诊断学理论与实践

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2019 , Vol. 18 >Issue 1: 56 - 60

DOI: https://doi.org/10.16150/j.1671-2870.2019.01.011

论著

α7烟碱型乙酰胆碱受体激动剂对TGF-β1介导的血管外膜成纤维细胞表型转化影响的体外研究

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  • 1. 上海交通大学医学院附属瑞金医院检验科，上海 200025
    2. 上海市高血压研究所，上海 200025

收稿日期: 2018-09-20

  网络出版日期: 2019-02-25

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Effect of α7 nicotinic acetylcholine receptor activation on transforming growth factor β1-induced phenotypic transformation of adventitia fibroblasts studied in vitro

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  • 1. Department of Clinical Laboratory, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, China
    2. Shanghai Institute of Hypertension, Shanghai 200025, China

Received date: 2018-09-20

  Online published: 2019-02-25

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摘要

目的 探讨α7烟碱型乙酰胆碱受体（α7 nicotinic acetylcholine receptor, α7 nAchR）与血管外膜成纤维细胞（adventitial fibroblasts, AF）表型转化间的关系。方法 选取雄性Sprague-Dawley（SD）大鼠，用组织切块法培养其胸主动脉外膜AF。为评估转化生长因子β1（transforming growth factor-β1, TGF-β1）刺激AF表型转化的作用，将体外培养的AF分为空白对照组A、TGF-β1处理24 h组和TGF-β1处理48 h组3组，处理结束后分别应用实时定量PCR和免疫印迹试验，检测相α7 nAchR基因和蛋白的表达以及α-肌动蛋白（α smooth muscle actin, α-SMA）和Ⅰ型胶原的表达情况。为进一步探讨α7 nAchR的激活能否抑制TGF-β1刺激表型转化及其分子机制，另将体外培养的AF分为空白对照组B、TGF-β1处理48 h组及TGF-β1处理48 h合并α7 nAchR激活剂PNU-282987处理组3组，处理48 h后用免疫印迹法检测α-SMA和Ⅰ型胶原的表达及细胞外调节蛋白激酶1/2（extracellular regulated protein kinases1/2, Erk1/2）磷酸化情况。结果 与空白对照A相比，TGF-β1可使α7 nAchR的基因和蛋白表达明显降低（P<0.05），同时TGF-β1可使α-SMA和Ⅰ型胶原的表达明显增加（P<0.05）。α7 nAchR激动剂PNU-282987可抑制TGF-β1刺激的AF中α-SMA和Ⅰ型胶原的表达，亦可抑制TGF-β1刺激的Erk1/2磷酸化。结论 α7 nAchR的抑制参与TGF-β1介导的血管AF表型转化，其机制可能与Erk1/2磷酸化有关。

关键词： α7烟碱型乙酰胆碱受体; 外膜成纤维细胞; 细胞表型转化; 机制

本文引用格式

魏坚, 高平进, 韩卫青 . α7烟碱型乙酰胆碱受体激动剂对TGF-β1介导的血管外膜成纤维细胞表型转化影响的体外研究[J]. 诊断学理论与实践, 2019 , 18(1) : 56 -60 . DOI: 10.16150/j.1671-2870.2019.01.011

Abstract

Objective: To investigate the correlation between α7 nicotinic acetylcholine receptor (α7 nAchR) and phenotypic transformation of adventitial fibroblasts (AF). Methods: AF were isolated and cultured by using thoracic aorta of male Sprague-Dawley (SD) rats. For evaluating the effect of transforming growth factor-β1 (TGF-β1) on phenotypic transformation of AF, cultured AF were divided into three groups: blank control group A, group with 24 h TGF-β1 treatment, and group with 48 h TGF-β1 treatment. After treatment, real-time quantitative PCR and western blotting were used to detect the mRNA level and protein level of α7 nAchR, and the expression of a smooth muscle actin (α-SMA) and collagen Ⅰ. To further explore whether the activation of α7 nAchR can inhibit TGF-β1-induced phenotypic transformation and the underlying molecular mechanism, AF cultured in vitro were divided into three groups: blank control group B, group with 48 h TGF-β1 treatment, and group with 48 h TGF-β1 +PNU-282987 (an α7 nAchR activator) treatment. After 48 h treatment, the expression of α-SMA and collagen Ⅰ and the phosphorylation of extracellular regulated protein kinase 1/2 (Erk1/2) were detected by immunoblotting. Results: Compared with the blank control group A, TGF-β1 significantly decreased the mRNA level and protein level of α7 nAchR (P<0.05), whereas TGF-β1 induced significant increase of a-SMA and collagen Ⅰ (P<0.05). PNU-282987, an α7 nAchR agonist, inhibited TGF-β1-induced expression of α-SMA and collagen typeⅠ. Meanwhile, PNU-282987 also significantly inhibited TGF-β1-induced Erk1/2 phosphorylation. Conclusions: The inhibition of α7 nAchR is involved in TGF-β1-induced phenotypic transformation in AF, and the mechanism may be related to Erk1/2 phosphorylation.

Key words： α7 nicotinic acetylcholine receptor; Adventitial fibroblasts; Cell phenotype transformation; Mechanism

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