目的 探讨细胞红蛋白(cytoglobin,CYGB)在缺氧缺血性脑损伤(hypoxic-ischemic brain damage,HIBD)中的神经保护作用. 方法 健康7日龄新生Sprague-Dawlay大鼠随机分为假手术组、HIBD组、HIBD+氯化高铁血红素(Hemin)组和HIBD+锌原卟啉(zinc protoporphyrin,ZnPP)组.HIBD组、HIBD+ Hemin组及HIBD+ ZnPP组大鼠分别腹腔注射0.5 ml生理盐水、Hemin(50 mg/kg)及ZnPP(50 mg/kg),12h后结扎并剪断左侧颈总动脉,缺氧2h,制成HIBD动物模型.建立模型后0、24、48 h免疫组织化学法检测各组大鼠海马及皮质中CYGB的表达情况,每组10只.HE染色观察建立模型后48 h的大鼠脑组织病理变化,每组8只.称重法检测建立模型后72h新生大鼠脑含水量,每组8只.建立模型后28 d采用Morris水迷宫实验检测海马学习记忆功能变化,每组10只.组间差异比较采用方差分析,组间两两比较使用Bonferroni法. 结果 (1)大鼠脑组织CYGB的表达水平:用平均灰度值(与蛋白表达水平成反比)表示.在0h,HIBD+ Hemin组和HIBD组皮质CYGB的平均灰度值分别为166.7±5.1和207.1±5.1,低于假手术组(232.3±3.4),而HIBD+ ZnPP组(234.9±4.5)高于假手术组(P＜0.05).随时间的增加,各HIBD组CYGB的平均灰度值递减,至48 h时,CYGB的平均灰度值HIBD+Hemin组最低(126.0±2.6),其次是HIBD组(150.9±4.5)和HIBD+ ZnPP组(163.7=6.3),最高是假手术组(232.1±5.8),差异均有统计学意义(P均＜0.01).CYGB在海马组织的表达变化与皮质一致.(2)各组脑组织病理变化:建立模型后48 h,HIBD组、HIBD+ Hemin组和HIBD+ ZnPP组新生大鼠脑组织可见典型的脑梗死和出血,HIBD+Hemin组梗死灶明显较HIBD+ZnPP组小,出血减轻.(3)脑组织含水量变化:建立模型后72 h,HIBD组和HIBD+ZnPP组左侧脑组织含水量分别为(86.5±0.4)％和(87.3±0.3)％,明显高于右脑[(85.6±0.2)％和(85.9±0.2)％],差异有统计学意义(t分别为12.57和11.32,P均＜0.01).(4)海马学习记忆功能变化:水迷宫实验中,5d总平均逃逸潜伏期HIBD+ ZnPP组最长[(76.7±29.8)s],其次是HIBD组[(71.0±30.5)s]和HIBD+ Hemin组[(46.7±34.0)s],最短为假手术组[(38.3±30.3)s],差异均有统计学意义(P均＜0.05).(5)远期脑组织病理变化:建立模型后34 d,脑组织萎缩率HIBD+ ZnPP组最高[(34.07±6.75)％],其次是HIBD组[(29.73±6.53)％],再次是HIBD+Hemin组[(18.33±4.52)％],均高于假手术组[(1.55±1.32)％],差异均有统计学意义(P均＜0.01).HIBD组及HIBD+ZnPP组大鼠脑组织可见锥体细胞层变薄,大片神经元缺失,HIBD+Hemin组仅见少量神经元缺失. 结论 CYGB的高表达可减轻HIBD脑组织近期及远期的病理损伤,保护远期的海马学习记忆功能.
目的 探討細胞紅蛋白(cytoglobin,CYGB)在缺氧缺血性腦損傷(hypoxic-ischemic brain damage,HIBD)中的神經保護作用. 方法 健康7日齡新生Sprague-Dawlay大鼠隨機分為假手術組、HIBD組、HIBD+氯化高鐵血紅素(Hemin)組和HIBD+鋅原卟啉(zinc protoporphyrin,ZnPP)組.HIBD組、HIBD+ Hemin組及HIBD+ ZnPP組大鼠分彆腹腔註射0.5 ml生理鹽水、Hemin(50 mg/kg)及ZnPP(50 mg/kg),12h後結扎併剪斷左側頸總動脈,缺氧2h,製成HIBD動物模型.建立模型後0、24、48 h免疫組織化學法檢測各組大鼠海馬及皮質中CYGB的錶達情況,每組10隻.HE染色觀察建立模型後48 h的大鼠腦組織病理變化,每組8隻.稱重法檢測建立模型後72h新生大鼠腦含水量,每組8隻.建立模型後28 d採用Morris水迷宮實驗檢測海馬學習記憶功能變化,每組10隻.組間差異比較採用方差分析,組間兩兩比較使用Bonferroni法. 結果 (1)大鼠腦組織CYGB的錶達水平:用平均灰度值(與蛋白錶達水平成反比)錶示.在0h,HIBD+ Hemin組和HIBD組皮質CYGB的平均灰度值分彆為166.7±5.1和207.1±5.1,低于假手術組(232.3±3.4),而HIBD+ ZnPP組(234.9±4.5)高于假手術組(P＜0.05).隨時間的增加,各HIBD組CYGB的平均灰度值遞減,至48 h時,CYGB的平均灰度值HIBD+Hemin組最低(126.0±2.6),其次是HIBD組(150.9±4.5)和HIBD+ ZnPP組(163.7=6.3),最高是假手術組(232.1±5.8),差異均有統計學意義(P均＜0.01).CYGB在海馬組織的錶達變化與皮質一緻.(2)各組腦組織病理變化:建立模型後48 h,HIBD組、HIBD+ Hemin組和HIBD+ ZnPP組新生大鼠腦組織可見典型的腦梗死和齣血,HIBD+Hemin組梗死竈明顯較HIBD+ZnPP組小,齣血減輕.(3)腦組織含水量變化:建立模型後72 h,HIBD組和HIBD+ZnPP組左側腦組織含水量分彆為(86.5±0.4)％和(87.3±0.3)％,明顯高于右腦[(85.6±0.2)％和(85.9±0.2)％],差異有統計學意義(t分彆為12.57和11.32,P均＜0.01).(4)海馬學習記憶功能變化:水迷宮實驗中,5d總平均逃逸潛伏期HIBD+ ZnPP組最長[(76.7±29.8)s],其次是HIBD組[(71.0±30.5)s]和HIBD+ Hemin組[(46.7±34.0)s],最短為假手術組[(38.3±30.3)s],差異均有統計學意義(P均＜0.05).(5)遠期腦組織病理變化:建立模型後34 d,腦組織萎縮率HIBD+ ZnPP組最高[(34.07±6.75)％],其次是HIBD組[(29.73±6.53)％],再次是HIBD+Hemin組[(18.33±4.52)％],均高于假手術組[(1.55±1.32)％],差異均有統計學意義(P均＜0.01).HIBD組及HIBD+ZnPP組大鼠腦組織可見錐體細胞層變薄,大片神經元缺失,HIBD+Hemin組僅見少量神經元缺失. 結論 CYGB的高錶達可減輕HIBD腦組織近期及遠期的病理損傷,保護遠期的海馬學習記憶功能.
목적 탐토세포홍단백(cytoglobin,CYGB)재결양결혈성뇌손상(hypoxic-ischemic brain damage,HIBD)중적신경보호작용. 방법 건강7일령신생Sprague-Dawlay대서수궤분위가수술조、HIBD조、HIBD+록화고철혈홍소(Hemin)조화HIBD+자원계람(zinc protoporphyrin,ZnPP)조.HIBD조、HIBD+ Hemin조급HIBD+ ZnPP조대서분별복강주사0.5 ml생리염수、Hemin(50 mg/kg)급ZnPP(50 mg/kg),12h후결찰병전단좌측경총동맥,결양2h,제성HIBD동물모형.건립모형후0、24、48 h면역조직화학법검측각조대서해마급피질중CYGB적표체정황,매조10지.HE염색관찰건립모형후48 h적대서뇌조직병리변화,매조8지.칭중법검측건립모형후72h신생대서뇌함수량,매조8지.건립모형후28 d채용Morris수미궁실험검측해마학습기억공능변화,매조10지.조간차이비교채용방차분석,조간량량비교사용Bonferroni법. 결과 (1)대서뇌조직CYGB적표체수평:용평균회도치(여단백표체수평성반비)표시.재0h,HIBD+ Hemin조화HIBD조피질CYGB적평균회도치분별위166.7±5.1화207.1±5.1,저우가수술조(232.3±3.4),이HIBD+ ZnPP조(234.9±4.5)고우가수술조(P＜0.05).수시간적증가,각HIBD조CYGB적평균회도치체감,지48 h시,CYGB적평균회도치HIBD+Hemin조최저(126.0±2.6),기차시HIBD조(150.9±4.5)화HIBD+ ZnPP조(163.7=6.3),최고시가수술조(232.1±5.8),차이균유통계학의의(P균＜0.01).CYGB재해마조직적표체변화여피질일치.(2)각조뇌조직병리변화:건립모형후48 h,HIBD조、HIBD+ Hemin조화HIBD+ ZnPP조신생대서뇌조직가견전형적뇌경사화출혈,HIBD+Hemin조경사조명현교HIBD+ZnPP조소,출혈감경.(3)뇌조직함수량변화:건립모형후72 h,HIBD조화HIBD+ZnPP조좌측뇌조직함수량분별위(86.5±0.4)％화(87.3±0.3)％,명현고우우뇌[(85.6±0.2)％화(85.9±0.2)％],차이유통계학의의(t분별위12.57화11.32,P균＜0.01).(4)해마학습기억공능변화:수미궁실험중,5d총평균도일잠복기HIBD+ ZnPP조최장[(76.7±29.8)s],기차시HIBD조[(71.0±30.5)s]화HIBD+ Hemin조[(46.7±34.0)s],최단위가수술조[(38.3±30.3)s],차이균유통계학의의(P균＜0.05).(5)원기뇌조직병리변화:건립모형후34 d,뇌조직위축솔HIBD+ ZnPP조최고[(34.07±6.75)％],기차시HIBD조[(29.73±6.53)％],재차시HIBD+Hemin조[(18.33±4.52)％],균고우가수술조[(1.55±1.32)％],차이균유통계학의의(P균＜0.01).HIBD조급HIBD+ZnPP조대서뇌조직가견추체세포층변박,대편신경원결실,HIBD+Hemin조부견소량신경원결실. 결론 CYGB적고표체가감경HIBD뇌조직근기급원기적병리손상,보호원기적해마학습기억공능.
Objective To investigate the protective function of cytoglobin (CYGB) on neurodevelopment of the neonatal rats with hypoxic-ischemic brain damage (HIBD).Methods Healthy seven day old Sprague-Dawlay rats were randomly divided into sham operated group,HIBD group,HIBD+Hemin group and HIBD+zinc protoporphyrin (ZnPP) group.The rats of HIBD,HIBD+Heminand HIBD+ZnPPgroup were given normal saline (0.5 ml),Hemin (50 mg/kg) and ZnPP (50 mg/kg) intraperitoneally respectively,and 12 hours later the left carotid arteries of these rats were ligated and cut off,then hypoxic treated for 2 hours to establish the HIBD models.At 0,24,48 h after HIBD models were established,the expressions of CYGB in the cerebral cortex and hippocampus were observed by immunohistochemistry analysis.At 48 h after HIBD,histopathological changes of brain were observed after HE staining.At 72 h after HIBD,the water content of the brain was observed.At 28 days after HIBD,long term study memory outcome was assessed by Morris water maze.Analysis of variance and Bonferroni test were applied as statistical methods.Results (1)The expression of CYGB in brain(expressed by average gray value which negatively correlated with protein levels):At 0 h,the average gray value of CYGB in the cerebral cortex in HIBD+ Hemin and HIBD group were 166.7±5.1 and 207.1±5.1,which were lower than that in sham operated group (232.3±3.4); but in HIBD+ZnPP group,it was higher (234.9±4.5)(P＜0.05).The average gray value of CYGB was decreased with the extension of hypoxic-ischemic time.At 48 h,the average gray value of CYGB was the lowest in HIBD+Hemin group (126.0± 2.6),followed by HIBD group (150.9±4.5) and HIBD+ZnPP group (163.7±6.3),and the highest was in sham operated group (232.1±5.8)(all P＜0.01).(2) Histopathologic changes of the brain:At 48 h,typical cerebral infarction and hemorrhage were seen in HIBD,HIBD+Hemin and HIBD+ZnPP group,but those were less severe in HIBD + Hemin group than in HIBD + ZnPP group.(3) The water content of the brain:At 72 h,the water content of the left brain in HIBD and HIBD+ZnPP group was (86.5±0.4)％ and (87.3±0.3)％,which was higher than that in right brain [(85.6±0.2)％ and (85.9±0.2)％] (t 12.57 and 11.32,P＜0.01,respectively).(4)Function of the hippocampus:Morris water-maze showed that the longest average escape latency in HIBD+ZnPP group [(76.7±29.8) s],followed by HIBD group [(71.0±30.5) s],HIBD+ Hemin group [(46.7±34.0) s],and sham operated group [(38.3±30.3) s] (all P＜0.01).(5) Long-term histopathologic changes of the brain:At 34 d,brain atrophy rate was the highest in HIBD+ZnPP group [(34.07± 6.75) ％],and then in HIBD group [(29.73± 6.53) ％] and HIBD+ Hemin group [(18.33±4.52)％],which were all higher than that in the sham operated group [(1.55±1.32)％](all P＜0.01).HE staining showed that the hippocampal stratum pgramidale was getting thinner and a large number of neurons was lost in HIBD and HIBD+ ZnPP group,but only a small amout of neurons was lost in HIBD+ Hemin group.Conclusions Increased expression of CYGB in HIBD brain could mitigate the short term and long term pathological injury,and protect the long-term study memory function of the hippocampus.
