Mechanical stimulation can modulate the chondrogenic differentiation of stem/progenitor cells and potentially benefit tissue engineering (TE) of functional articular cartilage (AC). Mechanical cues like hydrostatic pressure (HP) are often applied to cell-laden scaffolds, with little optimization of other key parameters (e.g. cell density, biomaterial properties) known to effect lineage commitment. In this study, we first sought to establish cell seeding densities and fibrin concentrations supportive of robust chondrogenesis of human mesenchymal stem cells (hMSCs). High cell densities (15*106 cells/ml) were more supportive of sGAG deposition on a per cell basis, while collagen deposition was higher at lower seeding densities (5*106 cells/ml). Employment of lower fibrin (2.5%) concentration hydrogels supported more robust chondrogenesis of hMSCs, with higher collagen type II and lower collagen type X deposition compared to 5% hydrogels. The application of HP to hMSCs maintained in identified chondro-inductive culture conditions had little effect on overall levels of cartilage-specific matrix production. However, if hMSCs were first temporally primed with TGF-β3 before its withdrawal, they responded to HP by increased sGAG production. The response to HP in higher cell density cultures was also associated with a metabolic shift towards glycolysis, which has been linked with a mature chondrocyte-like phenotype. These results suggest that mechanical stimulation may not be necessary to engineer functional AC grafts using hMSCs if other culture conditions have been optimised. However, such bioreactor systems can potentially be employed to better understand how engineered tissues respond to mechanical loading in vivo once removed from in vitro culture environments.