This research aimed to study the effect of microRNA-128-3p (miR-128-3p) on deep venous thrombosis (DVT). The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, Transwell chamber method, and flow cytometry technique were used in the cell experiments. Potential interconnection between miR-128-3p and silent information regulator sirtuin 1 (SIRT1) was revealed by luciferase activity. The concentration of miR-128-3p and mRNA SIRT1 was assessed by quantitative reverse transcription polymerase chain reaction (qRT-PCR). The receiver operating characteristic (ROC) curve was used to test the predictive effect of miR-128-3p in DVT. Decreased miR-128-3p expression was beneficial to cell proliferation and migration and inhibited inflammation, apoptosis, and adhesion of human umbilical vein endothelial cells (HUVECs). The impacts of miR-128-3p on HUVECs were achieved by targeting SIRT1. MiR-128-3p was upregulated in patients with DVT, and it was of great significance in differentiating patients with DVT. Overexpression of miR-128-3p might become a biomarker for patients with DVT.