摘要
目的探讨非竞争性N-甲基-D-天冬氨酸(NMDA)受体拮抗剂美金胺对缺血诱导未成熟脑白质内谷氨酸异常信号传输的体内外阻断效果。方法制备2日龄新生大鼠少突胶质细胞(OL)前体氧糖剥夺(OGD)细胞模型,分为对照组、OGD组及美金胺组。应用高效液相色谱仪测定OL前体胞外谷氨酸水平,流式细胞仪检测胞内钙离子浓度和OL前体凋亡率。制备缺血型脑室周围白质软化(PVL)动物模型,分为假手术组、PVL组及美金胺组。分别进行光镜下脑白质病理评估、免疫组织化学法检测脑白质髓鞘碱性蛋白(MBP)阳性OL表达率及电镜下进行脑白质髓鞘形成状况评估。结果体外与对照组相比,OGD组OL前体胞外谷氨酸水平增高[(24.60±2.42)μmol/L比(9.49±1.08)μmol/L,t=9.28,P<0.01],胞内钙离子浓度显著增加[(32.9±6.9)%比(6.9±3.5)%,t=4.41,P<0.01],OL前体凋亡率亦明显增高[(24.77±2.05)%比(6.65±1.39)%,t=15.01,P<0.01]。经美金胺处理后,胞外谷氨酸水平[(14.70±1.70)μmol/L]、胞内钙离子浓度[(23.1±2.0)%]和OL前体凋亡率[(11.80±2.06)%]均显著降低,差异均有统计学意义(t=5.68、6.13、5.18,均P<0.01)。体内与假手术组相比,PVL组大鼠脑白质均呈中至重度病变,脑白质内MBP阳性OL表达率[(5.94±1.37)%比(15.40±3.22)%,t=4.63,P<0.01]明显降低,髓鞘数目(4.00±1.00比14.67±2.70,t=6.11,P<0.01)明显减少,髓鞘厚度[(33.83±3.21)nm比(79.67±6.45)nm,t=10.43,P<0.01]明显变薄。予美金胺处理后,脑白质病理(Z=88.479,P<0.01)、髓鞘数目(10.30±1.50,t=6.01,P<0.01)及髓鞘厚度[(57.21±4.05)nm,t=7.47,P<0.01]均显著改善,脑白质内MBP阳性OL表达率[(11.02±1.35)%,t=4.40,P<0.05]亦明显增高。结论缺血诱导未成熟脑白质内谷氨酸异常信号传输是发生缺血性PVL的重要途径,美金胺可有效阻断此异常信号传输,有望为早产儿缺氧缺血PVL的防治提供新途径。
Objective To investigate the blocking effect of non-competitive N-methyl-D-aspartic acid(NMDA)receptor antagonist Memantine on glutamate abnormal signal transmission in immature white matter induced by ischemia in vitro and in vivo.Methods The oligodendrocyte(OL)precursor oxygen glucose deprivation(OGD)cell models of 2-day-old newborn rats were prepared and divided into the normal control group,the OGD group and the Memantine group.The extracellular glutamate level of the OL precursor was measured by high performance liquid chromatography,while the concentration of intracellular calcium and the apoptosis rate of OL precursor were detected by flow cytometry.The animal models of ischemic periventricular leukomalacia(PVL)were established and divided into the sham group,the PVL group and the Memantine group.The pathological evaluation of white matter was performed under light microscope.The positive OL expression rate of myelin basic protein(MBP)was detected by immunohistoche-mistry.The myelination of white matter was evaluated under electron microscope.Results Compared with the normal control groupin vitro,the OGD group had a higher extracellular glutamate level of the OL precursor[(24.60±2.42)μmol/L vs.(9.49±1.08)μmol/L,t=9.28,P<0.01],a higher intracellular calcium concentration[(32.9±6.9)%vs.(6.9±3.5)%,t=4.41,P<0.01],a higher apoptosis rate of the OL precursor[(24.77±2.05)%vs.(6.65±1.39)%,t=15.01,P<0.01].After treatment with Memantine,the extracellular glutamate level[(14.70±1.70)μmol/L,t=5.68,P<0.01],the intracellular calcium concentration[(23.1±2.0)%,t=6.13,P<0.01],and the apoptosis rate of the OL precursor[(11.80±2.06)%,t=5.18,P<0.01]decreased significantly.Compared with the sham group in vivo,the white matter of the PVL group showed mild or severe pathological changes,and the PVL group had a lower MBP-positive OL expression rate in the white matter[(5.94±1.37)%vs.(15.40±3.22)%,t=4.63,P<0.01]less myelin sheaths(4.00±1.00 vs.14.67±2.70,t=6.11,P<0.01)and thinner myelin sheaths[(33.83±3.21)nm vs.(
作者
何亚芳
张建华
陈惠金
钱龙华
He Yafang;Zhang Jianhua;Chen Huijin;Qian Longhua(Department of Pediatric Respiratory,Xinhua Hospital Affiliated to Shanghai Jiaotong University School of Medicine,Shanghai 200092,China;Perinatal Research Department,Shanghai Institute for Pediatric Research,Shanghai 200092,China)
出处
《中华实用儿科临床杂志》
CSCD
北大核心
2020年第14期1093-1097,共5页
Chinese Journal of Applied Clinical Pediatrics
基金
国家自然科学基金(30672246)
上海市卫计委课题(2010Y149)。
关键词
美金胺
缺血
未成熟脑白质
谷氨酸
异常信号传输
Memantine
Ischemia
Immature white matter
Glutamate
Abnormal signal transmission
