Impact of Herpud1 in the homocysteine-induced phenotypic switching of vascular smooth muscle cells
10.3760/cma.j.issn.0253-3758.2019.07.010
- VernacularTitle: Herpud1蛋白在同型半胱氨酸诱导的血管平滑肌细胞表型转化中的作用
- Author:
Hui LIN
1
;
Jie ZHANG
2
;
Liping MENG
1
;
Tingjuan NI
3
;
Feidan GAO
4
;
Jufang CHI
1
;
Hangyuan GUO
1
;
Fukang XU
1
Author Information
1. Department of Cardiology, Shaoxing People′s Hospital (Shaoxing Hospital, Zhejiang University School of Medicine), Shaoxing 312000, China
2. First Clinical and Information Engineering College of Wenzhou Medical University, Wenzhou 325000, China
3. Zhejiang University School of Medicine, Hangzhou 310000, China
4. Second Clinical Medical College, Zhejiang University of Traditional Chinese Medicine, Hangzhou 310000, China
- Publication Type:Journal Article
- Keywords:
Blood vessels;
Myocytes, smooth muscle;
Homocysteine;
Herpud1
- From:
Chinese Journal of Cardiology
2019;47(7):561-569
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the impact of homocysteine inducible endoplasmic reticulum(ER) protein with ubiquitin like domain 1 protein (Herpud1) in the homocysteine (Hcy) -induced phenotypic switching of vascular smooth muscle cells (VSMCs).
Methods:VSMCs were derived from thoracic aortic artery of male Sprague Dawley rats and cultured VSMCs (4-7 passage) were treated with various concentrations of Hcy (0, 100, 500 and 1 000 μmol/L) and applied to immunofluorescence to observe the morphological changes of VSMCs via SM-actin staining. Western blot was used to detect the expression of VSMCs phenotypic markers, including Osteopontin, Calponin and smooth muscle myosin heavy chain (SM-MHC) and the expression of endoplasmic reticulum stress (ERS) related proteins, including C/EBP-homologous protein (CHOP), inositol-requiring kinase 1 (IRE-1) and glucose regulating protein 78 (GRP78) in the absence and presence of non-selective inhibitor of ERS, 4-phenylbutyric acid (4-PBA, 2 mg/ml). The Herpud1 mRNA and protein levels were determined in Hcy-stimulated VSMCs treated with 4-PBA or transfected with specific siRNA targeting Herpud1.
Results:Compared with the control group, SM-actin staining results showed that the shape of VSMCs treated with different concentrations of Hcy for 24 hours changed from long fusiform into round form, arrangement of myofilament became irregular and the most significant alteration was found in the 500 μmol/L Hcy group. After intervention of 24 hours, various concentration of Hcy increased protein expression of Osteopontin, and reduced Calponin and SM-MHC protein expressions in VSMCs (all P<0.05). In addition, the results showed that Hcy increased the expression of CHOP, IRE-1 and GRP78 in a dose-dependent manner, which could be reversed by 4-PBA treatment (all P<0.05). However, 4-PBA inhibited Hcy induced upregulation of Osteopontin and downregulation of Calponin and SM-MHC, suggesting that ERS was involved in Hcy-induced phenotypic switching of VSMCs. Herpud1 protein was mostly expressed in the cytoplasm and was also expressed in the nucli, both in the control, Hcy and Hcy+4-PBA groups. Moreover, Hcy increased mRNA and protein levels of Herpud1 (P<0.05), whereas treatment with 4-PBA could significantly reduce Hcy-induced upregulation of Herpud1 (P<0.05). Furthermore, knockdown of Herpud1 abrogated the effects of Hcy on VSMCs phenotype markers.
Conclusion:Herpud1 plays an important role in Hcy-induced phenotypic switching of VSMCs.
