长时程亚低温通过抑制颅内压反弹促进颅脑创伤大鼠的神经保护作用

doi:10.11958/20231091

天津医药 ›› 2024, Vol. 52 ›› Issue (1): 68-72.doi: 10.11958/20231091

长时程亚低温通过抑制颅内压反弹促进颅脑创伤大鼠的神经保护作用

赵万勇¹(), 李晓红², 王景景³, 孙洪涛³^,^△()

1.山东大学齐鲁医院德州医院神经外科（邮编253000）
2.天津大学医学工程与转化医学研究院
3.武警特色医学中心神经创伤修复研究所，天津市神经创伤修复重点实验室

收稿日期:2023-09-01 出版日期:2024-01-15 发布日期:2024-01-18
通讯作者: ^△E-mail：chenmo333@163.com
作者简介:赵万勇（1989），男，主治医师，主要从事颅脑创伤及功能神经外科疾病方面研究。E-mail：dcrzhaowy@126.com
基金资助:
国家自然科学基金面上项目(32070791)

Long-term mild hypothermia promotes neuroprotection by antagonizing the rebound of intracranial pressure after traumatic brain injury in rats

ZHAO Wanyong¹(), LI Xiaohong², WANG Jingjing³, SUN Hongtao³^,^△()

1. Department of Neurosurgery, Qilu Hospital of Shandong University Dezhou Hospital, Dezhou 253000, China
2. Institute of Medical Engineering and Translational Medicine, Tianjin University
3. Tianjin Key Laboratory of Neurotrauma Repair, Institute of Neurotrauma Repair, Characteristic Medical Center of People’s Armed Police Forces

Received:2023-09-01 Published:2024-01-15 Online:2024-01-18
Contact: ^△E-mail：chenmo333@163.com

赵万勇, 李晓红, 王景景, 孙洪涛. 长时程亚低温通过抑制颅内压反弹促进颅脑创伤大鼠的神经保护作用[J]. 天津医药, 2024, 52(1): 68-72.

ZHAO Wanyong, LI Xiaohong, WANG Jingjing, SUN Hongtao. Long-term mild hypothermia promotes neuroprotection by antagonizing the rebound of intracranial pressure after traumatic brain injury in rats[J]. Tianjin Medical Journal, 2024, 52(1): 68-72.

摘要/Abstract

摘要：

目的探索长时程亚低温（MHT）治疗颅脑创伤（TBI）大鼠的最佳持续时间，并观察其对颅内压（ICP）变化和神经功能恢复的影响。方法 48只健康成年雄性SD大鼠采用随机数字表法分为常温治疗（NT）组、MHT4 h组、MHT24 h组和MHT48 h组，每组12只。制备大鼠TBI模型并植入ICP监测探头。造模完成后NT组给予常温（37 ℃）维持，其余组分别给予低温（33.0±1.0）℃治疗4 h、24 h和48 h。监测各组MHT治疗结束后的ICP并计算脑组织含水量（BWC）；伊文斯兰（EB）染色测定血脑屏障透通性；免疫荧光染色检测5-溴脱氧尿嘧啶核苷（BrdU）、神经元核抗原抗体（NeuN）和白细胞分化抗原86（CD86）表达情况；Western blot法检测B细胞淋巴瘤-2（Bcl-2）、Bcl-2相关X蛋白（Bax）、诱导型一氧化氮合成酶（iNOS）、白细胞介素（IL）-10和精氨酸酶1（Arg-1）蛋白表达情况。结果与NT组相比，MHT4 h组、MHT24 h组和MHT48 h组BWC、ICP、EB水平、CD86阳性细胞数，Bax、iNOS表达水平降低，海马区BrdU阳性细胞数和BrdU/NeuN双标记阳性细胞数增多，Bcl-2、IL-10和Arg-1表达水平升高（P<0.01）；与MHT24 h组比较，MHT48 h组BWC、ICP、EB水平、CD86阳性细胞数，Bax、iNOS表达水平降低，BrdU阳性细胞数和BrdU/NeuN双标记阳性细胞数增多，Bcl-2、IL-10和Arg-1表达水平升高（P<0.01）。结论长时程MHT能够通过抑制ICP反弹，促进神经元的增殖和分化，抑制细胞凋亡和减轻炎症反应，促进TBI后的神经保护作用。

关键词: 颅脑损伤, 颅内压, 低温, 人工, 血脑屏障, 亚低温, 长时程

Abstract:

Objective To explore the optimal duration of long-term mild hypothermia (MHT) for traumatic brain injury (TBI) in rats, and observe its effect on intracranial pressure (ICP) and neurological function. Methods Forty-eight healthy adult male SD rats were divided into the normal temperature treatment (NT) group, the MHT4 h group, the MHT24 h group and the MHT48 h group by random number table method, with twelve rats in each group. The TBI model of rats was prepared by electronic controllable cortical injury device, and ICP monitoring probe was implanted. After modeling, the NT group was treated with normal temperature (37 ℃), and the other groups were treated with low temperature (33.0±1.0 ) ℃ for 4 h, 24 h and 48 h, respectively. ICP was monitored and brain water content (BWC) was calculated after MHT treatment in each group. Blood-brain barrier permeability was determined by Evansland (EB) staining. The expression of 5-bromodeoxyuracil nucleoside (BrdU), neuronal nuclear antigen antibody (NeuN) and leukocyte differentiation antigen 86 (CD86) positive cells were detected by immunofluorescence staining. The expressions of B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax), inducable nitric oxide synthase (iNOS), interleukin (IL) -10 and arginase 1 (Arg-1) were detected by Western blot assay. Results Compared with the NT group, levels of BWC, ICP, EB, and CD86 positive cells, Bax and iNOS expression levels were decreased in the MHT4 h group, the MHT24 h group and the MHT48 h group, and the number of BrdU positive cells and BrdU/NeuN double-labeled positive cells were increased in hippocampus. The expression levels of Bcl-2, IL-10 and Arg-1 were increased (P<0.01). Compared with the MHT24 h group, levels of BWC, ICP and EB, and CD86 positive cells, Bax and iNOS expression were decreased, and the number of BrdU positive cells and BrdU/NeuN double-labeled positive cells were increased in the MHT48 h group, while levels of Bcl-2, IL-10 and Arg-1 expression were increased (P<0.01). Conclusion Long-term MHT can promote the proliferation and differentiation of neurons, inhibit apoptosis and reduce inflammation by suppressing ICP rebound, further promoting neuroprotection after TBI.

Key words: craniocerebral trauma, intracranial pressure, hypothermia, induced, blood-brain barrier, mild hypothermia, long time horizon

中图分类号:

R651.1+5

图/表 9

表1 各组大鼠脑组织BWC和ICP的变化

Tab.1 Changes of water content of brain tissue and ICP in each group （n=3，$\bar{x}±s$）

组别	BWC/%	ICP/mmHg
NT组	90.41±1.51	26.10±0.84
MHT4 h组	86.57±1.85^a	23.40±1.34^a
MHT24 h组	82.07±2.04^ab	18.50±0.92^ab
MHT48 h组	76.42±1.76^ac	11.40±0.98^ac
F	116.000^＊＊	347.900^＊＊

表2 Comparison of BrdU, BrdU/NeuN and CD86 positive cells between the four groups （n=3，个/mm2，$\bar{x}±s$）

Tab.2

组别	BrdU	BrdU/NeuN	CD86
NT组	39.83±5.27	27.33±4.76	81.33±6.95
MHT4 h组	65.50±9.42^a	55.00±7.56^a	59.33±5.65^a
MHT24 h组	53.83±6.08^ab	44.50±4.63^ab	70.83±7.99^ab
MHT48 h组	76.17±8.61^ac	64.50±8.26^ac	48.83±8.23^ac
F	25.690^＊＊	35.860^＊＊	22.460^＊＊

图1 各组大鼠海马区BrdU表达情况（免疫荧光染色，×200）

Fig.1 The BrdU expression in hippocampus in each group (immunofluorescence staining，×200)

图2 MHT24 h组和MHT48 h组大鼠海马区BrdU/NeuN双标记表达情况（免疫荧光染色）

Fig.2 BrdU/NeuN expression double markers in hippocampus in the MHT24 h group and the MHT48 h group (immunofluorescence staining)

图3 各组大鼠CD86表达情况（免疫荧光染色，×200）

Fig.3 The expression of CD86 in each group (immunofluorescence staining，×200)

表3 各组大鼠Bcl-2和Bax表达水平的比较

Tab.3 Comparison of expression levels of Bcl-2 and Bax between the four groups （n=3，$\bar{x}±s$）

组别	Bcl-2	Bax
NT组	0.59±0.07	1.47±0.15
MHT4 h组	1.10±0.15^a	1.08±0.13^a
MHT24 h组	0.80±0.11^ab	0.80±0.10^ab
MHT48 h组	1.34±0.12^ac	0.49±0.15^ac
F	25.230^＊＊	29.820^＊＊

图4 各组大鼠Bcl-2和Bax蛋白表达情况 A：NT组；B：MHT4 h组；C：MHT24 h组；D：MHT48 h组。

Fig.4 The expression of Bcl-2 and Bax protein in each group

表4 各组大鼠iNOS、IL-10和Arg-1表达水平的比较

Tab.4 Comparison of expression levels of iNOS, IL-10 and Arg-1 between the four groups （n=3，$\bar{x}±s$）

组别	iNOS	IL-10	Arg-1
NT组	1.07±0.21	0.35±0.10	0.38±0.07
MHT4 h组	0.64±0.12^a	0.63±0.10^a	0.73±0.10^a
MHT24 h组	0.83±0.13^ab	0.51±0.06^ab	0.59±0.09^ab
MHT48 h组	0.49±0.11^ac	0.82±0.06^ac	0.94±0.06^ac
F	16.980^＊＊	31.720^＊＊	48.480^＊＊

图5 各组大鼠iNOS、IL-10和Arg-1表达情况 A：NT组；B：MHT4 h组；C：MHT24 h组；D：MHT48 h组。

Fig.5 The expression of iNOS, IL-10 and Arg-1 in each group

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长时程亚低温通过抑制颅内压反弹促进颅脑创伤大鼠的神经保护作用

Long-term mild hypothermia promotes neuroprotection by antagonizing the rebound of intracranial pressure after traumatic brain injury in rats

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