Production of a de novo cartilage-like tissue construct is a goal for the repair of traumatic chondral defects. We aimed to enhance the matrix synthesis within a scaffold free, de novo cartilage-like tissue construct by way of mechanical load. A novel loading machine that enables the application of shear, as well as compression, was used to subject tissue engineered cartilage-like tissue to mechanical stress. The machine, which applies the load through a roller mechanism, can load up to 20 constructs with four different loading patterns simultaneously. The expression of mRNA encoding matrix products, and subsequent changes in matrix protein content, were analyzed after various loading regimes. The force applied to the immature tissue had a direct bearing on the short-term (first 4 h) response. A load of 0.5 N caused an increase in collagen 11 and aggrecan mRNA within an hour, with a peak at 2 h. This increased mRNA expression was translated into an increase of up to 60% in the glycosaminoglycan content of the optimally loaded constructs after 4 days of intermittent cyclical loading. Introducing pauses between load cycles reproducibly lead to an increase in GAG/DNA. In contrast, constant cyclical load, with no pause, lead to a decrease in the final glycosaminoglycan content compared with unloaded controls. Our data suggest that a protocol of mechanical stimulation, simulating in vivo conditions and involving shear and compression, may be a useful mechanism to enhance the properties of tissue engineered tissue prior to implantation. (c) 2006 Wiley Periodicals, Inc.