【摘要】：研究背景：伴隨世界老齡化進(jìn)程的加劇，具有年齡依賴性的阿爾茨海默病已成為21世紀(jì)威脅人類健康的重要疾病之一，因其發(fā)病機制復(fù)雜，發(fā)病率、死亡率高，尚無有效治療措施，其相關(guān)領(lǐng)域成為了科學(xué)界的研究熱點。近年來諸多研究表明蛋白聚集與神經(jīng)元的損傷及死亡之間有著密切的聯(lián)系；細(xì)胞凋亡與阿爾茨海默病密切相關(guān)，但其機制尚未十分清楚，可能與線粒體損傷、氧化應(yīng)激、鈣穩(wěn)態(tài)失衡等因素相關(guān)；同時，研究者發(fā)現(xiàn)內(nèi)質(zhì)網(wǎng)對于機體內(nèi)環(huán)境的改變比較敏感，當(dāng)細(xì)胞受到某些內(nèi)外因素的刺激時，細(xì)胞啟動內(nèi)質(zhì)網(wǎng)應(yīng)激機制，但長時間、持續(xù)的內(nèi)質(zhì)網(wǎng)應(yīng)激將激活細(xì)胞死亡級聯(lián)反應(yīng)，進(jìn)而參與阿爾茨海默病的發(fā)病機制和病理過程。臨床和動物學(xué)研究表明，全身麻醉能夠加重阿爾茨海默病的癥狀和病理過程，，降低其發(fā)病年齡，增加其發(fā)病率。相關(guān)文獻(xiàn)報道，臨床常用的吸入麻醉藥異氟醚可能通過增加腦內(nèi)異常蛋白的產(chǎn)生和聚集，激活凋亡級聯(lián)信號通路，誘導(dǎo)神經(jīng)細(xì)胞凋亡，介導(dǎo)阿爾茨海默病的發(fā)生。目前全世界每年有2.3億人在接受麻醉和手術(shù)，其中接受麻醉的阿爾茨海默病患者達(dá)800萬以上，存在巨大的麻醉安全風(fēng)險和隱患。因此，探究吸入麻醉藥異氟醚、阿爾茨海默病與細(xì)胞凋亡之間的內(nèi)在聯(lián)系具有重要的臨床和社會意義。 目的：探討吸入麻醉藥異氟醚對β淀粉樣蛋白（A）25-35誘導(dǎo)的大鼠PC12細(xì)胞凋亡和內(nèi)質(zhì)網(wǎng)應(yīng)激的影響，并闡明其機制。 方法：將PC12細(xì)胞隨機分為4組：正常對照組（C組）、A25-3510μmol/L處理組（A組）、2%異氟醚處理組（Iso組）和2%異氟醚和A25-3510μmol/L處理組（Iso+A組）。各組PC12細(xì)胞藥物處理24h后應(yīng)用四甲基偶氮唑鹽（MTT）比色法檢測細(xì)胞活性，Hoechst33342核染色法檢測PC12細(xì)胞凋亡形態(tài)，Western blot檢測內(nèi)質(zhì)網(wǎng)分子伴侶蛋白GRP78及其應(yīng)激相關(guān)凋亡信號蛋白CHOP、JNK和caspase-12表達(dá)。 結(jié)果：與C組比較，A組和Iso組PC12細(xì)胞存活率明顯降低（P 0.05），細(xì)胞凋亡率明顯增加（P 0.05），內(nèi)質(zhì)網(wǎng)分子伴侶蛋白GRP78表達(dá)明顯上調(diào)（P 0.05），內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)凋亡信號蛋白CHOP、p-JNK和caspase-12表達(dá)均明顯增強（P 0.05）；與A組比較，Iso+A組PC12細(xì)胞存活率明顯降低（P 0.05），細(xì)胞凋亡率明顯增加（P 0.05），內(nèi)質(zhì)網(wǎng)分子伴侶蛋白GRP78和內(nèi)質(zhì)網(wǎng)應(yīng)激相關(guān)凋亡信號蛋白CHOP、p-JNK和caspase-12表達(dá)均明顯增強（P 0.05）。 結(jié)論：異氟醚能夠促進(jìn)Aβ25-35誘導(dǎo)的大鼠PC12細(xì)胞凋亡，其機制與激活內(nèi)質(zhì)網(wǎng)應(yīng)激及其相關(guān)的凋亡信號通路有關(guān)。
[Abstract]:Background: with the aggravation of the aging process in the world, Alzheimer's disease with age dependence has become one of the most important diseases threatening human health in the 21st century, because of its complex pathogenesis, high morbidity and high mortality. There is no effective treatment, and its related field has become a hot spot in the scientific community. In recent years, many studies have shown that there is a close relationship between protein aggregation and neuronal injury and death. Apoptosis is closely related to Alzheimer's disease, but its mechanism is not very clear, which may be related to mitochondrial damage, oxidative stress, calcium homeostasis and other factors. At the same time, the researchers found that endoplasmic reticulum is more sensitive to changes in the body's internal environment. When cells are stimulated by some internal and external factors, the cells activate the endoplasmic reticulum stress mechanism, but for a long time. Persistent endoplasmic reticulum stress activates the cascade of cell death, and then participates in the pathogenesis and pathological process of Alzheimer's disease. Clinical and zoological studies have shown that general anesthesia can aggravate the symptoms and pathological process of Alzheimer's disease, reduce the age of onset and increase the incidence of Alzheimer's disease. It has been reported that inhaled anesthetic isoflurane may activate apoptotic cascade signaling pathway by increasing the production and aggregation of abnormal proteins in the brain, induce neuronal apoptosis and mediate the occurrence of Alzheimer's disease. At present, 230 million people in the world are undergoing anesthesia and surgery every year. Among them, more than 8 million patients with Alzheimer's disease are anesthetized, and there are huge risks and hidden dangers in anesthetic safety. Therefore, it is of great clinical and social significance to explore the relationship between inhaled anesthetic isoflurane, Alzheimer's disease and apoptosis. Aim: to investigate the effects of inhaled anesthetic isoflurane on apoptosis and endoplasmic reticulum stress of rat PC12 cells induced by 尾 -amyloid (A) 25-35, and to elucidate its mechanism. Methods: PC12 cells were randomly divided into 4 groups: normal control group (group C), A25-3510 渭 mol/L group (group A), 2% isoflurane treatment group (Iso group) and 2% isoflurane and A25-3510 渭 mol/L treatment group (Iso A group). The activity of PC12 cells was detected by tetramethylazolium (MTT) colorimetry 24 hours after drug treatment, and the apoptotic morphology of PC12 cells was detected by Hoechst33342 nuclear staining, and the endoplasmic reticulum molecular chaperone protein GRP78 and its stress-related apoptotic signal protein CHOP, were detected by Hoechst33342 nuclear staining. JNK and caspase-12 expression. Results: compared with group C, the survival rate of PC12 cells in group A and Iso was significantly lower than that in group C (P 0.05), the apoptosis rate was significantly increased (P 0.05), and the expression of ER molecular chaperone GRP78 was up-regulated (P 0.05). Endoplasmic reticulum stress-related apoptotic signal proteins (CHOP,p-JNK and caspase-12) were significantly increased (P 0.05). Compared with group A, the survival rate of PC12 cells in, Iso A group was significantly lower than that in group A (P 0.05), and the apoptosis rate was significantly increased (P 0.05). Endoplasmic reticulum molecular chaperone protein (GRP78) and endoplasmic reticulum stress-related apoptotic signal protein (CHOP,) were also significantly increased in, Iso A group. The expression of p-JNK and caspase-12 were significantly increased (P0. 05). Conclusion: isoflurane can promote apoptosis of rat PC12 cells induced by A 尾 25-35, and its mechanism is related to activation of endoplasmic reticulum stress and its related apoptosis signaling pathway.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R614

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