本文選題：衰老 + 線粒體��； 參考：《中國病理生理雜志》2013年07期

【摘要】：目的:利用自然衰老雄性SD大鼠和D-半乳糖(D-Gal)誘導(dǎo)的細(xì)胞衰老模型,研究在衰老細(xì)胞中線粒體順烏頭酸酶(mitochondrial aconitase,ACO2)活性的變化及其對能量合成的影響。方法:利用55 mmol/L D-Gal誘導(dǎo)人胚肺成纖維細(xì)胞MRC-5衰老,并采用蔗糖連續(xù)密度梯度離心法分離大鼠腦組織和MRC-5細(xì)胞的線粒體;ACO2活性根據(jù)反應(yīng)產(chǎn)物NADPH在340 nm的吸光度變化率進(jìn)行檢測,其表達(dá)水平分析分別用熒光定量PCR和Western blotting完成;利用試劑盒檢測細(xì)胞超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量以反映細(xì)胞氧化還原狀態(tài);組織鐵含量的檢測則通過菲咯嗪法進(jìn)行;線粒體膜電位及腺苷酸含量分別應(yīng)用JC-1熒光探針法和高效液相色譜法檢測,并計算能荷。結(jié)果:隨著大鼠月齡增加,ACO2活性逐漸下降,表達(dá)水平無明顯差異,組織鐵水平逐漸升高;體外氧化應(yīng)激實驗顯示ACO2活性隨H2O2處理濃度和時間增加而下降;細(xì)胞衰老模型中,SOD活性降低,MDA含量升高,ACO2活性下降而表達(dá)水平未變;自然衰老大鼠和細(xì)胞衰老模型中,線粒體膜電位、ATP合成與能荷呈明顯下降趨勢。結(jié)論:衰老過程中,隨著氧化壓力增加,氧化應(yīng)激敏感的ACO2活性下降,影響三羧酸循環(huán)效率,同時伴線粒體膜電位下降,進(jìn)而使得能量合成降低。
[Abstract]:Objective: To study the changes of mitochondrial CIS (mitochondrial aconitase, ACO2) activity in aging cells and the effect on the energy synthesis in aging SD rats and D- galactose (D-Gal). Methods: using 55 mmol/L D-Gal to induce the MRC-5 senescence of human embryonic lung fibroblasts and use sucrose. The mitochondria of rat brain tissue and MRC-5 cells were separated by continuous density gradient centrifugation; ACO2 activity was detected by the reaction product NADPH at 340 nm absorbance rate, and the expression level analysis was performed by fluorescence quantitative PCR and Western blotting, respectively. The activity of cell superoxide dismutase (SOD) and malondialdehyde (MDA) were detected by the reagent box. The content of the cells was redox state, and the content of the tissue was detected by the phenanthrazine method. The mitochondrial membrane potential and adenylate content were detected by JC-1 fluorescence probe and high performance liquid chromatography, and the energy charge was calculated. The results showed that with the increase of the age of the rats, the activity of ACO2 gradually decreased and the expression level was not significantly different. In vitro oxidative stress experiments showed that the activity of ACO2 decreased with the increase of concentration and time of H2O2 treatment. In the cell aging model, the activity of SOD decreased, the content of MDA increased, the activity of ACO2 decreased and the expression level was not changed. In the natural aging rats and cell aging models, the grain potential of the grain body, the synthesis of ATP and the energy charge were obviously decreased. Conclusion: during the aging process, the oxidative stress sensitive ACO2 activity decreased with the increase of oxidative stress, which affected the efficiency of the three carboxylic acid cycle, while the mitochondrial membrane potential decreased, which reduced the energy synthesis.
【作者單位】： 溫州醫(yī)學(xué)院檢驗醫(yī)學(xué)與生命科學(xué)學(xué)院;溫州醫(yī)學(xué)院環(huán)境與公共衛(wèi)生學(xué)院;
【基金】：國家973計劃項目(No.2007CB507402) 浙江省自然科學(xué)基金資助項目(No.Y2100582) 浙江省教育廳科研基金資助項目(No.Y200804470)
【分類號】：R363

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9 楊朝;，

本文編號：1929543