【摘要】：心搏驟停(cardiac arrest,CA)是世界范圍內(nèi)的主要致死原因,能夠造成全腦嚴(yán)重的缺氧缺血性損害。雖然心肺復(fù)蘇(cardio-pulmonary resuscitation,CPR)技術(shù)的提高和急救醫(yī)療體系的完善能挽回部分患者的自主呼吸和循環(huán),提高最初的復(fù)蘇成功率,但后期的存活率及生存質(zhì)量卻令人沮喪。研究表明,僅不足50%的患者在心搏驟停后得以長期生存,而其中大多患者會遺留有不同程度的認(rèn)知和行為功能障礙。盡管人們進(jìn)行了大量的基礎(chǔ)研究和臨床實(shí)踐,迄今為止仍沒有找到能夠有效減少復(fù)蘇后腦損傷的理想腦保護(hù)藥物,這已經(jīng)成為一個亟待解決的臨床醫(yī)學(xué)難題。近年來,有研究表明七氟烷后處理能夠顯著改善局灶性腦損傷的神經(jīng)缺損,而對心肺復(fù)蘇后的全腦損傷的保護(hù)作用卻鮮有報道。磷脂酰肌醇3-激酶/蛋白激酶B(phosphatidyl inositol 3-kinase/protein kinase B,PI3K/Akt)通路為體內(nèi)最為重要的信號通路之一,參與細(xì)胞的生長、存活、代謝及凋亡,在藥物處理的動物局灶性腦缺血損傷或全腦缺血損傷模型中發(fā)揮了重要的作用。糖原合成激酶-3β(glycogen synthase kinase-3β,GSK-3β)是PI3K/Akt通路的重要下游因子之一,它除了廣泛參與體內(nèi)細(xì)胞的生長和發(fā)育,還兼有調(diào)節(jié)血糖和促進(jìn)腫瘤的形成的作用。研究表明,GSK-3β不僅在四血管栓塞所致的全腦損傷中具有保護(hù)作用,而且在關(guān)于胰島素受體的大量研究中發(fā)現(xiàn),GSK-3β還可以通過多種機(jī)制來實(shí)現(xiàn)體內(nèi)糖代謝的調(diào)節(jié),涉及糖代謝的各個環(huán)節(jié),如影響胰島β細(xì)胞的功能、糖原合成等。因此,本實(shí)驗(yàn)建立大鼠窒息性心搏驟停模型,在復(fù)蘇即刻給大鼠吸入不同濃度七氟烷,通過觀察復(fù)蘇后大鼠血糖值、血清神經(jīng)元特異性烯醇化酶(neuron specific enolase,NSE)值、海馬凋亡蛋白P53表達(dá)情況、組織病理損傷情況以及大鼠整體認(rèn)知行為能力的變化,研究七氟烷后處理對窒息性心搏驟停后腦損傷的保護(hù)作用;在上述實(shí)驗(yàn)的基礎(chǔ)上,靜脈給予PI3K特異性拮抗劑,評價其對七氟烷后處理神經(jīng)保護(hù)作用的影響,揭示PI3K/Akt通路在七氟烷后處理腦保護(hù)中的作用,探究其與抗凋亡和糖代謝調(diào)節(jié)之間的關(guān)系,為心肺復(fù)蘇腦損傷的救治提供新的思路和方法。實(shí)驗(yàn)一:不同濃度七氟烷后處理對窒息性心搏驟停大鼠腦損傷的保護(hù)作用目的研究表明,在局灶性腦損傷模型中,七氟烷后處理能夠顯著改善神經(jīng)缺損,而對心肺復(fù)蘇后的全腦損傷的保護(hù)作用研究卻較少。本實(shí)驗(yàn)旨在評價不同濃度七氟烷后處理對大鼠心搏驟停后全腦缺血損傷的保護(hù)作用。方法將80只成年雄性SD大鼠隨機(jī)分為五組:假手術(shù)組(sham組)、對照組(control組)、0.5倍最低肺泡有效濃度(minimum alveolar concentration,MAC)、1.0 MAC和1.5 MAC七氟烷后處理組(0.5SP組、1.0SP組及1.5SP組)。Control組、0.5SP組、1.0SP組及1.5SP組建立8min窒息性心搏驟停模型,后給予心肺復(fù)蘇搶救;各七氟烷后處理組在復(fù)蘇即刻給予間斷吸入七氟烷2次,每次5min,間隔10min,使用氣體分析儀確保濃度分別為1.3%、2.5%和3.8%,而control組大鼠僅吸入100%氧氣。記錄各組大鼠復(fù)蘇成功率、復(fù)蘇后72h血糖值、血清神經(jīng)元特異性烯醇化酶(NSE)含量、海馬CA1區(qū)神經(jīng)元形態(tài)學(xué)變化、海馬區(qū)P53的表達(dá)、復(fù)蘇后24、72h和7d的神經(jīng)功能缺損評分(neurological deficit score,NDS),及復(fù)蘇后7d至11d的大鼠空間學(xué)習(xí)記憶能力。結(jié)果各處理組大鼠復(fù)蘇成功率沒有統(tǒng)計學(xué)差異,1.0SP組及1.5SP組在復(fù)蘇后72h的血糖值、NSE值明顯較control組低(P0.05),海馬CA1區(qū)有較多的存活神經(jīng)元、較少的凋亡蛋白P53表達(dá)(P0.05),神經(jīng)功能和空間學(xué)習(xí)記憶能力也較control組水平高(P0.05),而0.5SP組并未表現(xiàn)出明顯的神經(jīng)保護(hù)作用。結(jié)論較高濃度(1.0MAC和1.5MAC)的七氟烷后處理可以顯著改善大鼠窒息性心搏驟停所致的全腦損傷。實(shí)驗(yàn)二:七氟烷后處理減輕窒息性心搏驟停腦損傷過程中PI3K/Akt的作用研究目的PI3K/Akt通路是體內(nèi)最為重要的信號通路之一,參與細(xì)胞的生長、存活、代謝及凋亡。而其下游因子GSK-3β可以從糖原合成等多方面參與機(jī)體糖代謝調(diào)節(jié)過程,本文旨在探索七氟烷的腦保護(hù)作用是否與PI3K/Akt通路介導(dǎo)的抗凋亡及GSK-3β參與的糖代謝調(diào)節(jié)作用有關(guān)。方法80只健康雄性SD大鼠隨機(jī)均分為五組:假手術(shù)組(sham組)、對照組(control組)、1.0MAC七氟烷后處理組(SP組)、PI3K特異性抑制劑Wortmannin+七氟烷后處理組(W+SP組)和單純Wortmannin組(W組)。Sham組、control組和SP組具體處理方法同實(shí)驗(yàn)一;W+SP組于窒息前30min經(jīng)股靜脈給予Wortmannin,并按實(shí)驗(yàn)一方法建立心搏驟停模型,進(jìn)行心肺復(fù)蘇,吸入七氟烷并確保呼氣末濃度為2.5%;而W組僅于窒息前30min經(jīng)股靜脈給予Wortmannin、并建立心搏驟停模型。記錄各組大鼠復(fù)蘇成功率、復(fù)蘇后72h血糖值、海馬CA1區(qū)神經(jīng)元形態(tài)學(xué)變化,并觀察海馬區(qū)凋亡相關(guān)蛋白Bcl-2、Bax,通路相關(guān)蛋白p Akt/Akt、pGSK-3β/GSK-3β等的表達(dá)。結(jié)果各組大鼠復(fù)蘇成功率沒有統(tǒng)計學(xué)差異;較control組,SP組在復(fù)蘇后72h有較低的血糖值(P0.05),海馬CA1區(qū)有較多的存活神經(jīng)元(P0.05),較多的抗凋亡蛋白Bcl-2和較少的凋亡蛋白Bax(P0.05),以及上調(diào)了下游通路磷酸化蛋白p Akt及p GSK-3β(P0.05),而對總蛋白沒有顯著影響;而W+SP組較SP組各結(jié)果均有部分拮抗,如血糖值略有升高(P0.05),存活神經(jīng)元有所減少(P0.05),凋亡相關(guān)蛋白也有顯著的變化(P0.05),并且部分降低了p Akt和pGSK-3β蛋白的表達(dá)(P0.05)。結(jié)論在窒息性心搏驟停模型中,七氟烷對全腦損傷的保護(hù)作用不僅與PI3K/Akt介導(dǎo)的抑制凋亡作用有關(guān),可能還與GSK-3β參與的糖代謝調(diào)節(jié)作用密切相關(guān)。
[Abstract]:Cardiac arrest (CA) is the leading cause of death worldwide and can cause severe hypoxic-ischemic damage to the whole brain. Studies have shown that fewer than 50% of patients survive long-term after cardiac arrest, and most of them suffer from cognitive and behavioral impairments of varying degrees. Despite extensive basic research and clinical practice, no effective reduction has been found so far. In recent years, studies have shown that sevoflurane postconditioning can significantly improve the neurological deficit of focal brain injury, but the protective effect of sevoflurane postconditioning on whole brain injury after cardiopulmonary resuscitation is rarely reported. Phosphatidylinositol 3-kinase/protein kinase B (p Hosphatidyl inositol 3-kinase / protein kinase B (PI3K / Akt) pathway is one of the most important signaling pathways in vivo. It participates in cell growth, survival, metabolism and apoptosis, and plays an important role in drug-treated animal models of focal cerebral ischemia injury or global cerebral ischemia injury. E-3beta, GSK-3beta) is one of the important downstream factors of PI3K/Akt pathway. It not only participates in the growth and development of cells in vivo, but also regulates blood glucose and promotes tumor formation. Studies have shown that GSK-3beta not only plays a protective role in brain injury induced by four-vessel embolism, but also has a great deal of research on insulin receptors. It was found that GSK-3 beta can also regulate glucose metabolism in vivo through a variety of mechanisms, involving various aspects of glucose metabolism, such as affecting the function of islet beta cells, glycogen synthesis and so on. Serum neuron specific enolase (NSE), expression of apoptotic protein P53 in hippocampus, histopathological damage and changes of cognitive and behavioral abilities in rats were measured to investigate the protective effect of sevoflurane postconditioning on brain injury after asphyxiated cardiac arrest. Specific antagonists were used to evaluate the neuroprotective effect of PI3K/Akt pathway on sevoflurane postconditioning, reveal the role of PI3K/Akt pathway in brain protection after sevoflurane postconditioning, explore the relationship between PI3K/Akt pathway and anti-apoptosis and glucose metabolism regulation, and provide new ideas and methods for the treatment of brain injury after cardiopulmonary resuscitation (CPR). Objective To study the protective effects of sevoflurane postconditioning on brain injury in rats with asphyxiated cardiac arrest.The results showed that sevoflurane postconditioning could significantly improve the nerve defect in the model of focal brain injury,but the protective effects of sevoflurane postconditioning on brain injury after cardiopulmonary resuscitation were seldom studied. Methods 80 adult male SD rats were randomly divided into five groups: sham group, control group, 0.5 times minimum alveolar concentration (MAC), 1.0 MAC and 1.5 MAC sevoflurane postconditioning group (0.5 SP group, 1.0 SP group and 1.5 SP group). Control group, 0.5 SP group, 1.0 SP group and 1.5 SP group. Each sevoflurane treatment group was given intermittent inhalation of sevoflurane 2 times, 5 minutes each time, 10 minutes interval, using gas analyzer to ensure the concentration of 1.3%, 2.5% and 3.8% respectively, while the control group rats only inhaled 100% oxygen. Results The successful rate of resuscitation in each treatment group was 7 to 11 days after resuscitation. There was no significant difference in blood glucose, NSE, P53 expression, neurological function and spatial learning and memory ability between 1.0SP group and 1.5SP group at 72 hours after resuscitation (P 0.05), and there were more surviving neurons in CA1 area of hippocampus, less apoptotic protein P53 expression (P 0.05), but no significant neurological protection in 0.5SP group. Conclusion Sevoflurane postconditioning at high concentrations (1.0MAC and 1.5MAC) can significantly improve the whole brain injury induced by asphyxiated cardiac arrest in rats. Experiment 2: Effect of sevoflurane postconditioning on reducing PI3K/Akt in brain injury induced by asphyxiated cardiac arrest Objective PI3K/Akt pathway is one of the most important signaling pathways in vivo and participates in it. Cell growth, survival, metabolism and apoptosis. The downstream factor GSK-3beta may participate in the regulation of glucose metabolism from glycogen synthesis and other aspects. The aim of this study was to explore whether the brain protective effect of sevoflurane was related to PI3K/Akt pathway-mediated anti-apoptosis and GSK-3beta-mediated regulation of glucose metabolism. They were divided into five groups: sham group, control group, 1.0 MAC sevoflurane post-treatment group (SP group), PI3K specific inhibitor Wortmannin + sevoflurane post-treatment group (W + SP group) and pure Wortmannin group (W group). The specific treatment methods of Sham group, control group and SP group were the same as those of experiment 1. In addition, the cardiopulmonary resuscitation was performed by inhaling sevoflurane and ensuring 2.5% end-expiratory concentration. In group W, Wortmannin was administered via femoral vein 30 minutes before asphyxia, and the cardiac arrest model was established. Results There was no significant difference in the success rate of resuscitation among the groups. Compared with control group, SP group had lower blood glucose level 72 hours after resuscitation (P 0.05), hippocampal CA1 area had more surviving neurons (P 0.05), more anti-apoptotic proteins Bcl-2 and less. Apoptotic protein Bax (P 0.05), and up-regulation of phosphorylated protein P Akt and P GSK-3 beta (P 0.05) in the downstream pathway had no significant effect on total protein, but W+SP group had some antagonistic effects on the results of SP group, such as a slight increase in blood sugar (P 0.05), a decrease in survival neurons (P 0.05), and a significant change in apoptosis-related proteins (P 0.05). Conclusion The protective effect of sevoflurane on brain injury in asphyxiated cardiac arrest model is not only related to PI3K/Akt-mediated inhibition of apoptosis, but also closely related to GSK-3 beta-mediated regulation of glucose metabolism.
【學(xué)位授予單位】：第四軍醫(yī)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：R614

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 楊小霖;Edmond Eger;Manohar Sharma;;七氟烷在小鼠中的代謝研究[J];四川醫(yī)學(xué);2012年11期

2 李麗;呂國義;鄧^，

本文編號：2246541