Endometrial mesenchymal stem cells (eMSCs) are a novel and biologically potent source of multipotent stromal cells with potential beyond reproductive medicine. This study explored their phenotypic profile, trilineage differentiation, and the cytoprotective effects of their conditioned media (eMSCCM) on oxidatively stressed neonatal and adult chondrocytes. Canine eMSCs displayed typical fibroblast-like morphology and expressed high levels of mesenchymal surface markers CD29 and CD44, low hematopoietic markers CD34/CD45, and variable CD90, confirming a mesenchymal identity. Differentiation assays revealed osteogenic and chondrogenic differentiation, whereas adipogenic activity was limited. Using eMSCCM at 25% and 50% concentrations, chondrocyte viability was assessed after exposure to 200 µM HO. eMSCCM significantly enhanced the viability of HO-stressed chondrocytes in a dose-dependent manner, particularly at 50%, with marked effects at 24 and 48 h. Although metabolic activity declined at 72 h, the treated cells remained more metabolically active than untreated controls. These findings suggest that eMSCCM offers promising cytoprotective effects for cartilage-related oxidative stress conditions.