Deacetylated Sp1 improves β‑glycerophosphate‑induced calcification in vascular smooth muscle cells

Zhang,Zihao; Zhang,Xinyu; Wang,Chengwei; Zhou,Peng; Xiao,Jie; Zheng,Hui; Wang,Lei; Yan,Senbo; Zhang,Yue; Ji,Xiaoping

doi:10.3892/etm.2021.10586

Deacetylated Sp1 improves β‑glycerophosphate‑induced calcification in vascular smooth muscle cells

Authors:
- Zihao Zhang
- Xinyu Zhang
- Chengwei Wang
- Peng Zhou
- Jie Xiao
- Hui Zheng
- Lei Wang
- Senbo Yan
- Yue Zhang
- Xiaoping Ji
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Affiliations: Department of Cardiology, The Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education, Chinese National Health Commission and Chinese Academy of Medical Sciences, The State and Shandong Province Joint Key Laboratory of Translational Cardiovascular Medicine, Qilu Hospital of Shandong University, Jinan, Shandong 250012, P.R. China, Department of Cardiology, State Key Laboratory of Cardiovascular Disease, Fuwai Hospital, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100037, P.R. China
Published online on: August 10, 2021 https://doi.org/10.3892/etm.2021.10586
Article Number: 1152
Copyright: © Zhang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

The aging of the population has led to an annual increase in the incidence of vascular calcification (VC). Specific protein 1 (Sp1) is a transcriptional activator that serves an important role in VC. The deacetylation of transcription factors represses their binding to the promoters of downstream genes, thereby causing their downregulation. The present study aimed to investigate the role of deacetylated Sp1 in the development of VC. In the present study, western blotting and immunoprecipitation (IP) were performed to detect the protein levels of acetylated Sp1. Western blotting and immunofluorescence staining were used to analyze phenotypic switching in vascular smooth muscle cells (VSMCs). Alizarin red S, alkaline phosphatase (ALP) activity and calcium content assays were used to assess calcium deposition in VSMCs. Western blotting, flow cytometry, TUNEL staining and caspase3 activity assay were used to evaluate apoptosis of VSMCs. Chromatin immunoprecipitation (ChIP) assay was used to detect Sp1 binding to the BMP2 promoter. The results indicated that, in a β‑glycerophosphate (β‑GP)‑induced VSMC calcification model, the level of acetylated Sp1 was increased. Western blotting and immunofluorescence staining results showed that, compared with the Sp1 overexpression group (Sp1‑WT), deacetylated Sp1 (Sp1‑K704A) downregulated the expression of osteogenic markers runt‑related transcription factor 2 (Runx2) and bone morphogenetic protein 2 (BMP2), and upregulated the expression of contraction marker α‑smooth muscle actin (α‑SMA) and calponin 1. In addition, deacetylated Sp1 also reduced the ALP activity and calcium content of calcified VSMCs, and the Alizarin red S assay revealed that the calcium crystallization of Sp1‑K704A group was markedly decreased. Western blotting, flow cytometry, TUNEL staining and caspase‑3 activity assay were detected to indicate that the B‑cell lymphoma 2 (Bcl‑2)/Bcl‑2‑associated X protein ratio was increased, and caspase‑3 activity and the apoptotic rate of VSMCs were decreased, in the Sp1‑K704A group, as compared with the Sp1‑WT group. ChIP assay revealed that Sp1 binding to the BMP2 promoter was downregulated in the Sp1‑K704A group, compared with that in theSp1‑WT group. In conclusion, a deacetylated mutant of Sp1 decreased Sp1 binding to the BMP2 promoter, thus decreasing apoptosis, phenotypic switching and calcium deposition in calcified VSMCs. This finding may indicate potential therapeutic targets for VC.

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