USP7 Stabilizes USF1 to Aggravate ox-LDL-Induced Endothelial Injury Through the MYD88/NF-κB Pathway in Atherosclerosis.

Abstract

Atherosclerosis (AS) is a complex disease that involves the accumulation of lipids in the arterial wall, leading to vessel narrowing and increased risk of heart disease. Upstream stimulatory factor 1 (USF1) is an important regulatory factor that plays an important role in disease progression. Understanding the role and mechanism of USF1 in AS is crucial for unraveling the molecular underpinnings of this condition. Oxidized low-density lipoprotein (Ox-LDL) was used to stimulate human umbilical vein endothelial cells (HUVECs) to induce an AS-like cellular injury. Protein expression was evaluated using western blotting, while mRNA expression was assessed via quantitative real-time polymerase chain reaction. Cell viability and proliferation were analyzed using the cell counting kit-8 and 5-ethynyl-2'-deoxyuridine assays, respectively. Cell apoptosis was examined through flow cytometry. Angiogenic capacity was assessed by tube formation assay in human umbilical vein endothelial cells (HUVECs). Enzyme-linked immunosorbent assays were conducted to measure IL-6 and TNF-α levels, and MDA levels were determined using a lipid peroxidation MDA assay kit. SOD activity was measured using an SOD activity assay kit. Co-immunoprecipitation assay was performed to investigate the association between ubiquitin-specific peptidase 7 (USP7) and USF1, while dual-luciferase reporter assay and chromatin immunoprecipitation assay were conducted to identify the association between USF1 and myeloid differentiation primary response 88 (MYD88). USF1 expression was upregulated in AS patients when compared with healthy volunteers. Knockdown of USF1 protected HUVECs from injury induced by ox-LDL. USP7 was found to stabilize USF1 protein expression by deubiquitination, and its knockdown mitigated ox-LDL-induced HUVEC injury by reducing USF1 protein expression. USF1 was shown to transcriptionally activate MYD88 in HUVECs, and silencing of USF1 protected HUVECs from ox-LDL-induced injury by inhibiting MYD88 expression. Furthermore, USF1 knockdown inactivated the NF-κB pathway by suppressing MYD88 expression. USP7-dependent stabilization of USF1 exacerbated ox-LDL-induced injury in HUVECs by activating the MYD88/NF-κB pathway. These findings underscore the importance of the USP7-USF1-MYD88 axis in AS and suggest potential therapeutic targets for diseases related to AS.