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Platelet factor 4 (PF4) induces cluster of differentiation 40 (CD40) expression in human aortic endothelial cells (HAECs) through the SIRT1/NF-κB/p65 signaling pathway

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Abstract

PF4 is a pro-atherosclerotic molecule. Endothelial CD40, upon binding to its ligand CD40L, induces endothelial cell (EC) activation, which is a vital pathophysiological process in the initiation and progression of atherosclerosis. However, the relationship between PF4 and endothelial CD40 remains elusive. This study aims to investigate whether and how PF4 affects endothelial CD40 expression using primary HAECs. PF4 treatment down-regulated sirtuin 1 (SIRT1) expression but upregulated the expression of acetylated NF-κB p65 (Ac-p65) and CD40 in HAECs in a concentration- and time-dependent manner. Pretreatment with SIRT1 agonist (SRT1720 or RSV) or SIRT1-overexpressing lentivirus attenuated PF4-induced Ac-p65 and CD40 expression in HAECs, whereas preincubation with SIRT1 antagonist (NAM or EX527) or SIRT1 shRNA had the opposite effect. To investigate whether NF-κB/p65 signaling pathway modulates CD40 expression in PF4-treated HAECs, PDTC, a NF-κB inhibitor, and p65-shRNA were introduced. PDTC or p65-shRNA treatment down-regulated Ac-p65 expression in HAECs. PDTC or p65-shRNA preincubation suppressed CD40 expression in HAECs after PF4 treatment. To better determine whether SIRT1 regulates CD40 expression in PF4-treated HAECs via the NF-κB/p65 signaling pathway, p65-knockdown HAECs were preincubated with SIRT1 agonists before PF4 treatment. SIRT1 agonist preincubation further decreased CD40 expression in p65-knockdown HAECs treated with PF4. Moreover, PF4 treatment promoted p65 nuclear translocation in HAECs. The results of dual luciferase assay demonstrated that four NF-κB binding sites in the promoter of human CD40 gene were activated in PF4-treated HAECs. In conclusion, our findings suggest that PF4 treatment facilitates CD40 expression in HAECs through the SIRT1/NF-κB/p65 pathway.

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Data availability

The data supporting the findings of this study are available from the corresponding author upon reasonable request.

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Acknowledgements

We appreciate the Experimental Research Center of the First Affiliated Hospital of Chongqing Medical University for technical support and the National Natural Science Foundation of China for funding support (No. 8177021049).

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Authors and Affiliations

  1. Department of Vascular Surgery, the First Affiliated Hospital of Chongqing Medical University, No. 1 Youyi Road, Yuanjiagang, Yuzhong District, Chongqing, 400016, China

    Ming Zhong, Xue-Hu Wang & Yu Zhao

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  1. Ming Zhong
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Yu Zhao and Ming Zhong designed the study. Ming Zhong performed the experiments and data analysis with the assistance of Xue-Hu Wang and Yu Zhao. Ming Zhong wrote and Xue-Hu Wang and Yu Zhao reviewed the manuscript.

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Correspondence to Yu Zhao.

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Zhong, M., Wang, XH. & Zhao, Y. Platelet factor 4 (PF4) induces cluster of differentiation 40 (CD40) expression in human aortic endothelial cells (HAECs) through the SIRT1/NF-κB/p65 signaling pathway. In Vitro Cell.Dev.Biol.-Animal 59, 624–635 (2023). https://doi.org/10.1007/s11626-023-00808-9

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