Improvements in the treatment of cartilage require insights into the secretory profile of mesenchymal stem cells (MSCs). Apart from their differentiation potential, MSCs secrete a multitude of molecules with therapeutic properties that benefit chondrogenesis and immunomodulation. Previously, we employed a small-panel microarray to demonstrate differences within conditioned medium (CM) of MSCs that were mechanically stimulated within a joint-mimicking bioreactor and their unloaded controls. This study analyzed the proteomic content within CM from 4 week mechanically loaded MSCs with a larger protein microarray. We examined the chondrogenic effect of CM by administration to MSC and chondrocyte pellet cultures, as well as functional changes in T cell proliferation. CM from mechanically loaded samples shows a promising push towards chondrogenic phenotypes within both pellet cultures. Inhibition of T cell proliferation was also observed. This