【摘要】：背景每年全世界數(shù)百萬嬰幼兒接受全身麻醉和手術(shù),但全身麻醉藥對神經(jīng)系統(tǒng)處于突觸爆發(fā)期的兒童智力是否有影響,還不清楚。新生動物使用全身麻醉藥麻醉后會出現(xiàn)多個腦區(qū)神經(jīng)細胞凋亡、突觸發(fā)育的異常和神經(jīng)退行性變,有些在成年后還會遺留神經(jīng)認知或社交方面的障礙,其具體機制還不清楚。目的通過實驗研究七氟烷對核糖體蛋白S6的影響及其對其上下游通路的信號分子的影響,以研究七氟烷對神經(jīng)系統(tǒng)作用的機制。方法建立新生動物七氟烷吸入麻醉模型。選用P8的SD大鼠隨機分為兩組,麻醉組和氧氣對照組。以70%氧氣空氣混合氣為載氣,麻醉組大鼠給予七氟烷麻醉,分別在0.5h,1h,2h,4h取樣,當麻醉持續(xù)4h后停止麻醉,將鼠仔放回母親身邊,并于放回后0.5h和2h分別取樣。另一組是氧氣對照組,直接使用70%空氣氧氣混合氣吸入。并在相同時間點取樣。一方面,取出新生大鼠的海馬和皮層組織,抽提蛋白行蛋白印跡實驗,進行Caspase-3,rp S6,p-rp S6,AKT,pAKT,m TOR,p-m TOR,ERK,p-ERK,JNK,p-JNK,P38,p-P38檢測,以actin為內(nèi)參。另一方面將新生大鼠從左心室灌流,取腦切片,行免疫組化實驗,檢測Caspase-3和p-rp S6的表達分部狀況。此外,七氟烷麻醉4h和氧氣對照4h的新生大鼠剝?nèi)『ｑR和皮層,抽提RNA后送樣進行基因芯片檢測,觀察七氟烷新生動物麻醉對海馬和皮層基因表達的影響。結(jié)果七氟烷新生動物麻醉4h后,在海馬和皮層均可見Caspase-3的表達上調(diào);給予新生大鼠七氟烷持續(xù)麻醉,會造成大腦皮層和CA1區(qū)明顯時間依賴性rp S6磷酸化的抑制;七氟烷能明顯抑制大腦皮層和海馬區(qū)的AKT活性,但對AKT的下游的m TOR沒有影響;七氟烷對海馬和大腦皮層的ERK活性產(chǎn)生了比較矛盾的效應(yīng);七氟烷增加了皮層和海馬的PP1的活性;基因芯片提示七氟烷麻醉對多種基因的表達產(chǎn)生了上調(diào)或下調(diào)的效應(yīng),這些基因中許多是由于神經(jīng)系統(tǒng)發(fā)育,認知記憶密切相關(guān)的基因。結(jié)論七氟烷麻醉新生動物確實會引起中樞神經(jīng)系統(tǒng)的細胞凋亡等神經(jīng)損傷的表現(xiàn)。這一損傷可能是由于七氟烷作用于核糖體蛋白S6使其磷酸化受到抑制所介導(dǎo)的,這一抑制作用又是通過對不依賴于m TOR的AKT活性抑制和對PP1活性增強實現(xiàn)的�；蛐酒慕Y(jié)果提示,七氟烷對新生動物的神經(jīng)系統(tǒng)發(fā)育相關(guān)的多個基因產(chǎn)生強烈的抑制或激活作用,他們也可能是導(dǎo)致細胞退變的原因之一。
[Abstract]:Background millions of infants around the world are subjected to general anesthesia and surgery every year, but it is not clear whether general anesthesia has an effect on the intelligence of children whose nervous system is in synaptic outbreak. After anesthetized with general anesthetic, the newborn animals may have several neuronal apoptosis, abnormal synaptic development and neurodegenerative changes, and some will leave behind cognitive or social obstacles in adulthood, the specific mechanism is not clear. Aim to investigate the effects of sevoflurane on ribosomal protein S6 and signal molecules in its upstream and downstream pathways in order to study the mechanism of the effects of sevoflurane on nervous system. Methods the sevoflurane inhalation anesthesia model of newborn animals was established. SD rats with P 8 were randomly divided into two groups: anesthesia group and oxygen control group. The anesthetized rats were anesthetized by sevoflurane with 70% oxygen air mixture as carrier. The anesthetized rats were taken at 0.5 h for 1 h and 2 h for 4 h respectively. The anesthetic was stopped after 4 h of anesthesia, and the rats were put back to their mothers. The rats were sampled at 0.5 h and 2 h after the anesthesia was put back. The other group was oxygen control group, which was inhaled directly with 70% air oxygen mixture. And sampling at the same time. On the one hand, the hippocampal and cortical tissues of newborn rats were taken out, and the protein Bank Western blot test was carried out. The detection of p-ERKP38 was carried out by using actin as the internal control. On the other hand, the neonatal rats were perfused from the left ventricle, the brain sections were taken and the expression of Caspase-3 and p-rp S6 were detected by immunohistochemistry. In addition, sevoflurane anesthetized newborn rats were anesthetized for 4 h and oxygen control group for 4 h. The hippocampus and cortex of newborn rats were stripped. After RNA was extracted, the gene expression in hippocampus and cortex was detected by gene chip analysis. The effect of sevoflurane neonate anesthesia on gene expression in hippocampus and cortex was observed. Results the expression of Caspase-3 was up-regulated in hippocampus and cortex 4 hours after sevoflurane anesthesia in neonatal rats, and the inhibition of rpS6 phosphorylation in cerebral cortex and CA1 area was induced by continuous sevoflurane anesthesia in neonatal rats. Sevoflurane inhibited AKT activity in cerebral cortex and hippocampus, but had no effect on m TOR downstream of AKT. Sevoflurane had a contradictory effect on ERK activity in hippocampus and cerebral cortex. Sevoflurane increased the activity of PP1 in cortex and hippocampus, and gene chip suggested that sevoflurane anesthesia could up-regulate or down-regulate the expression of many genes, many of which were related to the development of nervous system and cognitive memory. Conclusion sevoflurane anesthetized newborn animals can induce neuronal damage such as apoptosis in the central nervous system. The damage may be mediated by the inhibition of phosphorylation of ribosomal protein S6 by sevoflurane, which is mediated by inhibition of AKT activity independent of m TOR and enhancement of PP1 activity. The microarray results suggest that sevoflurane strongly inhibits or activates multiple genes associated with the development of the nervous system in newborn animals, and they may also be one of the causes of cell degeneration.
【學(xué)位授予單位】：上海交通大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2014
【分類號】：R726.1

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