渭 mol/L MnCl2 for 24 h, the levels of MDA-SOD and GSH-PX in PC12 cells were significantly higher than those in the control group. There was a significant difference in the activity of MDA-SOD and GSH-PX in PC12 cells treated with P0.01-500 渭 mol/L MnCl2 for 48h. The results of P0.01- RP-HPLC- fluorescence assay showed that the content of dopamine decreased with the increase of mn concentration. Compared with the control group, the dopamine content was significantly decreased by manganese concentration of 300 渭 mol/L or above. The content of dopamine was significantly decreased after exposure to manganese for 6 h, 24 h and 48 h respectively, and the content of dopamine was significantly lower than that of the control group (P 0. 01%, P 0. 01%, P 0. 01, P 0. 01). The results of real-time fluorescence quantitative PCR and Western blot showed that the expression of PARK2 mRNA and protein was down-regulated at the concentration of 300 渭 mol/L or more than that of the control group. The expression of PARK2 mRNA and protein was down-regulated (P 0.05), and the expression of PARK2mRNA and protein was down-regulated (P 0.05). Conclusion: manganese can induce the damage of PC12 cell function, which may be related to the down-regulation of PARK2 expression induced by mn.
【學(xué)位授予單位】：遵義醫(yī)學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R114

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