The purpose of this study was to investigate the effect and mechanism of silencing long non-coding RNA (LncRNA) plasmacytoma variant translocation 1 (PVT1) on high-fat-induced vascular endothelial cell injury and endothelial dysfunction. Human umbilical vein endothelial cells (human umbilical vein endothelial cells, HUVECs) were divided into four groups, including control group, high-fat group, high-fat + si-NC group, and high-fat + si-PVT1 group. The expression level of PVT1 after transfection, the proliferation ability of cells and the level of reactive oxygen species (ROS) were detected by RT-qPCR, CCK-8 and wide-field high-content imaging, respectively. The levels of interleukin-6 (IL-6), superoxide dismutase (SOD), malondialdehyde (MDA), and endothelin-1 (ET-1) in cells were detected by ELISA. The expression levels of phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT), and nuclear factor κB (NF-κB) were detected by Western blot. The results showed that the expression level of PVT1 was significantly decreased, and the proliferation activity of HUVECs was significantly increased after silencing PVT1. Compared with the control group, the levels of ROS, MDA, IL-6, and ET-1 were significantly increased as well as the expression of PI3K, AKT, and NF-κB, and the SOD level was significantly decreased in the high-fat group. Compared with the high-fat + si-NC group, the contents of ROS, MDA, IL-6, and ET-1 were significantly decreased as well as the expression of PI3K, AKT, and NF-κB , and the content of SOD was significantly increased in the high-fat + si-PVT1 group. This study shows that inhibiting the expression of PVT1 can significantly inhibit the inflammatory response , reduce oxidative stress, and improve endothelial dysfunction induced by high fat. These results provided a scientific basis for LncRNA PVT1 as an important detection and treatment target for atherosclerosis and related diseases.