Effect and mechanism of clazosentan on phenotypic transformation of cerebrovascular smooth muscle cells after subarachnoid hemorrhage in rats

Shishuang JIANG; Zhaohui HE; Xiaohong GOU; Hongxia ZHANG; Xuenong HE

doi:10.16016/j.1000-5404.202103247

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Effect and mechanism of clazosentan on phenotypic transformation of cerebrovascular smooth muscle cells after subarachnoid hemorrhage in rats

Neurosciences | 更新时间：2026-01-09

- Effect and mechanism of clazosentan on phenotypic transformation of cerebrovascular smooth muscle cells after subarachnoid hemorrhage in rats
- Journal of Third Military Medical University Vol. 43, Issue 19, Pages: 1846-1853(2021)
- 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：
- 基金信息：
  
  the General Program of National Natural Science Foundation of China(81870927);the General Program of Natural Science Foundation of Chongqing(CSTC2019jcyj-msxmX0239)
- DOI：10.16016/j.1000-5404.202103247
  CLC： R743.35;R966;R972.9
- Received：29 March 2021，
  
  Revised：2021-05-20，
  
  Published：15 October 2021
- 稿件说明：
移动端阅览
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：

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

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