Optimization of a chondrogenic medium through the use of factorial design of experiments.

The standard culture system for in vitro cartilage research is based on cells in a three-dimensional micromass culture and a defined medium containing the chondrogenic key growth factor, transforming growth factor (TGF)-β1. The aim of this study was to optimize the medium for chondrocyte micromass culture. Human chondrocytes were cultured in different media formulations, designed with a factorial design of experiments (DoE) approach and based on the standard medium for redifferentiation. The significant factors for the redifferentiation of the chondrocytes were determined and optimized in a two-step process through the use of response surface methodology. TGF-β1, dexamethasone, and glucose were significant factors for differentiating the chondrocytes. Compared to the standard medium, TGF-β1 was increased 30%, dexamethasone reduced 50%, and glucose increased 22%. The potency of the optimized medium was validated in a comparative study against the standard medium. The optimized medium resulted in micromass cultures with increased expression of genes important for the articular chondrocyte phenotype and in cultures with increased glycosaminoglycan/DNA content. Optimizing the standard medium with the efficient DoE method, a new medium that gave better redifferentiation for articular chondrocytes was determined.