异氟醚通过激活FAS信号途径引起新生小鼠海马损伤和学习记忆障碍

发布时间：2018-06-18 06:06

本文选题：异氟醚 + 发育期大脑　；参考：《复旦大学》2014年硕士论文

【摘要】：第一部分异氟醚麻醉损伤新生小鼠海马并引起学习记忆障碍目的：探讨异氟醚对新生小鼠海马的损伤及学习记忆能力的影响。方法：健康新生7日龄C57BL/6小鼠,随机分为两组：异氟醚组和对照组。异氟醚组：出生第7日起开始进行浓度为1.5%的异氟醚麻醉,麻醉三天(出生第7、8、9日),每天2小时,吸入氧浓度100%；对照组在相应的时间仅吸入100%氧气,麻醉过程中保温保湿。两组在第9日麻醉后立即处死,解剖取脑,分离海马,行Western Blot检测海马组织凋亡标志蛋白Caspase-3; TUNEL染色计数海马区凋亡细胞；Morris水迷宫实验检测以后的学习记忆能力。结果：与对照组相比,异氟醚组小鼠海马组织Caspase-3表达显著增加,TUNEL染色海马区阳性细胞数明显增多,差异均有统计学意义(P0.05)；水迷宫实验结果,与对照组相比,异氟醚组小鼠逃避潜伏期明显延长,平台穿越次数显著减少,差异有统计学意义(P0.05)。结论：异氟醚麻醉新生小鼠可致其海马区神经细胞凋亡,引起海马损伤,影响大脑发育,进而引起其以后的学习记忆能力减退。第二部分异氟醚通过激活FAS信号途径损伤新生小鼠海马并引起学习记忆障碍目的：探讨异氟醚可通过激活FAS信号途径损伤新生小鼠海马损伤并引起学习记忆能力障碍。方法：健康新生7日龄C57BL/6小鼠,随机分为两组,分别为异氟醚组和空白对照组。两组麻醉方法同第一部分。在三天麻醉后,脱颈处死,取海马,行Western Blot检测FAS蛋白(CD95)和FAS配体蛋白(FASligand,FASL)。另取FAS或FASL基因敲除的B6. MRL-Faslpr/Jnju小鼠和B6Smn. C3-Faslgld/Jnju小鼠作为FAS基因缺陷组和FASL基因缺陷组,对两种小鼠分别做吸入异氟醚和单纯氧气的处理,用第一部分中相同方法麻醉和单纯吸氧三天。用Western Blot方法检测四组小鼠(FAS基因敲除异氟醚组、FAS基因敲除纯氧组、FASL基因敲除异氟醚组和FASL基因敲除纯氧组)的Caspase-3的表达。用TUNEL染色法观察FAS基因敲除异氟醚组和FASL基因敲除异氟醚组小鼠海马区阳性细胞数。用水迷宫实验检测FAS基因敲除异氟醚组和FASL基因敲除异氟醚组小鼠的学习记忆能力。将三个实验的结果与第一部分中相应健康异氟醚组和对照组的结果合并、对比观察并统计分析。结果：异氟醚组麻醉的健康新生小鼠其FAS蛋白和FASL蛋白均较空白对照组表达显著增加,差异有统计学意义(p0.05)。异氟醚健康组Caspase-3表达较FAS缺陷异氟醚组和FASL缺陷异氟醚组显著增加,差异有统计学意义(p0.05)；而对照组分别与FAS缺陷纯氧组、FAS缺陷异氟醚组、FASL缺陷纯氧组和FASL缺陷异氟醚组相比,Caspase-3表达差异均无统计学意义；Two-way ANOVA分析可得异氟醚麻醉与FAS或FASL基因敲除在Caspase-3的表达影响上有联系,FAS或FASL基因敲除可能减弱了异氟醚所引起的Caspase-3表达的增加。异氟醚健康组TUNEL染色阳性细胞数较FAS缺陷异氟醚组和FASL缺陷异氟醚组均显著增加；而对照组与FAS缺陷异氟醚组和FASL缺陷异氟醚组的TUNEL阳性细胞数相比,差异无统计学意义。异氟醚健康组的逃避潜伏期较FAS缺陷异氟醚组和FASL缺陷异氟醚组明显延长,平台穿越次数明显减少,差异有统计学意义(p0.05)；而对照组与该两组小鼠的差异均无统计学意义。结论：异氟醚可通过激活FASL-FAS信号途径引起健康新生小鼠海马区细胞的凋亡,从而损伤海马并引起其以后的学习记忆能力下降。对于切断FASL-FAS信号通路的小鼠则可减轻该种损伤,学习记忆能力也有明显改善。从而证明FAS通路是异氟醚损伤新生小鼠海马的机制之一。
[Abstract]:Part 1: the effects of isoflurane on the hippocampus and learning and memory impairment in newborn mice: the effects of isoflurane on hippocampal damage and learning and memory ability of newborn mice. Methods: 7 days old C57BL/6 mice were randomly divided into two groups: isoflurane group and control group: isoflurane group: the concentration of isoflurane began to be concentrated on the seventh day of birth. 1.5% isoflurane anesthesia, three days of anesthesia (7,8,9 day of birth), 2 hours a day, inhaled oxygen concentration 100%; the control group was only inhaled 100% oxygen at the corresponding time and kept warm and moisturizing during the anesthesia. The two groups died immediately after ninth days of anesthesia, dissected the brain, separated the hippocampus, and Western Blot was used to detect the apoptosis marker protein Caspase-3 of the hippocampus; T UNEL staining was used to count the apoptotic cells in the hippocampus and the learning and memory ability after the Morris water maze test. Results: compared with the control group, the expression of Caspase-3 in the hippocampus of the isoflurane group increased significantly, and the number of positive cells in the hippocampus of the TUNEL stained hippocampus increased significantly (P0.05). The results of water maze experiment and control were compared with the control group. Compared with the group, the escape latency of the isoflurane group was significantly prolonged, and the number of platform crossing times decreased significantly (P0.05). Conclusion: Isoflurane can induce neuronal apoptosis in the hippocampus, damage the hippocampus, affect the brain development, and induce the impairment of learning and memory. Second parts of the isoflurin can be induced by isoflurane. Ether can damage the hippocampus of newborn mice by activating FAS signal pathway and cause learning and memory impairment: Isoflurane can damage hippocampus damage in newborn mice by activating FAS signal pathway and cause learning and memory impairment. Methods: healthy newborn 7 day old C57BL/6 mice were randomly divided into two groups, isoflurane group and blank control group, respectively. The two groups were anesthetized with the first part. After three days of anesthesia, they were removed from the neck and took the hippocampus, and the FAS protein (CD95) and the FAS ligand protein (FASligand, FASL) were detected by Western Blot. The B6. MRL-Faslpr/Jnju mice of FAS or FASL knockout were taken as the gene defect group and the gene defect group, and two mice were used. The expression of Caspase-3 in four groups of mice (FAS knockout isoflurane group, FAS knockout pure oxygen group, FASL gene knockout isoflurane group and FASL gene knockout pure oxygen group) was detected by Western Blot method, with the same method of inhaling isoflurane and simple oxygen, respectively. The expression of Caspase-3 in the FAS gene knockout isoflurane group, the FAS knockout isoflurane group, the FASL gene knockout isoflurane group and the FASL gene knockout group of pure oxygen group. The number of positive cells in the hippocampus of FAS gene knockout isoflurane group and FASL gene knockout isoflurane group. The learning and memory ability of FAS gene knockout isoflurane group and FASL gene knockout isoflurane group was detected by water maze test. The results of the three experiments were combined with the results of the Xiang Yingjian Kang isoflurane group and the control group in the first part, and the results were combined with the results of the first part. Results: the expression of FAS protein and FASL protein in the healthy newborn mice anaesthetized by isoflurane increased significantly than that in the blank control group. The difference was statistically significant (P0.05). The expression of Caspase-3 in isoflurane healthy group was significantly higher than that of FAS deficient isoflurane group and FASL deficient isoflurane group (P0.05). Compared with the FAS defect pure oxygen group, the FAS deficient isoflurane group, the FASL defect pure oxygen group and the FASL deficient isoflurane group, there was no significant difference in the Caspase-3 expression, and the Two-way ANOVA analysis could be linked to the expression of FAS or FASL gene knockout in Caspase-3, and the FAS or FASL gene knockout might be weakened. The expression of Caspase-3 induced by isoflurane increased. The number of positive TUNEL staining cells in isoflurane group was significantly higher than that in the FAS deficient isoflurane group and the FASL deficient isoflurane group, while the control group had no statistical difference compared with the number of TUNEL positive cells in the FAS deficient isoflurane group and the FASL deficient isoflurane group. The escape latency was significantly longer than that of FAS defect isoflurane group and FASL deficient isoflurane group, and the number of platform crossing times decreased significantly (P0.05), but there was no significant difference between the control group and the two groups of mice. Conclusion: Isoflurane can induce the cells in the hippocampus of healthy newborn mice by activating FASL-FAS signal pathway. Apoptosis, which can damage the hippocampus and cause the decline of learning and memory ability in the future, can reduce the damage and improve the learning and memory ability in mice that cut off the FASL-FAS signaling pathway, which proves that the FAS pathway is one of the mechanisms of isoflurane damage to the hippocampus of newborn mice.
【学位授予单位】：复旦大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R614.2

【共引文献】