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Utility of a rotation/revolution-type agitator for chondrocyte isolation during preparation of engineered cartilage

Oda, Akinori, Takamiya, Ryota, Kaneko, Rin, Yoshida, Haruna, Yanagita, Yuta, Sekiguchi, Hatsumi, Nobe, Yoshihito, Muramatsu, Kazuaki
Journal of bioscience and bioengineering 2019 v.128 no.1 pp. 117-122
agitation, agitators, cartilage, cell proliferation, chondrocytes, collagenase, culture dishes, manufacturing, rats, thermolysin, tissue engineering
During the manufacture of cell- and tissue-based products, such as engineered cartilage for autologous chondrocyte implantation, maximizing the number of cells isolated from donor tissue substantially improves the productivity of these products. The method used for agitating tissues with digestive fluid and enzymes can considerably affect both the quality and quantity of isolated cells. This study aimed to investigate the effectiveness of a rotation/revolution-type agitator for chondrocyte isolation following the enzymatic digestion of rat costal cartilage. Cartilage tissue cut into 1 mm3-thick sections was equally divided between two groups and placed in 50-mL conical tubes; sections in both groups were digested using 0.1 mg/mL liberase TH (collagenase/thermolysin) at 37 °C for 4 h with either rotation/revolution or conventional orbital agitation method. Compared with using conventional orbital agitator, using the rotation/revolution-type agitator resulted in a significant (>two-fold) increase in the number of isolated cells. In subsequent primary cultures, chondrocytes obtained by rotation/revolution agitation showed superior initial attachment to tissue culture dish on day 1 and 2 compared with those obtained by conventional agitation; however, no differences in cell proliferation or cartilage-related molecule expression patterns were observed between cells derived from either method after 3 days of subculture. These findings suggested that there are no disadvantages to the proposed rotation/revolution agitation method. Rotation/revolution-type agitators are a promising apparatus for preparing chondrocytes for primary cultures and cartilage tissue engineering.