克拉生坦对大鼠蛛网膜下腔出血后脑血管平滑肌细胞表型转化的影响及机制研究

蒋世双; 何朝晖; 苟小红; 张红霞; 贺学农

doi:10.16016/j.1000-5404.202103247

您当前的位置：

首页 >

文章列表页 >

克拉生坦对大鼠蛛网膜下腔出血后脑血管平滑肌细胞表型转化的影响及机制研究

神经科学 | 更新时间：2026-01-09

- 克拉生坦对大鼠蛛网膜下腔出血后脑血管平滑肌细胞表型转化的影响及机制研究
- Effect and mechanism of clazosentan on phenotypic transformation of cerebrovascular smooth muscle cells after subarachnoid hemorrhage in rats
- 第三军医大学学报 2021年43卷第19期页码：1846-1853
- 作者机构：
  
  1.402160 重庆，重庆医科大学附属永川医院: 重症医学科
  2.400016 重庆，重庆医科大学附属第一医院神经外科
  3.402160 重庆，重庆医科大学附属永川医院: 神经外科
- 作者简介：
  
  [贺学农，E-mail: 875791142@qq.com HE Xuenong, E-mail: 875791142@qq.com
- 基金信息：
  
  ‍基‍金‍项‍目‍］‍ ‍国家自然科学基金面上项目(81870927);重庆市自然科学基金面上项目(CSTC2019jcyj-msxmX0239)
- DOI：10.16016/j.1000-5404.202103247
  中图分类号： R743.35;R966;R972.9
- 收稿：2021-03-29，
  
  修回：2021-05-20，
  
  纸质出版：2021-10-15
- 稿件说明：
移动端阅览
蒋世双, 何朝晖, 苟小红, 等. 克拉生坦对大鼠蛛网膜下腔出血后脑血管平滑肌细胞表型转化的影响及机制研究[J]. 第三军医大学学报, 2021,43(19):1846-1853.

Shishuang JIANG, Zhaohui HE, Xiaohong GOU, et al. Effect and mechanism of clazosentan on phenotypic transformation of cerebrovascular smooth muscle cells after subarachnoid hemorrhage in rats[J]. Journal of Third Military Medical University, 2021, 43(19): 1846-1853.
蒋世双, 何朝晖, 苟小红, 等. 克拉生坦对大鼠蛛网膜下腔出血后脑血管平滑肌细胞表型转化的影响及机制研究[J]. 第三军医大学学报, 2021,43(19):1846-1853. DOI： 10.16016/j.1000-5404.202103247.

Shishuang JIANG, Zhaohui HE, Xiaohong GOU, et al. Effect and mechanism of clazosentan on phenotypic transformation of cerebrovascular smooth muscle cells after subarachnoid hemorrhage in rats[J]. Journal of Third Military Medical University, 2021, 43(19): 1846-1853. DOI： 10.16016/j.1000-5404.202103247.

摘要

目的

探究分析克拉生坦(clazosentan

Cla)对大鼠蛛网膜下腔出血(subarachnoid hemorrhage，SAH)后脑血管平滑肌细胞(vascular smooth muscle cells，VSMC)表型转化的影响及作用机制。

方法

采用SD大鼠脑基底动脉及中动脉VSMC构建SAH细胞模型，加入1、5、10 mg克拉生坦干预，Western blot检测空白对照组(Control)、模型组(Model)、克拉生坦低剂量组(L-Cla)、克拉生坦中剂量组(M-Cla)及克拉生坦高剂量组(H-Cla)细胞VSMC表型转化标志蛋白表达；ELISA检测内皮素-1(ET-1)水平；CCK-8检测细胞增殖；划痕实验检测细胞迁移。构建SD大鼠SAH模型，鞘内注射克拉生坦，改良Garcia神经功能缺陷评分评估各组大鼠SAH建模后72 h神经功能状态；HE染色检测基底动脉直径及血管厚度；Western blot检测VSMC表型转化标志蛋白表达；ELISA检测ET-1蛋白表达。

结果

与Control组比较，Model组和各Cla干预组VSMC细胞中α-SMA、SM-22α蛋白表达降低，MGP、OPN蛋白表达升高，细胞ET-1水平、不同时点细胞增殖活性和迁移能力则较高(

均

0.05)；与Model组比较，各Cla干预组VSMC细胞中α-SMA、SM-22α蛋白表达升高，MGP、OPN蛋白表达降低，细胞ET-1水平、不同时间点细胞增殖活性和迁移能力较低(

均

0.05)，且随着克拉生坦剂量的增加，各指标改变更明显。与Control组大鼠比较，Model组及各Cla干预组大鼠Garcia评分升高，基底动脉直径较小，血管壁厚度较厚，α-SMA、SM-22α、MGP、OPN蛋白表达及ET-1水平与体内研究结果一致(

均

0.05)；与Model组大鼠比较，各Cla干预组大鼠Garcia评分较高，基底动脉直径明显较大，血管壁厚度明显较小，分子水平检测结果与体内研究结果一致，随着克拉生坦干预剂量的增加，各项观察指标进一步改善(

均

0.05)。

结论

克拉生坦可通过促进收缩型标记蛋白，抑制合成分泌型标记蛋白表达，影响细胞增殖及迁移活性，抑制血红蛋白诱导的脑VSMC表型转化，同时可通过改善基底动脉病理性血管重塑，抑制SAH模型大鼠VSMC表型转化，促进神经功能损伤修复。

Abstract

Objective

To explore the effect and mechanism of clazosentan (Cla) on phenotype transformation of vascular smooth muscle cells (VSMC) after subarachnoid hemorrhage (SAH) in rats.

Methods

Cell model of SAH was established by VSMC derived from basilar artery and middle artery of SD rats and cultured in the medium containing 10

-6

mol/L oxyhemoglobin from autologous whole blood. Then 1

5 and 10 mg Cla were used to treat the cell model. Western blotting was adopted to detect the expression of phenotype transformation marker proteins of VSMC in control group

model group

l-Cla group

m-Cla group and h-Cla group. The content of endothelin-1 (ET-1) was detected by ELISA. CCK-8 assay and cell scratch test were respectively employed to detect cell proliferation and cell migration. The SAH model was established in SD rats

and Cla was injected intrathecally. The neurological function of rats in each group was evaluated 72 h after SAH modeling by modified Garcia neurological deficit scores. The diameter and thickness of basilar artery were detected by HE staining. Western blotting was used to detect the expression of VSMC phenotype transformation marker proteins. The content of ET-1 protein

was measured by ELISA.

Results

When compared with the VSMC cells of the control group

the cells in the model group and Cla intervention groups had significantly lower expression levels of α-SMA and SM-22α

increased levels of MGP and OPN

and elevated ET-1 content and stronger proliferation activity and migration ability at different time points (

0.05). Opposite phenomena were observed when the Cla intervention groups compared with the model group (

0.05)

and the differences were more significant with the increment of Cla dose. The Garcia scores were higher

diameter of basilar artery was smaller

and thickness of vascular wall was thicker in the model group and the Cla intervention groups of SD rats than the control rats

and the expression trends of α-SMA

SM-22α

MGP

OPN and ET-1 were similar to those in the

in vitro

study (all

0.05). In the rats of the Cla intervention groups

the above indicators showed similar trends as the cell model when compared with the rats of the model group (all

0.05).

Conclusion

Cla can inhibit the phenotypic transformation of brain VSMC induced by hemoglobin by promoting the expression of contractile marker proteins and inhibiting the expression of synthetic secretory marker proteins. It can also inhibit the phenotypic transformation of VSMC in SAH model rats and promote the repair of neural function injury by promoting pathological vascular remodeling of basilar artery.

关键词

Keywords

references

董漪, 郭珍妮, 李琦, 等. 中国脑血管病临床管理指南(节选版): 蛛网膜下腔出血临床管理[J]. 中国卒中杂志, 2019, 14(8): 814-818.

DONG Y, GUO Z N, LI Q, et al . Chinese stroke association guidelines for clinical management of cerebrovascular disorders (excerpts): clinical management of subarachnoid hemorrhage[J ] . Chin J Stroke, 2019, 14(8): 814-818.

LIAO W H, SU Y J, LIU C H. Junctional bradycardia caused by subarachnoid hemorrhage[J]. Interdiscip Neurosurg, 2021, 23: 100986. DOI:10.1016/j.inat.2020.100986.

刘思齐, 张艳, 孙娟, 等. 蛛网膜下腔出血后继发性脑血管痉挛的研究进展[J]. 解剖学报, 2019, 50(4): 537-542. DOI:10.16098/j.issn.0529-1356.2019.04.023.

LIU S Q, ZHANG Y, SUN J, et al . Progress of mechanism and therapy on cerebral vasospasm after subarachnoid hemorrhage[J ] . Acta Anat Sin, 2019, 50(4): 537-542. DOI:10.16098/j.issn.0529-1356.2019.04.023.

肖雪, 侯翠柳, 王筱筱, 等. NKT细胞通过下调PDGFRβ抑制Ang Ⅱ诱导的小鼠血管平滑肌细胞表型转换[J]. 中国病理生理杂志, 2020, 36(8): 1345-1350.

XIAO X, HOU C L, WANG X X, et al . NKT cells inhibit Ang Ⅱ-induced phenotypic transformation of mouse vascular smooth muscle cells by down-regulating PDGFRβ[J ] . Chin J Pathophysiol, 2020, 36(8): 1345-1350.

邓娟, 于晓江, 杨洋, 等. 血管平滑肌细胞表型转化在心血管疾病中作用的研究进展[J]. 生理科学进展, 2020, 51(2): 107-112.

DENG J, YU X J, YANG Y, et al . Research progress of vascular smooth muscle cell phenotypic modulation in cardiovascular diseases[J ] . Prog Physiol Sci, 2020, 51(2): 107-112.

孙慧, 侯玉立, 崔林林, 等. 克拉生坦防治动脉瘤性蛛网膜下腔出血后脑血管痉挛的疗效及安全性的系统评价[J]. 中华临床医师杂志(电子版), 2016, 10(19): 2887-2891.

br > SUN H, HOU Y L, CUI L L, et al . Efficacy and safety of clazosentan for cerebral vasospasm after aneurysmal subarachnoid hemorrhage: a systematic review[J ] . Chin J Clin Electron Ed, 2016, 10(19): 2887-2891.

KONCZALLA J, KASHEFIOLASL S, BRAWANSKI N, et al . Cerebral vasospasm and delayed cerebral infarctions in 225 patients with non-aneurysmal subarachnoid hemorrhage: the underestimated risk of Fisher 3 blood distribution[J ] . J Neurointerv Surg, 2016, 8(12): 1247-1252. DOI:10.1136/neurintsurg-2015-012153.

VARSOS G V, BUDOHOSKI K P, CZOSNYKA M, et al . Cerebral vasospasm affects arterial critical closing pressure[J ] . J Cereb Blood Flow Metab, 2015, 35(2): 285-291. DOI:10.1038/jcbfm.2014.198.

DINC N, QUICK-WELLER J, TRITT S, et al . Vasospasm of the basilar artery following spontaneous SAH-clinical observations and implications for vascular research[J ] . Neurosurg Rev, 2019, 42(4): 983-989. DOI:10.1007/s10143-018-1015-4.

KONCZALLA J, KASHEFIOLASL S, BRAWANSKI N, et al . Cerebral vasospasm-dependent and cerebral vasospasm-independent cerebral infarctions predict outcome after nonaneurysmal subarachnoid hemorrhage: a single-center series with 250 patients[J ] . World Neurosurg, 2017, 106: 861-869.e4. DOI:10.1016/j.wneu.2017.07.017.

KELLER E, KRAYENBVHL N, BJELJAC M, et al . Cerebralvasospasm: results of a structured multimodal treatment[J ] . Acta Neurochir Suppl, 2005, 94: 65-73. DOI:10.1007/3-211-27911-3_11.

HANSEN-SCHWARTZ J. Cerebral vasospasm: a consideration of the various cellular mechanisms involved in the pathophysiology[J]. Neurocrit Care, 2004, 1(2): 235-246. DOI:10.1385/NCC: 1: 2: 235.

HALD E S, ALFORD P W. Smooth muscle phenotype switching in blast traumatic brain injury-induced cerebral vasospasm[J]. Transl Stroke Res, 2014, 5(3): 385-393. DOI:10.1007/s12975-013-0300-3.

CHENG M F, SONG J N, LI D D, et al . The role of rosiglitazone in the proliferation of vascular smooth muscle cells after experimental subarachnoid hemorrhage[J ] . Acta Neurochir(Wien), 2014, 156(11): 2103-2109. DOI:10.1007/s00701-014-2196-4.

ZHANG H R, JIANG L, GUO Z D, et al . PPARβ/δ, a novel regulator for vascular smooth muscle cells phenotypic modulation and vascular remodeling after subarachnoid hemorrhage in rats[J ] . Sci Rep, 2017, 7: 45234. DOI:10.1038/srep45234.

HE J C, LIU M D, LIU Z, et al . Recombinant osteopontin attenuates experimental cerebral vasospasm following subarachnoid hemorrhage in rats through an anti-apoptotic mechanism[J ] . Brain Res, 2015, 1611: 74-83. DOI:10.1016/j.brainres.2015.03.015.

袁瑞军, 唐杨烽, 刘晓红, 等. 转录因子SP1调控血管平滑肌细胞表型转化在主动脉夹层发病中的作用[J]. 国际心血管病杂志, 2016, 43(6): 365-368, 374.

YUAN R J, TANG Y F, LIU X H, et al . The effects of transcription factor SP 1 on phenotypic switching of vascular smooth muscle cells in pathogenesis of aortic dissection[J ] . Int J Cardiovasc Dis, 2016, 43(6): 365-368, 374..

SCHUBERT G A, SCHILLING L, THOMÉC. Clazosentan, an endothelin receptor antagonist, prevents early hypoperfusion during the acute phase of massive experimental subarachnoid hemorrhage: a laser Doppler flowmetry study in rats[J]. J Neurosurg, 2008, 109(6): 1134-1140. DOI:10.3171/JNS.2008.109.12.1134.

MACDONALD R L. Clazosentan: an endothelin receptor antagonist for treatment of vasospasm after subarachnoid hemorrhage[J]. Expert Opin Investig Drugs, 2008, 17(11): 1761-1767. DOI:10.1517/13543784.17.11.1761.

BECK J, RAABE A. Clazosentan: prevention of cerebral vasospasm and the potential to overcome infarction[J]. Acta Neurochir Suppl, 2011, 110(Pt 2): 147-150. DOI:10.1007/978-3-7091-0356-2_26.

KÄSTNER S, OERTEL M F, SCHARBRODT W, et al . Endothelin-1 in plasma, cisternal CSF and microdialysate following aneurysmal SAH[J ] . Acta Neurochir (Wien), 2005, 147(12): 1271-1279;discussion1279. DOI:10.1007/s00701-005-0633-0.

LEI Q, LI S, ZHENG R B, et al . Endothelin-1 expression and alterations of cerebral microcirculation after experimental subarachnoid hemorrhage[J ] . Neuroradiology, 2015, 57(1): 63-70. DOI:10.1007/s00234-014-1435-y.

VERGOUWENM D. Effect of endothelin-receptor antagonists on delayed cerebral ischemia after aneurysmal subarachnoid hemorrhage remains unclear[J]. Stroke, 2009, 40(12): e714. DOI:10.1161/STROKEAHA.109.565887.

PARENT R, LAVALLÉE M. Endothelin-dependent effects limit flow-induced dilation of conductance coronary vessels after blockade of nitric oxide formation in conscious dogs[J]. Cardiovasc Res, 2000, 45(2): 470-477. DOI:10.1016/s0008-6363(99)00366-1.

MACDONALD R L, HIGASHIDA R T, KELLER E, et al . Clazosentan, an endothelin receptor antagonist, in patients with aneurysmal subarachnoid haemorrhage undergoing surgical clipping: a randomised, double-blind, placebo-controlled phase 3 trial (CONSCIOUS-2)[J ] . Lancet Neurol, 2011, 10(7): 618-625. DOI:10.1016/s1474-4422(11)70108-9.

浏览量

下载量

CSCD

文章被引用时，请邮件提醒。

提交

工具集

关联资源

Geminin基因过表达对VSMCs表型转化的影响

动脉瘤性蛛网膜下腔出血后迟发性脑缺血早期识别：一项多因素临床预测模型的建立与验证

血管紧张素Ⅱ-1型受体自身抗体-AT1R-Bmal1轴促进血管平滑肌细胞表型转换和血管纤维化

蛛网膜下腔出血后早期脑损伤中ROS和PDGFR活化的研究

血管平滑肌SCAP过表达抑制自噬促进脂质积聚的机制研究