Updated project metadata. The interaction between monocytes and endothelial cells in inflammation is central to chemoattraction, adhesion, and transendothelial migration. Key players such as selectins and their ligands, integrins and other adhesion molecules and their function in these processes are well studied. Toll-like receptor 2 (TLR2), expressed on monocytes, is critical for sensing invading pathogens and initiating a rapid and effective immune response. However, the extended role of TLR2 in monocyte adhesion and migration has only been partially elucidated. To address this question, we performed several functional cell-based assays using monocyte-like wild type (WT), TLR2-knock-out (KO) and, TLR2-knock-in (KI) THP-1 cells. We found that TLR2 promotes faster and stronger adhesion of monocytes to the endothelium and a more intense endothelial barrier disruption after endothelial activation. In addition, we performed quantitative mass-spectrometry, STRING protein analysis, and RT-qPCR, which revealed the association of TLR2 with specific integrins, but also uncovered novel proteins affected by TLR2. In conclusion, we could show that unstimulated TLR2 affects cell adhesion, endothelial barrier disruption, migration, and actin polymerization.