Erythropoietin increases glutathione peroxidase enzyme activity and decreases lipid peroxidation levels in hypoxic-ischemic brain injury in neonatal rats

BACKGROUND: We have previously shown that erythropoietin (Epo) exerts neuroprotective effects in the Rice-Vannucci model of neonatal hypoxic-ischemic brain injury. However, the mechanisms of Epo protection in this model are still unclear. OBJECTIVES: In the present study, we studied the effects of systemically administered Epo on lipid peroxidation levels and antioxidant enzyme (superoxide dismutase and glutathione peroxidase) activities following hypoxic-ischemic brain injury in neonatal rats. METHODS: Seven-day-old Wistar rat pups were subjected to left carotid artery occlusion followed by 2.5 h of hypoxic exposure. Brain lipid peroxidation levels and antioxidant enzyme activities were measured in the injured hemispheres 24 h after the hypoxic-ischemic insult. RESULTS: Hypoxic-ischemic injury significantly increased the thiobarbituric acid-reactive substance levels in the injured hemispheres as compared to the control group. In addition, glutathione peroxidase activity was significantly elevated in Epo-treated animals compared to saline-treated animals and the control group. CONCLUSIONS: These results suggest that Epo exerts neuroprotective effects against hypoxic-ischemic brain injury at least partially via the modulation of antioxidant enzyme activity.     

新生大鼠缺氧缺血性脑损伤COX-2 mRNA表达变化

研究新生大鼠缺氧缺血性脑损伤(HIBD)时环氧合酶-2(COX-2)mRNA的表达变化,应用、制备新生大鼠左脑HIBD的模型.用逆转录多聚酶链反应(RT-PRC)检测HIBD后6 h、24h、48 h、5 d不同时点脑皮层COX-2 mRNA的表达情况.经凝胶成像及分析系统扫描RT-PCR产物,用内参半定量分析COX-2 mRNA的动态变化.结果显示七日龄Wistar大鼠对照组即有COX-2 mRNA的低表达,HIBD 6 h表达开始升高,HIBD 24～48 h为表达高峰(与对照组相比有显著性差异,P＜0.01),HIBD 5 d表达下降.结论新生大鼠HIBD可诱导COX-2基因表达,其可能参与HIBD后的神经毒性损伤.     

8-异前列腺素F2α及其在新生儿缺氧缺血性脑损伤中的作用

1概述 新生儿缺氧缺血性脑损伤（HIBD）多由围生期窒息所致，是新生儿死亡及儿童伤残的重要原因之一。如何对HIBD患儿病情早期做出正确判断并实旅有效的干预，防止或减少HIBD的发生，降低致残率，是围生医学领域的重大课题。迄今为止，HIBD的发病机制尚未完全明确，也缺乏有效评定患儿脑损伤程度的指标．但一般认为缺血再灌注后的氧化应激在HIBD的发生发展中具有重要作用。氧化应激是体内自由基产物超过抗氧化防御代谢时的一种不平衡状态，可发生脂质过氧化作用、膜蛋白损伤及DNA损害。     

Effects of dexamethasone in hypoxic-ischemic brain injury in the neonatal rat

To clarify the effects of corticosteroids in neonatal hypoxic-ischemic brain injury, 7-day-old rats were subjected to unilateral common carotid artery ligation and hypoxia (Levine procedure) after being injected subcutaneously with saline, low-dose dexamethasone (4 mg/kg) or high-dose dexamethasone (40 mg/kg). Neither low-dose nor high-dose dexamethasone ameliorated the brain edema, lactacidemia, or hypoglycemia associated with hypoxia-ischemia. In addition, dexamethasone did not alter the pattern of neuropathologic damage or reduce the fall in brain high-energy phosphates. Finally, high-dose dexamethasone-treated animals experienced significantly more mortality than did either saline- or low-dose dexamethasone-treated animals. In this model of neonatal hypoxia-ischemia, dexamethasone did not confer any significant cerebral protection.     

川芎嗪对新生鼠缺氧缺血性脑损伤c-fos基因表达影响的研究

目的：探讨川芎嗪对缺氧缺血脑损伤(HIBD)脑组织c-fos基因表达的影响。方法：通过结扎左侧颈总动脉后置入氮氧混合气中2 h建立HIBD模型,采用RT-PCR方法分别半定量检测空白对照组、生理盐水组、及川芎嗪干预组海马及皮质c-fos mRNA水平。结果：生理盐水组海马、皮质c-fos mRNA明显高于空白对照组(P<0.05),川芎嗪干预组海马、皮质c-fos mRNA水平明显低于生理盐水组(P<0.05)。结论：HIBD新生鼠经川芎嗪干预后可明显降低脑组织海马、皮质c-fos基因的表达水平。     

间充质干细胞移植治疗缺氧缺血性脑损伤

缺氧缺血性脑损伤(hypoxie-ischaemie brain injury,HIBI)是导致新生儿死亡和儿童神经功能障碍的主要原因,目前临床尚无有效的治疗方法.间充质干细胞(mesenchymal stem cells,MSC)移植治疗为减少缺氧缺血性脑损害带来了希望.     

间充质干细胞移植治疗缺氧缺血性脑损伤

缺氧缺血性脑损伤(hypoxie-ischaemie brain injury,HIBI)是导致新生儿死亡和儿童神经功能障碍的主要原因,目前临床尚无有效的治疗方法.间充质干细胞(mesenchymal stem cells,MSC)移植治疗为减少缺氧缺血性脑损害带来了希望.     

Neuroprotective effect of erythropoietin on hypoxic-ischemic brain injury in neonatal rats

Erythropoietin (Epo) prevents ischemia and hypoxia-induced neuronal death in vitro. Recent studies have shown that this cytokine also has in vivo neuroprotective effects in cerebral and spinal ischemia in adult rodents. In this study, we aimed to investigate the effect of systemically administered recombinant human Epo on infarct volume and apoptotic neuronal death in a newborn rat hypoxic-ischemic brain injury model. Our results showed that a single dose of intraperitoneal Epo treatment (1,000 U/kg) significantly decreased the mean infarct volume as compared to the control group. In contrast to the Epo-treated group, histopathological examination by positive terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling of the affected brain in control animals revealed widespread neuronal injury associated with numerous apoptotic cells. Morphometric analysis to determine the extent of damage quantitatively ascertained that the mean infarct volume was significantly lower in the Epo-treated group (p < 0.003). These results suggest the beneficial neuroprotective effect of Epo in this model of neonatal hypoxic-ischemic brain injury. To our knowledge, this is the first study that demonstrates a protective effect of Epo against hypoxia-ischemia in the developing brain.     

新生大鼠缺氧缺血性脑损伤时细胞凋亡抑制蛋白1的变化

目的探讨在新生大鼠缺氧缺血性脑损伤(HIBD)细胞凋亡抑制蛋白1(cIAPl)的变化。方法建立新生大鼠HIBD动物模型，采用RT-PCR技术检测缺氧缺血(HI)后不同时间点缺血侧脑组织中cIAP1基因表达的变化。结果正常组中有cIAPl基因表达，HI组cIAPl基因表达明显下调，且随HI后时间的延长，cIAPl基因表达在HI后6h开始下调，24h下调最明显，48-72h有所回升。结论脑缺氧缺血引起的神经细胞凋亡可能与clAPl基因表达下调有关。     

缺氧缺血性脑损伤大鼠脑内bcl-2基因表达与细胞凋亡的关系

目的 观察bcl-2基因在缺氧缺血性脑损伤(HIBD)后细胞凋亡中的表达，探讨新生大鼠HmD后神经细胞凋亡的分子机制。方法 54只7日龄新生SD大鼠随机分为1个对照组和8个实验组。给动物吸入含有92％氮气和8％氧气混合气体建立新生大鼠HIBD动物模型，分另q在脑损伤后不同时间点(0．5、1、3、6、12、24、48、72h等)断头处死动物，取海马组织，用免疫组织化学方法检测海马脑神经细胞凋亡及bcl-2基因表达情况。结果 HIBD1h内未见凋亡细胞出现，3h开始出现，并在48h达高峰，之后逐渐下降。bcl-2蛋白阳性细胞在HIBD后立即出现，6h达到高峰，之后渐下降。结论 HIBD后bd-2基因表达在脑神经细胞凋亡过程中起一定作用。在一定时间范围内,bcl-2基因表达与脑神经细胞凋亡呈负相美。     

Combination of systemic hypothermia and N-acetylcysteine attenuates hypoxic-ischemic brain injury in neonatal rats

Hypoxic ischemic (HI) injury in neonates may have devastating, long-term consequences. Recently completed clinical trials in HI neonates indicate that hypothermia within 6 h of birth results in modest improvement in the combined outcome of death or severe disability. The aim of this study was to investigate the effects of combining hypothermia and N-acetylcysteine (NAC) on brain injury, neonatal reflexes and myelination after neonatal HI. Seven-day-old rats were subjected to right common carotid artery ligation and hypoxia (8% oxygen) for 2 h. Systemic hypothermia (30 + 0.5 degrees C) was induced immediately after the period of HI and was maintained for 2 h. NAC (50 mg/kg) was administered by intraperitoneal injection daily until sacrifice. Brain infarct volumes were significantly reduced at 48 h post-HI in the hypothermia plus NAC group (21.5 +/- 3.84 mm3) compared with vehicle (240.85 +/- 4.08 mm3). Neonatal reflexes were also significantly improved by combination therapy at days 1 and 7. There was a significant loss of right hemispheric brain volume in the untreated group at 2 and 4 wk after HI insult. Brain volumes were preserved in hypothermia plus NAC group and were not significantly different when compared with the sham group. Similarly, increased myelin expression was seen in brain sections from hypothermia plus NAC group, when stained for Luxol Fast Blue (LFB), Myelin Basic Protein (MBP) and Proteolipid protein (PLP). These results indicate that hypothermia plus NAC combination therapy improves infarct volume, myelin expression and functional outcomes after focal HI injury.     

Agmatine suppresses nitric oxide production and attenuates hypoxic-ischemic brain injury in neonatal rats

Nitric oxide and excitatory amino acids contribute to hypoxic-ischemic brain injury. Agmatine, an endogenous neurotransmitter or neuromodulator, is an inhibitor of nitric oxide synthase and an antagonist of N-methyl-D-aspartate receptors. Does agmatine reduce brain injury in the rat pup hypoxic-ischemic model? Seven-day old rat pups had right carotid arteries ligated followed by 2.5 h of hypoxia (8% oxygen). Agmatine or vehicle was administered by i.p. injection at 5 min after reoxygenation and once daily thereafter for 3 d. Brain damage was evaluated by weight deficit of the right hemisphere at 22 d after hypoxia by a blinded observer. Agmatine treatments significantly reduced weight loss in the right hemisphere from -30.5 +/- 3.6% in vehicle-treated pups (n = 22) to -15.6 +/- 4.4% in the group treated with 50 mg/kg (n = 18, p < 0.05) and to -15.0 +/- 3.7% in the group treated with 100 mg/kg (n = 18, p < 0.05), but the group treated with 150 mg/kg showed no reduction. Other pups received agmatine or vehicle at 5 min after reoxygenation, and brain biochemistry was assessed. Levels of endogenous brain agmatine rose 2- to 3-fold owing to hypoxic-ischemic (3 h), whereas pups treated with agmatine (100 mg/kg) showed 50-fold higher brain agmatine levels (3 h). Agmatine (100 mg/kg) blocked a hypoxia-induced increase in brain nitric oxide metabolites at 6 h (vehicle-treated, +60.2 +/- 15.2%; agmatine-treated, +4.2 +/- 8.4%; p < 0.05). Agmatine thus reduces brain injury in the neonatal rat hypoxic-ischemic model, probably by blunting the rise in nitric oxide metabolites normally seen after hypoxia.     

新生大鼠脑缺氧缺血对脑组织谷氨酸及环核苷酸的影响

目的　研究新生大鼠缺氧缺血性脑损伤 (HIBD)时谷氨酸 (Glu)和环核苷酸 (cAMP和cGMP)的代谢改变及意义。方法　选 7日龄Wistar新生大鼠 ,随机分为对照组和缺氧缺血组。采取结扎左侧颈总动脉并吸入低氧的混合气体制作HIBD动物模型。用氨基酸自动分析仪与放免法测定不同时间脑组织Glu、cAMP和cGMP含量。结果　脑缺氧缺血后即刻Glu(17.5 6± 5 .32mol/g　P <0 .0 1)和cAMP(15 .0 1± 3.80nmol/g　P<0 .0 5 )较对照组显著升高 ,随后逐渐下降 ,于 6 0～ 180min降至正常 ,cGMP则显著升高 (0 .88± 0 .19nmol/ g　P <0 .0 5 )。结论　新生大鼠HIBD时脑组织Glu、cAMP、cGMP含量发生变化 ,引起细胞兴奋性毒性和代谢改变是加重脑损害的重要因素     

环氧化酶-2在新生大鼠缺氧缺血性脑损伤的表达

目的 探讨环氧化酶-2(COX-2)在新生儿缺氧缺血性脑损伤(HIBD)中的作用.方法 新生7日龄大鼠制成HIBD模型,缺氧时间为2h,RT-PCR半定量分析COX-2mRNA,免疫组化方法测定COX-2蛋白表达,光镜下观察神经元坏死情况.结果 HIBD后6、24、48h,5d COX-2mRNA表达出现不同程度的表达增强,分别为(0.461±0.052),(0.887±0.0816),(0.660±0.119),(0.474±0.104),与对照组(0.321±0.175)比较,差异有统计学意义(P＜0.01),其中HIBD 24 hCOX-2mRNA表达为最高峰,同其余各组相比,差异均有统计学意义(P＜0.01),免疫组化结果与之一致.结论 COX-2在新生儿HIBD形成中发挥一定的作用.     

环氧化酶-2在新生大鼠缺氧缺血性脑损伤的表达

目的探讨环氧化酶-2(COX-2)在新生儿缺氧缺血性脑损伤(HIBD)中的作用。方法新生7日龄大鼠制成HIBD模型,缺氧时间为2h,RT-PCR半定量分析COX-2mRNA,免疫组化方法测定COX-2蛋白表达,光镜下观察神经元坏死情况。结果HIBD后6、24、48h,5dCOX-2mRNA表达出现不同程度的表达增强,分别为(0·461±0·052),(0·887±0·0816),(0·660±0·119),(0·474±0·104),与对照组(0·321±0·175)比较,差异有统计学意义(P<0·01),其中HIBD24hCOX-2mRNA表达为最高峰,同其余各组相比,差异均有统计学意义(P<0·01),免疫组化结果与之一致。结论COX-2在新生儿HIBD形成中发挥一定的作用。     

Timing of neutrophil depletion influences long-term neuroprotection in neonatal rat hypoxic-ischemic brain injury

In neonatal rats, neutrophils do not accumulate in ischemic brain parenchyma to the extent that they do in adult rodents. They are also confined to the intravascular compartment during the first few hours of recovery. However, neonatal rats rendered neutropenic have less brain swelling after a hypoxic-ischemic (HI) insult. In this study, we used the Rice-Vannucci model of HI brain injury in 7-d-old rats, and we depleted neutrophils before injury in one group and 4-8 h after injury in another group to determine 1) whether neutrophils contribute to cerebral atrophy, 2) whether neutropenia induced within 8 h after recovery from HI is neuroprotective, and 3) whether neutropenia preserved energy metabolites during the HI insult. Brain energy metabolites were measured at 0 h and 6 h of recovery. Brain atrophy was measured morphometrically on brain slices at 2 wk of recovery. In 67 rats, we found that neutropenia induced before the HI insult, but not after HI, reduced brain swelling at 42 h of recovery by about 75% (p < 0.001). In another 60 rats, we found that cerebral atrophy was reduced by 61% provided that neutropenia was induced before HI (p < 0.05). Total adenine nucleotides were better preserved in the neutropenic rats at the end of the HI insult (0 h recovery); p < 0.05. We conclude that neutrophils do contribute to vascular dysfunction either during the HI insult or early hours (<4-8 h) of recovery. Antineutrophil strategies initiated after this time are unlikely to be protective in the neonatal rat.     

性别对缺氧缺血性脑损伤新生大鼠脑发育的影响

目的探讨不同性别缺氧缺血性脑损伤(HIBI)新生大鼠脑发育的差异。方法2018年1月1日至12月31日,新生7日龄SD大鼠60只(雌性30只、雄性30只),按随机数字表法将大鼠分为HIBI组40只(雄性20只、雌性20只),对照组20只(雌性10只、雄性10只)。HIBI组大鼠采用Rice-Vannucci方法制作缺氧缺血性脑损伤模型,分离左侧颈总动脉,双重结扎后切断,再置于8%O2和92%N2的混合气体的缺氧箱中90 min;对照组不进行任何处理。缺氧缺血处理后2周,应用步迹分析评价各组大鼠21日龄运动发育功能,头颅磁共振成像(MRI)测量大鼠脑组织的残存脑容量,透射电镜下分析运动区域神经元突触结构的破坏程度。采用方差分析和χ^2检验进行组间统计学比较,同组步长及趾间距比较采用配对t检验。结果HIBI-雄性组大鼠的病死率明显高于HIBI-雌性组[20%(4/20)比10%(2/20),χ^2=40.000,P=0.001];HIBI-雄性组和HIBI-雌性组大鼠脑损伤对侧(右侧)步长及趾间距均明显小于对照组[(7.5±0.3)和(7.9±0.5)比(8.2±0.5)cm,(0.9±0.1)和(1.0±0.0)比(1.1±0.1)cm,F=9.605、71.437,P均<0.01];且HIBI-雄性组均明显小于雌性组(P均<0.01)。HIBI-雄性组和HIBI-雌性组大鼠右侧步长及趾间距均明显小于同组左侧步长[(8.3±0.4)和(8.3±0.5)cm,t=5.289、10.580,P=0.001、0.010]及趾间距[(1.1±0.1)和(1.1±0.1)cm,t=7.953、6.435,P均<0.01]。HIBI-雄性组与HIBI-雌性组的残存脑容量明显小于对照组[(67±4)%和(75±5)%比100%,F=406.122,P<0.01],其中HIBI-雄性组小于HIBI-雌性组(t=-5.281,P<0.01)。HIBI-雄性组和HIBI-雌性组左侧中央前回神经元突触间隙均明显大于对照组[(23.4±1.3)和(19.7±1.6)比(18.9±0.6)nm,F=71.719,P<0.01],其中HIBI-雄性组大于HIBI-雌性组(t=7.645,P<0.01)。结论雄性新生大鼠更易受到HIBI的危害,具有更严重的脑损伤程度及偏瘫症状,对不同性别HIBI患儿应采用不同的更合适的治疗策略。