目的 研究盐酸法舒地尔对蛛网膜下腔出血(SAH)大鼠学习记忆及海马CA1区自噬的影响。方法 雄性Sprague-Dawley大鼠54只，随机分为假手术组、SAH组、药物组，各18只。颈内动脉刺破法造模，药物组造模成功后予盐酸法舒地尔10 mg/kg腹腔注射，每天1次，假手术组和SAH组腹腔注射等量生理盐水，干预后6 h、24 h、72 h予穿梭箱试验；取海马CA1区组织HE染色观察细胞形态，免疫组织化学染色观察Beclin-1和微管相关蛋白l轻链3-Ⅱ(LC3-Ⅱ)的表达。结果 与假手术组相比，SAH组大鼠各时间点穿梭箱试验逃避反应次数减少(P<0.05)，逃避反应时间延长(P<0.05)；海马CA1区神经细胞数目减少(P<0.05)，Beclin-1、LC3-Ⅱ表达增高(P<0.05)。与SAH组相比，药物组各时间点逃避反应次数增多(P<0.05)，逃避反应时间减少(P<0.05)；海马CA1区神经细胞增多(P<0.05)，Beclin-1、LC3-Ⅱ表达进一步增多(P<0.05)。结论 盐酸法舒地尔可改善SAH大鼠学习记忆功能，减少神经细胞损伤，增强海马CA1区自噬。
ObjectiveTo investigate the effects of fasudil hydrochloride on learning and memory, and the autophagy in hippocampal CA1 neurons in subarachnoid hemorrhage (SAH) rats. MethodsFifty-four male Sprague-Dawley rats were randomly divided into sham group (n=18), SAH group (n=18) and drug group (n=18). Subarachnoid hemorrhage model was established with internal carotid artery puncture. The drug group was injected fasudil hydrochloride 10 mg/kg intraperitoneally after modeling per 24 hours, while the sham group and SAH group were injected the same volume of saline. They were tested with shuttle box test 6, 24 and 72 hours after intervention, then the hippocampal CA1 area was stained with HE and immunohistochemistry to observe the morphology of cells and the expression of Beclin-1 and microtubule-associated protein 1 light chain 3 II (LC3-II). ResultsCompared with the sham group, the frequence of avoidance decreased in SAH group at each time point (P<0.05), while the avoidance reaction time increased (P<0.05); the survival of neurons in hippocampal CA1 area decreased (P<0.05), and the expression of Beclin-1 and LC3-II increased (P<0.05). Compared with SAH group, the frequence of avoidance increased in the drug group at each time point (P<0.05), while the avoidance reaction time decreased (P<0.05); the survival of neurons in hippocampal CA1 area increased (P<0.05) and the expression of Beclin-1 and LC3-II increased further (P<0.05). ConclusionFasudil hydrochloride can improve learning and memory ability and protect neurons from damage, which may associate with the excess of autophagy activation in hippocampal CA1 areas in SAH rats.
[1] 周良辅. 现代神经外科学[M]. 2版. 上海:复旦大学出版社, 2015: 1009-1014.
[2] Nishizawa S. The roles of early brain injury in cerebral vasospasm following subarachnoid hemorrhage: from clinical and scientific aspects [J]. Acta Neurochir Suppl, 2013, 115: 207-211.
[3] Washington CW, Derdeyn CP, Dacey RG Jr, et al. Analysis of subarachnoid hemorrhage using the Nationwide Inpatient Sample: the NIS-SAH severity score and outcome measure [J]. J Neurosurg, 2014, 121(2): 482-489.
[4] Sheng R, Zhang LS, Han R, et al. Autophagy activation is associated with neuroprotection in a rat model of focal cerebral ischemic preconditioning [J]. Autophagy, 2010, 6(4): 482-494.
[5] Clark RS, Bayir H, Chu CT, et al. Autophagy is increased in mice after traumatic brain injury and is detectable in human brain after trauma and critical illness [J]. Autophagy, 2008, 4(1): 88-90.
[6] Tamura M, Nakao H, Yoshizaki H. Development of specific Rho-kinase inhibitors and their clinical application [J]. Biochim Biophys Acta, 2005, 1754(1-2): 245-252.
[7] Muroi C, Fujioka M, Marbacher S. Mouse model of subarachnoid hemorrhage: technical note on the filament perforation model [J]. Acta Neurochir Suppl, 2015, 120: 315-320.
[8] 王艮卫,吴楠,牛光明,等. 法舒地尔联合尼莫地平治疗蛛网膜下腔出血后脑血管痉挛的临床研究[J]. 中国临床药理学杂志, 2016, 32(11): 693-695.
[9] 彭智涛,陈建良. Rho激酶抑制剂在神经外科疾病中应用进展[J]. 医药前沿, 2015, 5(26): 131-132.
[10] 徐继伟,赵雅宁,李建民,等. 依达拉奉对SAH大鼠海马区神经细胞自噬的影响[J]. 安徽医科大学学报, 2016, 51(4): 531-535.
[11] 任宝鑫,马云峰,刘殿伟,等. Wnt3a在大鼠蛛网膜下腔出血后早期脑损伤中对神经细胞自噬和凋亡的影响[J]. 山东大学学报, 2016, 56(10): 11-15.
[12] Wan H, AlHarbi BM, Macdonald RL. Mechanisms, treatment and prevention of cellular injury and death from delayed events after aneurysmal subarachnoid hemorrhage [J]. Expert Opin Pharmacother, 2014, 15(2): 231-243.
[13] Chrissobolis S, Sobey CG. Recent evidence for an involvement of Rho-kinase in cerebral vascular disease [J]. Stroke, 2006, 37(8): 2174-2180.
[14] Naraoka M, Munakata A, Matsuda N, et al. Suppression of the Rho/Rho-kinase pathway and prevention of cerebral vasospasm by combination treatment with statin and fasudil after subarachnoid hemorrhage in rabbit [J]. Transl Stroke Res, 2013, 4(3): 368-374.
[15] Mizushima N, Yamamoto A, Matsui M, et al. In vivo analysis of autophagy in response to nutrient starvation using transgenic mice expressing a fluorescent autophagosome marker [J]. Mol Biol Cell, 2004, 15(3): 1101-1111.
[16] Dupont N, Nascimbeni AC, Morel E. Molecular mechanisms of noncanonical autophagy [J]. Int Rev Cell Mol Biol, 2017, 328: 1-23.
[17] 叶红伟,方婷婷,谷小雨,等. Rho激酶和自噬在法舒地尔抗心肌缺血/再灌注损伤中的作用[J]. 南方医科大学学报, 2016, 36(12): 1706-1711.
[18] Gao H, Hou F, Dong R, et al. Rho-kinase inhibitor fasudil suppresses high glucose-induced H9c2 cell apoptosis through activation of autophagy [J]. Cardiovasc Ther, 2016, 34(5): 352-359.
[19] Zhao H, Ji Z, Tang D, et al. Role of autophagy in early brain injury after subarachnoid hemorrhage in rats [J]. Mol Biol Rep, 2013, 40(2): 819-827.
