Rapid and large-scale formation of chondrocyte aggregates by rotational culture.
Chondrocytes in articular cartilage synthesize collagen type II and large sulfated proteoglycans, whereas the same cells cultured in monolayer (2D) dedifferentiate into fibroblastic cells and express collagen type I and small proteoglycans. On the other hand, a pellet culture system was developed as a method for preventing the phenotypic modulation of chondrocytes and promoting the redifferentiation of dedifferentiated ones. Because the pellet culture system forms only one cell aggregate each tube by a centrifugator, the pellet could not be applied to produce a tissue-engineered cartilage. Therefore, we tried to form chondrocyte aggregates by a rotational culture, expecting to form a large number of aggregates at once. In order to increase cell-cell interactions and decrease chondrocyte-material interaction, dishes with low retention of protein adsorption and cell adhesiveness were used. In addition, rotational shaking of the dish including cells was attempted to increase the cell-cell interaction. The shaking speed was set at 80 rpm, so the cells would be distributed in the center of the dish to augment the frequency of cell-cell contact. Under these conditions, bovine articular chondrocytes started aggregating in a few hours. At 24-36 h of rotational culture, aggregates with smooth surfaces were observed. Parameters such as increase of culture time and addition of TGF-beta controlled diameters of the aggregates. There were many fusiform cells at the periphery of the aggregates, where the cells tended to form a multilayered zone in cross sections. In addition, lacune-like structure, which was almost the same as pellet culture, was observed. It was found that the internal structure of the aggregates was similar to that of pellets reported previously. Therefore, the aggregates formed by a rotational culture could become an essential component to make tissue-engineered artificial cartilage.
Furukawa
Katsuko S
KS
Biomedical Engineering Laboratory, Department of Mechanical Engineering, Graduate School of Engineering, School of Engineering, University of Tokyo, Bunkyo-ku, Tokyo, Japan. furukawa@te.t.u-tokyo.ac.jp
Suenaga
Hideyuki
H
Toita
Kenshi
K
Numata
Akiko
A
Tanaka
Junzo
J
Ushida
Takashi
T
Sakai
Yasuyuki
Y
Tateishi
Tetsuya
T
D000818:Animals; D002356:Cartilage / Q000166:cytology; D002417:Cattle; D002449:Cell Aggregation; D002450:Cell Communication; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D002478:Cells, Cultured; D002498:Centrifugation / Q000379:methods*; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism*; D003094:Collagen / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D010641:Phenotype; D012333:RNA, Messenger / Q000378:metabolism; D013997:Time Factors; D023822:Tissue Engineering / Q000379:methods; D016212:Transforming Growth Factor beta / Q000378:metabolism
D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D003094:Collagen