Hippo信号通路在颈动脉结扎所致大鼠动脉血管重构中的表达及其意义

doi:10.12047/j.cjap.5717.2019.006

摘要/Abstract

摘要： 目的:在建立大鼠血管重构模型基础上探讨Hippo信号通路在该模型中的表达及意义。方法:模型组(n=40)经颈部正中切口游离出左侧颈总动脉,用6-0不可吸收线在尽量靠近近心端处结扎,完全阻断血流。对照组(n=20)仅将手术线穿过颈总动脉而不结扎,闭合切合。14 d后处死所有动物,经原手术路径分离颈总动脉,收集结扎处至远心端的动脉。用HE以及MASSON染色观察血管形态以及纤维化,免疫组织化学染色法检测颈动脉中α-肌动蛋白(α-MSA)和增殖细胞核抗原(PCNA)的表达,Western blot检测yes相关蛋白(YAP),PDZ结合基序的转录辅激活子(TAZ),TEAD1,Bax,Bcl-2的表达。结果:与对照组相比,造模组HE染色提示血管重构明显,新生内膜/中膜比例明显增加,MASSON染色提示纤维化明显增加;免疫组织化学染色法提示造模组血管α-MSA及PCNA表达明显增加;Western blot提示造模组血管YAP,TAZ,TEAD1,Bcl-2表达增加,而Bax表达降低,Bax/Bcl-2蛋白比例明显降低。结论:本研究成功建立颈动脉结扎介导的大鼠血管重构模型,另外证明Hippo信号通路在颈动脉结扎介导的大鼠血管重构模型中明显激活,以及可能介导增殖、凋亡相关的Bax/Bcl-2比值的改变,进而参与平滑肌细胞增殖促进血管重构。

关键词: 大鼠, 颈动脉结扎, 血管重构, Hippo信号通路

Abstract: Objective:To establish an arterial remodeling model of rats and to investigate the expression and role of Hippo signaling pathway in this model. Methods: In the model group (n=40), the left common carotid artery was removed through the median incision of the neck. The 6-0 non-absorbable line was used to ligate the carotid artery near the proximal end as far as possible, completely blocking the blood flow. The common carotid artery of rats in control group (n=20) was not ligated using the operative line. After 14 days, the animals were sacrificed and the common carotid arteries were separated through the original surgical pathway and the arteries from the ligature to the distal end were collected. Arterial morphology and fibrosis were observed by HE and MASSON staining. Immunohistochemical staining was used to detect the expressions of anti-α smooth muscle actin (α-MSA) and proliferating cell nuclear antigen (PCNA) in the carotid artery. Western blot was used to detect the expressions of yes associated protein (YAP), transcriptional coactivator with PDZ-binding motif (TAZ), TEAD1, Bcl-2-like protein 4 (Bax), and B-cell lymphoma-2 (Bcl-2). Results: Compared with the control group, the HE staining showed that the vascular remodeling was obvious, the ratio of the neointima/middle membrane was increased significantly, and the MASSON staining indicated that the fibrosis was significantly increased in model group. The immunohistochemical staining suggested that the expressions of α-SMA and PCNA were increased significantly; Western blot suggested that the expressions of YAP, TAZ, TEAD1, and Bcl-2 were increased in carotid artery of the model group. While the expression of Bax and the ratio of Bax/Bcl-2 were decreased. Conclusion: A rat model of arterial remodeling mediated by carotid artery ligation was established successfully in this study. Hippo signaling pathway was proved to be activated in the arterial remodeling model induced by carotid artery ligation in rats, and might regulate the change of Bax/Bcl-2 ratio related to proliferation and apoptosis, and subsequently involved in the proliferation of smooth muscle cells to promote vascular remodeling.

Key words: rat, carotid artery ligation, vascular remodeling, Hippo signaling pathway

朱宁, 陈皓, 赵旭勇, 叶凡豪, 王毅. Hippo信号通路在颈动脉结扎所致大鼠动脉血管重构中的表达及其意义[J]. 中国应用生理学杂志, 2019, 35(1): 23-27.

ZHU Ning, CHEN Hao, ZHAO Xu-yong, YE Fan-hao, WANG Yi. The expression and meaning of Hippo signaling pathway in carotid artery ligation induced arterial remodeling model of rats[J]. CJAP, 2019, 35(1): 23-27.

参考文献

[1] Assimes TL, Roberts R. Genetics: implications for prevention and management of coronary artery disease[J]. J Am Coll Cardiol, 2016, 68(25): 2797-2818.
[2] Meraj PM, Jauhar R, Singh A. Bare metal stents versus drug eluting stents: where do we stand in 2015[J]? Curr Treat Options Cardiovasc Med, 2015, 17(8): 393.
[3] Stone GW, Ellis SG, Cannon L, et al. Comparison of a polymer-based paclitaxel-eluting stent with a bare metal stent in patients with complex coronary artery disease: a randomized controlled trial[J]. JAMA, 2005, 294(10): 1215-1223.
[4] 尹扬光, 黄岚, 赵晓辉, 等. 非显微外科手术建立小鼠颈动脉损伤模型[J]. 中国应用生理学杂志, 2008, 24(1): 34-35.
[5] Plouffe SW, Hong AW, Guan KL. Disease implications of the Hippo/YAP pathway[J]. Trends Mol Med, 2015, 21(4): 212-222.
[6] Stone GW, Midei M, Newman W, et al. Comparison of an everolimus-eluting stent and a paclitaxel-eluting stent in patients with coronary artery disease[J]. JAMA, 2008, 299(16): 1903-1913.
[7] Indolfi C, De Rosa S, Colombo A. Bioresorbable vascular scaffolds-basic concepts and clinical outcome[J]. Nat Rev Cardiol, 2016, 13(12): 719-729.
[8] Li P, Zhu N, Yi B, et al. MicroRNA-663 regulates human vascular smooth muscle cell phenotypic switch and vascular neointimal formation[J]. Circ Res, 2013, 113(10): 1117-1127.
[9] Alexander MR, Owens GK. Epigenetic control of smooth muscle cell differentiation and phenotypic switching in vascular development anddisease[J]. Annu Rev Physiol, 2012, 74(1): 13-40.
[10]Xie C, Guo Y, Zhu T, et al. Yap1 protein regulates vascular smooth muscle cell phenotypic switch by interaction with myocardin[J]. J Biol Chem, 2012, 287(18): 14598-14605.
[11]周昌钻, 潘孙雷, 林辉, 等. 瑞舒伐他汀对同型半胱氨酸诱导的小鼠血管平滑肌细胞去分化和内质网应激的影响及其机制研究[J]. 中国应用生理学杂志, 2018, 34(1): 43-48.
[12]朱宁, 赵旭勇, 向贻佳, 等. 大鼠肺细小动脉平滑肌细胞培养新方法的建立及血小板衍生因子对其增殖迁移的影响[J]. 中国应用生理学杂志, 2016, 32(4): 343-346.
[13]Davis-Dusenbery BN, Wu C, Hata A. Micromanaging vascular smooth muscle cell differentiation and phenotypic modulation[J]. Arterioscler Thromb Vasc Biol, 2011, 31(11): 2370-2377.
[14]Yu FX, Zhao B, Guan KL. Hippo pathway in organ size control, tissue homeostasis, and cancer[J]. Cell, 2015, 163(4): 811-828.
[15]Ono H, Ichiki T, Ohtsubo H, et al. Critical role of Mst1 in vascular remodeling after injury[J]. Arterioscler Thromb Vasc Biol, 2005, 25(9): 1871-1876.
[16]Kimura TE, Duggirala A, Smith MC, et al. The Hippo pathway mediates inhibition of vascular smooth muscle cell proliferation by cAMP[J]. J Mol Cell Cardiol, 2016, 90: 1-10.
[17]Zhao B, Ye X, Yu J, et al. TEAD mediates YAP-dependent gene induction and growthcontrol[J]. Genes Dev, 2008, 22(14): 1962-1971.