C/EBP α-Mediated Transcriptional Activation of PIK3C2A Regulates Autophagy, Matrix Metalloproteinase Expression, and Phenotypic of Vascular Smooth Muscle Cells in Aortic Dissection

Heng Lu; Yi Chen; Yinhai Chen; Lingchen Huang; Liangwan Chen

doi:10.1155/2022/7465353

C/EBP α-Mediated Transcriptional Activation of PIK3C2A Regulates Autophagy, Matrix Metalloproteinase Expression, and Phenotypic of Vascular Smooth Muscle Cells in Aortic Dissection

J Immunol Res. 2022 Sep 12:2022:7465353. doi: 10.1155/2022/7465353. eCollection 2022.

Authors

Heng Lu^{1

2}, Yi Chen^{1

2}, Yinhai Chen^{1

2}, Lingchen Huang^{1

2}, Liangwan Chen^{1

2}

Affiliations

¹ Department of Cardiovascular Surgery, Fujian Medical University Union Hospital, Fuzhou, China.
² Key Laboratory of Cardio-Thoracic Surgery, Fujian Medical University, Fujian Province University, Fuzhou, China.

Abstract

Purpose: To investigate the function of C/EBPα in the development of aortic dissection (AD) and the underlying mechanism.

Methods: Aortic vascular smooth muscle cells (VSMCs) were isolated, cultured, and identified from AD rats. Then, C/EBPα and PIK3C2A were knockdown or overexpressed by siRNA or plasmid transfection, respectively. Rapamycin or 3-MA was utilized to stimulate and restrain autophagy of VSMCs, respectively. Western blot was used to evaluate the expression levels of C/EBPα, PIK3C2A, LC3, Beclin-1, p62, MMP-2, MMP-9, α-SMA, SM-MHC, and OPN. The pathological status of aortic ring was evaluated by stretch stress, and ChIP assay was used to analyze the binding between C/EBPα and PIK3C2A. C/EBPα shRNA was injected into tail vein to observe the effect of C/EBPα knockdown in vivo on phenotype, autophagy of aortic vascular tissue by immunohistochemical staining and Western blot.

Results: The protein levels of C/EBPα, PIK3C2A, MMP-2, MMP-9, and LC3 in the aorta of AD rats were all upregulated significantly. C/EBPα and rapamycin promoted notable upregulation of the synthesized proteins (OPN), PIK3C2A, matrix metalloproteinases, LC3, and Beclin-1 in VSMCs, while suppressed contractile proteins (α-SMA and SM-MHC) and p62. The opposite results were observed in the C/EBPα-knockdown VSMCs, PIK3C2A-knockdown VSMCs, or VSMCs treated with 3-MA. C/EBPα, PIK3C2A, and LC3 were dramatically upregulated by the stimulation of 3 g and 5 g stretch stress. The downregulated contractile proteins, upregulated synthetic proteins, activated autophagy, and aggravated pathological state in 5 g stretch stress-treated aortic rings were significantly reversed by the knockdown of C/EBPα. ChIP results indicated that there was a binding site for C/EBPα in the promoter of PIK3C2A. C/EBPα also downregulated α-SMA level and upregulated OPN levels in AD rats in vivo.

Conclusion: Our data indicated that during the development of AD, C/EBPα regulated the transition of VSMC phenotype and extracellular matrix remodeling by activating autophagy through regulating the transcriptional activity of PIK3C2A promoter.

MeSH terms

Animals
Aortic Dissection* / genetics
Autophagy / genetics
Beclin-1 / genetics
Beclin-1 / metabolism
Beclin-1 / pharmacology
CCAAT-Enhancer-Binding Protein-alpha / genetics
CCAAT-Enhancer-Binding Protein-alpha / metabolism
CCAAT-Enhancer-Binding Protein-alpha / pharmacology
Cells, Cultured
Matrix Metalloproteinase 2 / metabolism
Matrix Metalloproteinase 9 / genetics
Matrix Metalloproteinase 9 / metabolism
Muscle, Smooth, Vascular* / metabolism
Muscle, Smooth, Vascular* / pathology
Phenotype
Phosphatidylinositol-4-Phosphate 3-Kinase / metabolism*
RNA, Small Interfering / genetics
RNA, Small Interfering / metabolism
Rats
Sirolimus / pharmacology
Transcriptional Activation

Substances

Beclin-1
CCAAT-Enhancer-Binding Protein-alpha
RNA, Small Interfering
PIK3C2A protein, rat
Phosphatidylinositol-4-Phosphate 3-Kinase
Matrix Metalloproteinase 2
Matrix Metalloproteinase 9
Sirolimus