摘要
目的探讨亚低温对脑出血大鼠神经再生的作用机制.方法选取165只健康成年SD大鼠为研究对象,依据随机数字表法分为A组(假手术组)、B组(脑出血组)、C组(脑出血+亚低温组).采用胶原酶脑内立体定向注射制作脑出血模型,C组大鼠伤后立即接受33.5℃的亚低温治疗4h.5-溴脱氧尿嘧啶核苷(BrdU)腹腔注射标记有丝分裂细胞,采用Morris水迷宫试验对各组大鼠的学习和记忆能力进行评估,采用神经功能缺损脊髓损伤运动功能(BBB)评分和斜板试验对各组大鼠的运动能力进行评估免疫荧光染色法观察各组大鼠不同时刻损伤侧海马齿状回区BrdU、神经元特异性核蛋白(NeuN)、微管相关蛋白(DCX)、剪切活化半胱氨酸天冬氨酸蛋白酶(Caspase)-3及Caspase-3表达情况.结果 B组的逃避潜伏期(24.36±5.98比9.81 ±2.24)长于A组,平台穿越次数(1.97±0.83比5.09±1.64)、目标象限停留时间(24.87±6.43比41.87±7.65)均短于A组,存在显著认知障碍.C组的逃避潜伏期(9.81±2.24比24.36 ±5.98)短于B组,平台穿越次数(3.54±1.23比1.97±0.83)、目标象限停留时间(31.77 ±4.63比24.87±6.43)均长于B组,差异有统计学意义(F =4.612、6.632、8.353,P<0.05).伤后7d和28 d时B组和C组的神经功能缺损BBB评分[(2.44±0.60)、(13.61 ±0.22)分比(3.95±1.06)、(15.34 ±0.16)分比(20.81±0.23)、(21.02±0.01)分]和斜板试验[(21.79±2.10)°、(28.21±1.25)°比(24.52 ±2.17)°(38.57±2.01)°比(42.48 ±2.22)°、(42.50±1.98)°]均低于A组,且B组低于C组,差异有统计学意义(F=5.837、8.231、4.151、6.389,P<0.05).B组和C组伤后7d和28 d损伤侧海马齿状回区BrdU+细胞数[(32.19±3.64)、(22.11±2.43)个比(44.47 ±5.54)、(58.76±10.08)个比(9.50±3.11)、(5.19±1.72)个]均多于A组,且C组多于B组;伤后28d C组的BrdU+细胞数多于伤后7d,B组的BrdU+细胞数少于伤后7d,差异有统计学意义(F=8.158、6.035,P<0.05).伤后7d时B组和C组损伤侧海马BrdU+DCX+新生神经元细胞数和剪切活化Caspase-3+ NeuN+凋亡神经元细胞数多于于A组,且C组多于B组;B组和C组的Caspase-3+ NeuN+凋亡神经元细胞数少于A组,且C组少于B组,差异有统计学意义(F =4.273、7.276、8.531,P<0.05).结论亚低温可能通过B细胞淋巴瘤/白血病-2(bcl-2)、剪切活化Caspase-3、Fas/FasL途径抑制海马神经元细胞的凋亡,促进海马齿状区内神经元细胞再生及成熟,进而改善认知和运动功能.
Objective To analyze the mechanism of hypothermia on nerve regeneration in rats with cerebral hemorrhage. Methods A total of 165 healthy adult SD rats were selected as the study subjects. They were divided into group A (sham operation group), group B (brain hemorrhage group) and group C (brain hemorrhage + mild hypothermia group) according to the random number table method. The model of cerebral hemorrhage was made by stereotactic injection of collagenase in the brain. The rats in group C were treated with mild hypothermia at 33.5 ℃ for 4 hours immediately after injury. 5-Bromodeoxyuridine (BrdU) was intraperitoneally injected with mitotic cells. Morris water maze test was used to evaluate the learning and memory ability of each group of rats. The nerve function defect Basso-Beattie-Bresnahan (BBB) score and slant plate test were used to treat each group of rats. Exercise capacity was evaluated by immunofluorescence staining to observe BrdU, neuron-specific nuclear protein (NeuN), microtubule-associated protein (DCX), and shear-activated cysteine in the dentate gyrus of hippocampus in different groups of rats at different times. Expression of cysteinyl aspartate-specific protease (Caspase)-3 and Caspase-3. Results The escape latency of group B was longer than that of group A (24.36±5.98 vs. 9.81±2.24), the number of platform crossings (1.97±0.83 vs. 5.09±1.64) and the target quadrant residence time (24.87±6.43 vs. 41.87±7.65) were shorter than those of group A, and there were significant cognitive impairments. The escape latency of group C was shorter than that of group B (9.81±2.24 vs. 24.36±5.98), the number of platform crossings (3.54±1.23 vs. 1.97±0.83) and the target quadrant residence time (31.77±4.63 vs. 24.87±6.43) were longer than those of group B (F=4.612, 6.632, 8.353, P<0.05). BBB scores of neurological deficits in group B and C at 7 and 28 days after damage (2.44±0.60/13.61±0.22 3.95±1.06/15.34±0.16 vs. 20.81±0.23/21.02±0.01) and inclined plate test (21.79±2.10/28.21±1.25 24.52±2.17/38.57±2.01 vs. 42.48±2.22/42.50±1.98), and the difference was = 5.837, 8.231, 4.151, 6, 389, P<0.05). The number of BrdU±cells in the dentate gyrus of hippocampus in group B and C (32.19±3.64/22.11±2.43 44.47±5.54/58.76 ±10.08) was higher than that in group A at 7 and 28 days after injury, and the number of BrdU + cells in group C was higher than that in group B at 7 days after injury, and the number of BrdU + cells in group B was lower than that at 7 days after injury (F=8.158, 6.035, P<0.05). At 7 days after injury, the number of BrdU+ DCX+ neonatal neurons and shear activated Caspase-3+ NeuN+ apoptotic neurons in hippocampus of group B and C was more than that of group A, and the number of Caspase-3±NeuN±apoptotic neurons in group C was less than that in group A, and the number of Caspase-3±NeuN±apoptotic neurons in group C was less than that in group B, with significant differences (F=4.273, 7.276, 8.531, P<0.05). Conclusion Mild hypothermia may inhibit the apoptosis of hippocampal neurons through B cell lymphoma/leukemia-2 (bcl-2), splicing and activation of Caspase-3 and Fas/FasL pathways, promote the regeneration and maturation of neuronal cells in the dentate dentate region, and improve cognitive and motor function.
作者
韩刚
王载忠
杨扬
李力
张磊
Han Gang;Wang Zaizhong;Yang Yang;Li Li;Zhang Lei(Department of Neurology,Zhumadian Central Hospital,Zhumadian 463000,China;ICU,Department of Neurosurgery,Hospital of Cerebrovascular Disease,Henan People’s Hospital,Zhengzhou 450000,China)
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2019年第10期1810-1814,共5页
Chinese Journal of Experimental Surgery
关键词
脑出血
亚低温
海马区
神经再生
Cerebral hemorrhage
Mild hypothermia
Hippocampus
Neural regeneration
