促红细胞生成素对2日龄未成熟大鼠缺血性脑白质损伤后学习记忆能力的影响

发布时间：2018-09-17 12:23

【摘要】：目的：建立未成熟大鼠脑白质损伤（white matter damage,WMD）动物模型，观察促红细胞生成素（Erythropoietin，EPO）对2日龄缺血性脑白质损伤大鼠脑细胞髓鞘碱性蛋白（MBP）表达及远期学习记忆能力的影响，探讨EPO对未成熟大鼠脑白质损伤后神经保护作用的可能机制。方法：将2日龄未成熟新生大鼠随机分为正常对照组即假手术组（Sham，n=36）、模型组(BCAO，n=36)、EPO治疗组(EPO，n=36)。采用双侧颈总动脉结扎术（Bilateral carotid artery occlusion，BCAO）建立缺血性脑白质损伤动物模型。对照组只分离双侧颈总动脉，但不结扎。EPO治疗组于BCAO后即刻给予腹腔注射EPO（5000u/Kg），对照组、BCAO组注射等体积生理盐水。三组大鼠于术后24h、48h、72h和7d、14d、35d分别测量体重、脑重，HE染色观察脑组织病理形态学改变，免疫组化染色检测脑组织MBP蛋白的表达。术后72h采用干湿法测定脑含水量；术后35天采用Morris水迷宫试验、跳台试验、闭暗试验进行神经行为学检测，观察EPO治疗对脑白质损伤未成熟新生大鼠体重、脑重、脑含水量、早期神经行为、脑组织病理形态学、学习记忆能力及MBP蛋白表达的影响。结果：（1）模型组大鼠体重、脑重增长不良，各时间点EPO治疗组大鼠体重、脑重增长明显高于模型组，与模型组比较差异有显著性（P＜0.05）；各时间点EPO治疗组大鼠体重、脑重与假手术组无显著性差异（P＞0.05）。（2）模型组脑组织含水量明显高于假手术组（P＜0.05），EPO治疗后脑组织含水量明显低于模型组（P＜0.05）。（3）HE染色模型组大鼠出现脑室周围白质损伤，光镜下表现为脑室周围白质细胞肿胀、囊性坏死、组织疏松；EPO治疗组脑白质细胞肿胀、囊性坏死等病理形态学改变较模型组减轻，具有凋亡特征的神经细胞明显减少。假手术组脑组织未见异常。（4）术后24h、48h、72h三组大鼠MBP平均积分光密度比较差异无显著性意义（P＞0.05）；术后7d模型组大鼠MBP积分光密度与对照组、治疗组比较均明显降低，差异有显著性意义（P＜0.05）；EPO治疗后MBP积分光密度较模型组增高，与假手术组比较差异无显著性（P＞0.05）；术后14d、35d差异更为显著。（5）35日龄时，模型组大鼠自主活动减少，反应迟钝，灵活性降低，Morris水迷宫试验、闭暗试验、跳台试验的结果均差于对照组和治疗组，差异有显著性（P＜0.05）。EPO治疗后大鼠学习、记忆能力明显改善。结论：成功建立了2日龄未成熟新生大鼠脑白质损伤动物模型；在此基础上，应用EPO干预治疗脑白质损伤未成熟新生大鼠，可明显减轻脑白质损伤大鼠脑水肿，促进体重、脑重增长，降低早期神经行为异常发生率，减少细胞凋亡，，进一步证实外源性EPO可提高大鼠学习、记忆能力，推测可能与促进未成熟新生大鼠脑白质MBP蛋白表达，减少少突胶质细胞凋亡有关，为临床应用EPO治疗早产儿脑损伤提供了实验依据。
[Abstract]:Objective:
To investigate the effects of erythropoietin (EPO) on the expression of myelin basic protein (MBP) and long-term learning and memory ability in brain cells of 2-day-old rats with ischemic brain white matter damage (WMD), and to explore the neuroprotective effect of EPO on immature rats after WMD. The possible mechanism.
Method:
Two-day-old immature neonatal rats were randomly divided into sham operation group (Sham, n = 36), model group (BCAO, n = 36) and EPO treatment group (EPO, n = 36). Bilateral carotid artery occlusion (BCAO) was used to establish an animal model of ischemic white matter injury. EPO (5000u/Kg) was injected into the abdominal cavity immediately after BCAO in the treatment group, and the same volume of normal saline was injected into the control group and the BCAO group. The body weight and brain weight were measured 24 hours, 48 hours, 72 hours, 7 days, 14 days and 35 days after BCAO. The pathological changes of brain tissue were observed by HE staining, and the expression of MBP protein was detected by immunohistochemical staining. The body weight, brain weight, brain water content, early neurobehavioral changes, histopathology, learning and memory ability and MBP protein expression in immature neonatal rats with white matter injury were observed by Morris water maze test, step-down test and dark test 35 days after operation.
Result:
(1) The weight and brain weight of rats in model group increased badly. The weight and brain weight of rats in EPO treatment group were significantly higher than those in model group at each time point (P < 0.05), and there was no significant difference between EPO treatment group and sham operation group at each time point (P > 0.05).
(2) The water content of brain tissue in model group was significantly higher than that in sham operation group (P < 0.05). The water content of brain tissue after EPO treatment was significantly lower than that in model group (P < 0.05).
(3) HE staining model group showed periventricular white matter injury, light microscopy showed that periventricular white matter cells swelling, cystic necrosis, tissue loosening; EPO treatment group white matter cells swelling, cystic necrosis and other pathological changes than the model group alleviated, neurons with apoptotic characteristics were significantly reduced. Often.
(4) There was no significant difference in the mean integral optical density of MBP among the three groups 24 hours, 48 hours and 72 hours after operation (P > 0.05); the integral optical density of MBP in the model group was significantly lower than that in the control group 7 days after operation (P < 0.05); the integral optical density of MBP after EPO treatment was higher than that in the model group, and the difference was significant (P < 0.05). There was no significant difference (P > 0.05), and the difference between 14d and 35d was more significant.
(5) At the age of 35 days, the autonomic activity of rats in the model group decreased, the response was slow and the flexibility was reduced. The results of Morris water maze test, dark test and step-down test were worse than those in the control group and the treatment group, and the difference was significant (P < 0.05). After EPO treatment, the learning and memory abilities of rats were significantly improved.
Conclusion:
The animal model of brain white matter injury in 2-day-old immature neonatal rats was successfully established. On this basis, EPO intervention in the treatment of brain white matter injury in immature neonatal rats can significantly reduce brain edema, promote body weight, brain weight growth, reduce the incidence of early neurobehavioral abnormalities, reduce cell apoptosis, and further confirm the exogenous. Sexual EPO can improve the learning and memory abilities of rats, which may be related to the promotion of MBP protein expression in the white matter of immature neonatal rats and the reduction of oligodendrocyte apoptosis, providing experimental basis for clinical application of EPO in the treatment of brain injury of premature infants.
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R722.1

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