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目的探讨重组促红细胞生成素(rEPO)对3日龄脑白质损伤(WMD)新生大鼠的神经保护作用。方法新生3日龄SD大鼠120只随机分为对照组、WMD组和EPO组各40只,对照组仅分离左侧颈总动脉,不结扎不缺氧;EPO组结扎左侧颈总动脉,术后6%氧气+94%氮气混合气中缺氧2.5h,WMD模型建立后立即腹腔注射rEPO5000IU/kg一次;WMD组大鼠造模方法同EPO组,模型建立后腹腔注射等量生理盐水。实验中测量各组大鼠不同时间点体重变化,并于术后24h、生后7天、21天断头取脑,测量脑重量,石蜡包埋切片,行HE染色、髓鞘碱性蛋白(MBP)免疫组化染色,生后30天进行神经行为学检测。结果 WMD组和EPO组大鼠缺氧缺血后均出现行为异常改变,WMD组术后体重(g)增长明显落后于EPO组和对照组[术后24h:(7.2±0.3)比(8.2±0.8)、(9.5±0.4),生后7天:(10.1±0.5)比(13.4±0.5)、(15.3±0.3),生后21天:(30.5±0.4)比(43.7±0.5)、(48.4±0.5),P均<0.05]。HE染色示WMD组术后24h皮质下白质出现不同程度疏松改变,生后21天可见侧脑室扩大;EPO组脑白质疏松明显减轻,侧脑室无明显扩大;WMD组MBP免疫组化染色较EPO组和对照组明显减少,可见髓鞘脱失。WMD组神经行为学与EPO组和对照组比较差异均有统计学意义[悬吊试验(min):(1.8±0.4)比(3.8±0.4)、(4.0±0.2),斜坡试验(s):(9.3±0.8)比(4.4±0.7)、(3.8±0.5),旷场试验(分):(4.8±0.9)比(11.4±2.2)、(11.9±1.8),拒俘反应试验(分):(1.0±0.6)比(3.1±1.0)、(3.4±1.1),P均<0.05]。结论 rEPO可减轻WMD大鼠脑白质损伤,改善少突胶质细胞的成熟障碍及髓鞘发育延迟,促进神经生长修复,提高后期的神经行为表现。
Objective To investigate the neuroprotective effect of recombinant human erythropoietin (rEPO) on neonatal rats with white matter damage (WMD) at 3 days. Methods 120 newborn 3-day-old Sprague-Dawley rats were randomly divided into control group, WMD group and EPO group, 40 rats in each group. Only the left common carotid artery was isolated from the control group and no hypoxia was not ligated. In the EPO group, left common carotid artery was ligated, After hypoxia for 2.5 hours in 6% oxygen + 94% nitrogen gas mixture, rEPO 5000 IU / kg was injected intraperitoneally immediately after the establishment of WMD model. The rats in WMD group were given the same EPO as the modeling method. Body weight changes of rats in different time points were measured in the experiment, and the brain was taken at 24 h, 7 d and 7 d after birth. The brain weight was measured and the paraffin embedded sections were obtained. HE staining, myelin basic protein MBP) immunohistochemical staining, 30 days after birth neurobehavioral testing. Results After hypoxia and ischemia, the behavioral changes of WMD group and EPO group were all changed. The postoperative weight (g) of WMD group lagged behind that of EPO group and control group [7.2 ± 0.3 vs 0.8, 9.5 ± 0.4, and 7 days after birth were (10.1 ± 0.5) vs (13.4 ± 0.5), (15.3 ± 0.3) and 21 days after birth (30.5 ± 0.4) 48.4 ± 0.5), all P <0.05]. Hematoxylin and eosin (HE) staining showed that the subcortical white matter of the WMD group showed varying degrees of looseness 24 hours after operation, and enlarged ventricles were observed 21 days after birth in the WMD group. The leukoaraiosis of the EPO group was significantly reduced and no significant enlargement of the lateral ventricle. The MBP immunohistochemical staining of WMD group was more obvious than that of the EPO group And control group decreased significantly, showing myelin loss. The neurobehavioral changes in WMD group were statistically significant compared with those in EPO group and control group [pendant (1.8 ± 0.4) vs (3.8 ± 0.4), (4.0 ± 0.2), slope test (s) (9.3 ± 0.8) vs (4.4 ± 0.7), (3.8 ± 0.5), open field test (4.8 ± 0.9) vs (11.4 ± 2.2), (11.9 ± 1.8) : (1.0 ± 0.6), (3.1 ± 1.0), (3.4 ± 1.1), P <0.05 respectively. Conclusion rEPO can alleviate the white matter damage of WMD rats, improve the maturation of oligodendrocytes, delay the development of myelin, promote the growth and repair of neurons, and improve the neurobehavioral performance of WMD rats.