Angiogenesis, the formation of new blood vessels from pre-existing vasculature, is essential for physiological processes such as development and wound healing, but its dysregulation contributes to a range of pathological conditions including cancer, diabetic retinopathy, and chronic inflammation. In recent years, marine-derived compounds have emerged as promising multitarget agents with anti-angiogenic potential. Posidonia oceanica, a Mediterranean seagrass traditionally used in folk medicine, is increasingly recognized for its pharmacological properties, including antioxidant, anti-inflammatory, and anti-invasive activities. This study investigated the effects of a hydroethanolic extract from P. oceanica leaves (POE) on human Endothelial Colony-Forming Cells (ECFCs), a subpopulation of endothelial progenitor cells with high proliferative and vessel-forming capacity, and a relevant model for studying pathological angiogenesis. ECFCs were treated with POE (4–8 µg/mL), and cell viability, morphology, migration, invasion, tube formation, oxidative stress, and activation markers were evaluated. POE did not alter ECFC morphology or viability, as confirmed by Trypan Blue and MTT assays. However, functional assays revealed that POE significantly impaired ECFC migration, invasion, and in vitro angiogenesis in a dose-dependent manner. Under VEGF (Vascular endothelial growth factor) stimulation, POE reduced intracellular ROS accumulation and downregulated key redox-regulating genes (hTRX1, hTRX2, PRDX2, AKR1C1, AKR1B10). Western blot analysis showed that POE inhibited VEGF-induced phosphorylation of KDR, mTOR and p-ERK, while p-AKT remained elevated, indicating selective disruption of VEGF downstream signaling. Furthermore, POE reduced the expression of pro-inflammatory and pro-coagulant markers (VCAM-1, ICAM-1, TF) and partially reversed TNF-α–induced endothelial activation. These findings suggest that POE exerts anti-angiogenic effects through a multitargeted mechanism, supporting its potential as a natural therapeutic agent for diseases characterized by aberrant angiogenesis.