Purpose: Cell-based therapies using mesenchymal stem cells from adipose tissue (ASC) showed promising results for the treatment of osteoarthritis (OA), that is a whole joint disease characterized by progressive degeneration of articular cartilage, inflammation of the synovium, and subchondral bone changes. ASC are commonly utilized in various cell therapies and recently in phase I clinical trials to counteract the development of knee osteoarthritis. Administered-ASC in the knee joints enter in close contact with the OA inflammatory milieu and hypoxic environment, which could modulate their characteristics. Little is known about the OA milieu factors that could enhance the migration and tissue specific engraftment of exogenously injected ASC for successful regenerative cell therapy. Therefore, we tested the effect of OA milieu on good manufactured practice (GMP)-ASC, both in normoxic and hypoxic conditions and evaluated their migration properties and cytokine receptors. Methods: We analyzed CXCL8/IL8, CXCL10/IP10, CXCL12/SDF-1, CCL2/MCP1, CCL3/MIP1α, CCL4/MIP1β, CCL5/RANTES and IL6 cytokines in two different OA milieu: OA synovial fluid (SF) and OA-conditioned medium (CM) from synoviocytes. These milieu adequately represent the OA environment for ASC injection and the milieu containing all the soluble factors released by the target tissue of ASC, respectively. The same cytokines were analyzed also on basal GMP-ASC, both in normoxic and hypoxic conditions. Afterwards, GMP-ASC were treated either with OA-SF or OA-CM for 48hours and tested for migration and cytokine receptor expression (CXCR1, CXCR2, CXCR3, CXCR4, CXCR7, CCR1, CCR2, CCR3, CCR5 and IL6R), as well as the cytokine secretion, both in normoxia or hypoxia (1,5%O2). Healthy SF was used as control. Results: In OA-SF we detected higher amount of CXCL10/IP10 than in OA-CM while CCL2/MCP1 and CCL4/MIP1β were higher in OA-CM compared to OA-SF. All the other cytokines were detected in the same amounts in both OA milieu. We demonstrated that GMP-ASC show an increase in proliferation, migration, no changes in the amount of cytokines released and modulation of CXCR1, CXCR3, CCR1 and CCR5 receptors in hypoxic condition. Moreover, GMP-ASC migration increased 15-fold when treated either with OA-SF or OA-CM compared to healthy SF, both in normoxic and hypoxic conditions. GMP-ASC treated in OA milieu showed an increase of CXCR3, CCR3 and IL6R and a decrease of CCR1 receptors. CXCL10/IP10 was the only chemokine of the OA milieu down-modulated after treatment with GMP-ASC. Conclusions: We demonstrated specific effects of OA milieu on both GMP-ASC migration and cytokine receptor expression that were strictly dependent by the inflammatory and hypoxic environment. The use of characterized OA milieu might contributes to define the therapeutic effects of GMP-ASC and indicates that CXCL10/IP10-CXCR3 axis is partially involved in GMP-ASC effect on synovial macrophages.