七氟醚暴露致新生大鼠未成熟脑损伤的研究
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摘要
目的探讨七氟醚早期暴露对新生大鼠脑发育的影响。方法选取新生6日龄SD大鼠80只,随机分为四组,每组20只,分别为空白对照组(C组),1%七氟醚暴露2 h组(Sev1组),2%七氟醚暴露2 h组(Sev2组),4%七氟醚暴露2 h组(Sev3组);七氟醚暴露组分别置于1%、2%、4%七氟醚中每天暴露2 h,连续3 d;出生后第9天采用0.5%戊巴比妥钠麻醉后处死新生大鼠取出脑组织;采用电子天平秤检测各组大鼠在出生后6、9、15、28 d体重和脑重的变化;采用荧光定量PCR法(RT-PCR)检测Cx30、IL-6及BDNF基因mRNA表达量;采用免疫荧光法检测GFP蛋白含量;出生后28 d Morris水迷宫评价新生大鼠学习记忆能力。结果与C组比较,Sev1、Sev2和Sev3组新生大鼠体质量和脑质量均有所减轻,且随着日期延长这种减轻更为明显,Cx30 mRNA和BDNF mRNA表达量明显降低,IL-6mRNA表达量明显升高(P<0.05),GFP细胞数量明显减少(P<0.05);与Sev1组比较,Sev2和Sev3组新生大鼠体质量和脑质量均明显减轻,GFP数量明显减少(P<0.05);与C组比较,Sev1、Sev2和Sev3组逃避潜伏期和游动距离均明显延长(P<0.05),目标象限停留时间明显缩短(P<0.05),穿越虚拟平台次数明显减少(P<0.05)。结论新生大鼠接受七氟醚暴露可致未成熟脑损伤及远期学习记忆能力下降,且这种脑损伤程度与七氟醚浓度呈正相关;其机制可能与其脑内Cx30、BDNF基因表达水平改变有关。
Objective To investigate the effect and mechanism of sevoflurane exposure on immature brain injury in neonatal rats. Methods Eighty 6-day-SD neonatal rats were randomly divided into four groups, twenty rats in each group: blank control group(group C), 1% sevoflurane 2 h-exposure group(group sev1), and 2% sevoflurane 2 h-exposure group(group sev2), 4% sevoflurane 2 h-exposure group(group Sev3). Sevoflurane exposure group were placed in 1%, 2%, 4% sevoflurane exposure 2 h daily for 3 days respectivelly. The neonatal rats were sacrificed after anesthesia with 0.5% pentobarbital sodium and brain tissues were removed until the 9 th day after birth. The body weights and brain weights were measured on the 6 th, 9 th, 15 th, and 28 th day after birth. The expression of Cx30, IL-6 and BDNF mRNA were identified by fluorescence quantitative PCR(RT-PCR). The expressions of GFP protein were detected by immunofluorescence. The ability of memorize and study in these rats were assessed using Morris water maze postnatal day 28. Results Compared with group C, the body weight and brain mass of the neonatal rats in groups Sev1, Sev2 and Sev3 were decreased, and the reduction was more pronounced with the extension of the date; the expression of Cx30 mRNA and BDNF mRNA was down-regulated, and the expression of IL-6 mRNA was up-regulated significantly(P < 0.05); the number of GFP~(+ )cells was decreased significantly(P < 0.05). Compared with group Sev1, body weight and brain mass of newborn rats in groups Sev2 and Sev3 also decreased, and the number of GFP~+ cells was decreased significantly(P < 0.05). Compared with group C, the time of escape latency of groups Sev1, Sev2 and Sev3 after trains was significantly longer(P < 0.05); the time in target quadrant were significantly shorter(P<0.05); the number of times through virtual platform were significantly lower(P < 0.05). Conclusion Exposure of sevoflurane could result in immature brain development disorders and long-term learning and memory deficits, and the extent of brain damage is positively correlated to the concentration of sevoflurane; the mechanism may be related to the changes of intracerebral Cx30 and BDNF expression.
Objective To investigate the effect and mechanism of sevoflurane exposure on immature brain injury in neonatal rats. Methods Eighty 6-day-SD neonatal rats were randomly divided into four groups, twenty rats in each group: blank control group(group C), 1% sevoflurane 2 h-exposure group(group sev1), and 2% sevoflurane 2 h-exposure group(group sev2), 4% sevoflurane 2 h-exposure group(group Sev3). Sevoflurane exposure group were placed in 1%, 2%, 4% sevoflurane exposure 2 h daily for 3 days respectivelly. The neonatal rats were sacrificed after anesthesia with 0.5% pentobarbital sodium and brain tissues were removed until the 9 th day after birth. The body weights and brain weights were measured on the 6 th, 9 th, 15 th, and 28 th day after birth. The expression of Cx30, IL-6 and BDNF mRNA were identified by fluorescence quantitative PCR(RT-PCR). The expressions of GFP protein were detected by immunofluorescence. The ability of memorize and study in these rats were assessed using Morris water maze postnatal day 28. Results Compared with group C, the body weight and brain mass of the neonatal rats in groups Sev1, Sev2 and Sev3 were decreased, and the reduction was more pronounced with the extension of the date; the expression of Cx30 mRNA and BDNF mRNA was down-regulated, and the expression of IL-6 mRNA was up-regulated significantly(P < 0.05); the number of GFP~(+ )cells was decreased significantly(P < 0.05). Compared with group Sev1, body weight and brain mass of newborn rats in groups Sev2 and Sev3 also decreased, and the number of GFP~+ cells was decreased significantly(P < 0.05). Compared with group C, the time of escape latency of groups Sev1, Sev2 and Sev3 after trains was significantly longer(P < 0.05); the time in target quadrant were significantly shorter(P<0.05); the number of times through virtual platform were significantly lower(P < 0.05). Conclusion Exposure of sevoflurane could result in immature brain development disorders and long-term learning and memory deficits, and the extent of brain damage is positively correlated to the concentration of sevoflurane; the mechanism may be related to the changes of intracerebral Cx30 and BDNF expression.
引文
[1]孙杰,王忠云,丁正年.七氟醚麻醉诱导在患儿全身麻醉中的应用.临床麻醉学杂志,2008,24(6):468-470.
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[7]Katiyar KS,Winter CC,Gordián-Vélez WJ,et al.Three-dimensional tissue engineered aligned astrocyte networks to recapitulate developmental mechanisms and facilitate nervous system regeneration.J Vis Exp,2018,131:3791-3798.
[8]Fominykh V,Vorobyeva A,Onufriev MV,et al.Interleukin-6,s-nitrosothiols,and neurodegeneration in different central nervous system demyelinating disorders:is there a relationship.J Clin Neurol,2018,14(3):327-332.
[9]Fletcher JL,Murray SS,Xiao J.Brain-derived neurotrophic factor in central nervous system myelination:a new mechanism to promote myelin plasticity and repair.Int J Mol Sci,2018,19(12):1912-1916.
[10]Hamzei-Sichani F,Kamasawa N,Janssen WG,et al.Gap junctions on hippocampal mossy fiber axons demonstrated by thin-section electron microscopy and freeze fracture replica immunogold labeling.Proc Natl Acad Sci U S A,2007,104(30):12548-12553.
[11]Gosejacob D,Dublin P,Bedner P,et al.Role of astroglial connexin30 in hippocampal gap junction coupling.Glia,2011,59(3):511-519.
[12]Millin PM,Rickert GM.Effect of a strawberry and spinach dietary supplement on spatial Learning in early and late middleaged female rats.Antioxidants(Basel),2018,8(1):126-131.
[13]Ahmadi M,Rajaei Z,Hadjzadeh MA,et al.Crocin improves spatial learning and memory deficits in the Morris water maze via attenuating cortical oxidative damage in diabetic rats.Neurosci Lett,2017,642:1-6.
[2]Wilder RT,Flick RP,Sprung J,et al.Early exposure to anesthesia and learning disabilities in a population-based birth cohort.Anesthesiology,2009,110(4):796-804.
[3]Liang LQ,Jiao YQ,Guo SL.Effects of sevoflurane inhalation anesthesia on cognitive and immune function in elderly patients after abdominal operation.Eur Rev Med Pharmacol Sci,2018,22(24):8932-8938.
[4]Kostovic'I,Sedmak G,Juda2 M.Neural histology and neurogenesis of the human fetal and infant brain.Neuroimage,2018,10(18):32187-32196.
[5]Krmer TJ,Sakas W,Jussen D,et al.Thrombin contributes to the injury development and neurological deficit after acute subdural hemorrhage in rats only in collaboration with additional blood-derived factors.BMC Neurosci,2018,19(1):81-87.
[6]Clancy B,Finlay BL,Darlington RB,et al.Extrapolating brain development from experimental species to humans.Neurotoxicology,2007,28(5):931-937.
[7]Katiyar KS,Winter CC,Gordián-Vélez WJ,et al.Three-dimensional tissue engineered aligned astrocyte networks to recapitulate developmental mechanisms and facilitate nervous system regeneration.J Vis Exp,2018,131:3791-3798.
[8]Fominykh V,Vorobyeva A,Onufriev MV,et al.Interleukin-6,s-nitrosothiols,and neurodegeneration in different central nervous system demyelinating disorders:is there a relationship.J Clin Neurol,2018,14(3):327-332.
[9]Fletcher JL,Murray SS,Xiao J.Brain-derived neurotrophic factor in central nervous system myelination:a new mechanism to promote myelin plasticity and repair.Int J Mol Sci,2018,19(12):1912-1916.
[10]Hamzei-Sichani F,Kamasawa N,Janssen WG,et al.Gap junctions on hippocampal mossy fiber axons demonstrated by thin-section electron microscopy and freeze fracture replica immunogold labeling.Proc Natl Acad Sci U S A,2007,104(30):12548-12553.
[11]Gosejacob D,Dublin P,Bedner P,et al.Role of astroglial connexin30 in hippocampal gap junction coupling.Glia,2011,59(3):511-519.
[12]Millin PM,Rickert GM.Effect of a strawberry and spinach dietary supplement on spatial Learning in early and late middleaged female rats.Antioxidants(Basel),2018,8(1):126-131.
[13]Ahmadi M,Rajaei Z,Hadjzadeh MA,et al.Crocin improves spatial learning and memory deficits in the Morris water maze via attenuating cortical oxidative damage in diabetic rats.Neurosci Lett,2017,642:1-6.