In vitro differentiation of chondrogenic ATDC5 cells is enhanced by culturing on synthetic hydrogels with various charge densities.

We investigated the behavior of chondrogenic ATDC5 cells on synthetic polymer gels with various charge densities: negatively charged poly(2-acrylamido-2-methyl-1-propanesulfonic acid) (PAMPS) gel, neutral poly(dimethylacrylamide) (PDMAAm) gel, and copolymer gels of 2-acrylamido-2-methyl-1-propanesulfonic acid and dimethylacrylamide P(AMPS-co-DMAAm) with different compositions (molar fractions of AMPS, F=0.25, 0.5, 0.75). In insulin-free maintenance medium, the ATDC5 cells cultured on the highly negatively charged gels - PAMPS gel and the P(AMPS-co-DMAAm) copolymer gels (F=0.75) - spread and became confluent at day 7, and interestingly formed nodules at day 14, expressing type II collagen and proteoglycan. This result demonstrates that the highly negatively charged gels can induce chondrogenic differentiation of ATDC5 cells even in insulin-free maintenance medium, in which the ATDC5 cells cultured on the standard polystyrene dish cannot differentiate into chondrocytes. In insulin-supplemented differentiation medium, ATDC5 cells cultured on the PDMAAm gel made focal adhesions, rapidly aggregated and formed large nodules within 7 days, expressing significantly greater levels of type II collagen and proteoglycan than cells cultured on the polystyrene dish and the negatively charged gels. These results showed that the neutral gel accelerated chondrogenic differentiation of ATDC5 cells cultured in the differentiation medium. We suggest that the highly negatively charged PAMPS gel and the neutral PDMAAm gel are interesting biomaterials for cartilage tissue engineering as a scaffold with the potential to induce chondrogenic differentiation.