Mitofusin-2在大鼠蛛网膜下腔出血后大脑动脉内表达的变化

doi:10.16098/j.issn.0529-1356.2017.02.004

摘要/Abstract

摘要：

目的研究Mitofusin-2(Mfn2)在大鼠蛛网膜下腔出血（SAH）后大脑动脉中表达的变化，寻找Mfn2基因与脑血管痉挛(CVS)之间的关系。方法蛛网膜下腔出血模型由刺破颈内动脉的颅内动脉分叉处诱导。146只SD大鼠被随机分为6组：假手术组（sham组），SAH后24h、48h、72h、7d 和14d各组。计算动物死亡率，测定神经功能学评分及脑水含量。组织学检测观察形态学变化，采用Western blotting及RT-PCR 分别检测SAH后不同时间点的大鼠主要大脑动脉Mfn2蛋白及mRNA 水平的表达变化。结果围绕在基底动脉周围的血块随着时间逐渐消失，形态学观察显示SAH 24h组基底动脉出现严重的痉挛。SAH7d组的基底动脉中层无阳性的免疫组织化学阳性染色。Western blotting结果显示，Mfn2蛋白表达 sham组与SAH48h组和SAH72h组相比，均显著增加(P<0.05)，SAH 7d出现显著性降低(P<0.05)，SAH14d 恢复至 sham 和SAH24h 组水平。而mRNA 水平变化与蛋白水平变化相类似。Mfn2基因参与到SAH后血管的自我调节过程中，表达随着时间增加而增加，并在72h到达高峰，7d时显著下降，14d左右恢复至sham组水平。结论 Mfn2在SAH后的早期以及迟发型脑血管痉挛中起重要作用，为揭示SAH后脑血管痉挛的机理提供实验依据。

关键词: 脑血管痉挛, Mitofusin-2, 蛛网膜下腔出血, 免疫印迹, 大鼠

Abstract:

Objective To investigate the dynamic changes of mitofusin-2(Mfn2) expression in rat cerebral arteries after experimental subarachnoid hemorrhage (SAH) and to reveal the relationship between Mfn2 and cerebral vasospasm (CVS). Methods A SAH model was induced by endovascular perforation of the intracranial portion of the internal carotid artery. One hundred and forty six male SD rats were randomly divided into six groups: sham group and SAH groups which were sacrificed at different time points (24 hours, 48 hours, 72 hours, 7days and 14days). Mortality, neurobehavioral score and brain water content were measured. Histology was conducted to observe the morphological changes. Western blotting and RT-PCR were performed to measure the Mfn2 protein and mRNA changes of the major cerebral arteries at different time points after SAH. Results Blood clot surrounded the basilar artery gradually dissipated after SAH. HE staining showed that the most severe morphological vasospasm in basilar arteries was observed at the 24th hour after SAH. No positive immunohistochemical staining of Mfn2 was shown in the media layer of basilar artery at the 7th day after SAH. Western blotting showed that Mfn2 protein was remarkably increased at the 48th hour and the 72th hour after SAH compared to sham groups (P<0.05) and significantly decreased at the 7th day after SAH (P<0.05). The protein level at the 14th day after SAH was almost the same level with the sham and SAH 24 hours groups. The mRNA level changed in the same tendency as the protein level. Conclusion This study indicate that Mfn2 plays essential roles in both acute and delayed CVS which may provide a theoretical basis for understanding of the mechanism of the CVS after SAH.

Key words: Cerebral vasospasm, Mitofusin-2, Subarachnoid hemorrhage, Western blotting, Rat

杨晓梅张艳孙娟彭舒晨周长满陈春花. Mitofusin-2在大鼠蛛网膜下腔出血后大脑动脉内表达的变化[J]. 解剖学报, 2017, 48(2): 142-149.

YANG Xiao-mei ZHANG Yan SUN Juan PENG Shu-chen ZHOU Chang-man CHEN Chun-hua. Changes of mitofusin-2 expression after experimental subarachnoid hemorrhage in rats[J]. Acta Anatomica Sinica, 2017, 48(2): 142-149.

参考文献

［1］Rothoerl RD, Ringel F. Molecular mechanisms of cerebral vasospasm following aneurysmal SAH ［J］. Neurol Res, 2007, 29(7): 636-642.

［2］Chen S, Feng H, Sherchan P, et al. Controversies and evolving new mechanisms in subarachnoid hemorrhage ［J］. Prog Neurobiol, 2014, 115:64-91.

［3］Chen S, Wu H, Tang J, et al. Neurovascular events after subarachnoid hemorrhage: focusing on subcellular organelles［J］. Acta Neurochir Suppl, 2015, 120:39-46.

［4］Hansen-Schwartz J, Vajkoczy P, Macdonald RL, et al. Cerebral vasospasm: looking beyond vasoconstriction ［J］. Trends Pharmacol Sci, 2007, 28(6): 252-256.

［5］Zhou C, Yamaguchi M, Kusaka G, et al. Caspase inhibitors prevent endothelial apoptosis and cerebral vasospasm in dog model of experimental subarachnoid hemorrhage ［J］. J Cereb Blood Flow Metab, 2004, 24(4): 419-431.

［6］Borel CO, McKee A, Parra A, et al. Possible role for vascular cell proliferation in cerebral vasospasm after subarachnoid hemorrhage ［J］. Stroke, 2003, 34(2): 427-433.

［7］Chen KH, Guo X, Ma D, et al. Dysregulation of HSG triggers vascular proliferative disorders ［J］. Nat Cell Biol, 2004, 6(9): 872-883.

［8］Zhou C, Yamaguchi M, Colohan AR, et al. Role of p53 and apoptosis in cerebral vasospasm after experimental subarachnoid hemorrhage ［J］. J Cereb Blood Flow Metab, 2005, 25(5): 572-582.

［9］Garcia JH, Wagner S, Liu KF, et al. Neurological deficit and extent of neuronal necrosis attributable to middle cerebral artery occlusion in rats. Statistical validation ［J］. Stroke, 1995, 26(4): 627-634; discussion 635.

［10］Gursoy-Ozdemir Y, Bolay H, Saribas O, et al. Role of endothelial nitric oxide generation and peroxynitrite formation in reperfusion injury after focal cerebral ischemia ［J］. Stroke, 2000, 31(8): 1974-1980; discussion 1981.

［11］Yan J, Chen C, Lei J, et al. 2-methoxyestradiol reduces cerebral vasospasm after 48 hours of experimental subarachnoid hemorrhage in rats ［J］. Exp Neurol, 2006, 202(2): 348-356.

［12］Yang X, Chen C, Hu Q, et al. Gammasecretase inhibitor (GSI1) attenuates morphological cerebral vasospasm in 24 hour after experimental subarachnoid hemorrhage in rats ［J］. Neurosci Lett, 2010, 469(3): 385-390.

［13］Zhao XD, Zhou YT, Zhang X, et al. Expression of NF-E2-related factor 2 (Nrf2) in the basilar artery after experimental subarachnoid hemorrhage in rabbits: a preliminary study ［J］. Brain Res, 2010, 1358: 221-227.

［14］Macdonald RL. Pathophysiology and molecular genetics of vasospasm ［J］. Acta Neurochir, 2001, 77: 7-11.

［15］Verlooy J, Van Reempts J, Haseldonckx M, et al. The course of vasospasm following subarachnoid haemorrhage in rats. A vertebrobasilar angiographic study ［J］. Acta Neurochir (Wien), 1992, 117(1-2): 48-52.

［16］Vorkapic P, Bevan JA, Bevan RD. Longitudinal in vivo and in vitro time-course study of chronic cerebrovasospasm in the rabbit basilar artery ［J］. Neurosurg Rev, 1991, 14(3): 215-219.

［17］Rojo M, Legros F, Chateau D, et al. Membrane topology and mitochondrial targeting of mitofusins, ubiquitous mammalian homologs of the transmembrane GTPase Fzo ［J］. J Cell Sci, 2002, 115(Pt 8): 1663-1674.

［18］Santel A, Fuller MT. Control of mitochondrial morphology by a human mitofusin ［J］. J Cell Sci, 2001, 114(Pt5): 867-874.

［19］Xia Y, Wu Y, He X, et al. Effects of mitofusin-2 gene on cell proliferation and chemotherapy sensitivity of MCF-7 ［J］. Huazhong Univ Sci Technolog Med Sci, 2008, 28(2):185-189.

［20］de Brito OM, Scorrano L. Mitofusin 2: a mitochondria-shaping protein with signaling roles beyond fusion［J］. Antioxid Redox Signal, 2008, 10(3):621-633.

［21］Li PF, Guo YH, Li Q, et al. Adenovirus-mediated gene transfer of rHSG-1 inhibits proliferation of vascular smooth muscle cells from spontaneously hypertensive rats ［J］. Beijing Da Xue Xue Bao, 2004, 36(3): 259-262.

［22］Wang YQ, Brooks G, Harper J, et al. Inhibition of vascular smooth muscle cell proliferation by nonsteroidal antiinflammatory drugs ［J］. Shi Yan Sheng Wu Xue Bao, 2003, 36(2): 85-90.

［23］Malik N, Francis SE, Holt CM, et al. Apoptosis and cell proliferation after porcine coronary angioplasty ［J］. Circulation, 1998, 98(16): 1657-1665.

［24］Kozniewska E, Michalik R, Rafalowska J, et al. Mechanisms of vascular dysfunction after subarachnoid hemorrhage ［J］. J Physiol Pharmacol, 2006, 57 Suppl 11: 145-160.

［25］Song JN, Chen H, Zhang M, et al. Dynamic change in cerebral microcirculation and focal cerebral metabolism in experimental subarachnoid hemorrhage in rabbits ［J］. Metab Brain Dis, 2013, 28(1): 33-43.

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