[Tissue reconstitution by isolated articular chondrocytes in vitro].
It is well known that isolated chondrocytes can form a tissue resembling hyaline cartilage when reaggregated in vitro. Recently, this tissue has been experimentally transplanted as an allograft into the defect of articular cartilage. This phenomenon, however, has not been investigated in detail up to this time. The present study was designed to examine the tissue reformed by isolated articular chondrocytes in comparison with normal articular cartilage. Chondrocytes were released by enzymatic digestion from slices of articular cartilage taken from 8 week old white rabbits. The cells were suspended in culture medium (Ham's F12/10% fetal calf serum, 1% penicillin-streptomycin, 80 micrograms/ml ascorbic acid), and were inoculated in a large number into a 0.28 cm2 stainless cylinder on the Millipore filter. After 12 hours these chondrocytes were layered by gravity onto the Millipore filter and were cultured in the same medium during 7 days. Subsequently the cell aggregate was transferred to organ culture system and then was fed every other day. Aggregates of cells were sampled at 1, 2, 4, 8, 12 weeks in culture for morphological and biochemical studies. The results were obtained as follows: One week in culture, deposition of metachromatic matrix was observed under a light microscope only at the periphery of the aggregate of cells. Matrix formation in the whole aggregate occurred after 2 weeks in culture. The tissue reformed in this culture consisted of metachromatic hyaline cartilage like matrix and chondrocytes within lacunae but for cells at the surface arranged in a tangential flattened layer. The collagen in this tissue was of type II mixed with a very small amount of type I. In view of the above facts, it seemed most reasonable to conclude that the tissue reconstituted in vitro by freshly isolated chondrocytes had characteristics of hyaline cartilage except over the surface. Compared with normal articular cartilage, the cells in this tissue were distributed randomly and the intercellular hyaline matrix was poor under a light microscope, and collagen fibrils in the matrix observed under an electron microscope were much thinner than those of normal articular cartilage. The number of collagen fibrils in the intercellular matrix gradually increased under an electron microscope. Similarly, hydroxyproline content of the tissue increased until 8 weeks in culture. But the maximum value was only approximately a quarter of that of normal articular cartilage of an 8 week-old rabbit. Freshly isolated chondrocytes were thus shown to form hyaline cartilage in this culture method. This method thus provides a tissue culture model of cartilage organization. On the other hand, there were several points of difference between the tissue formed in vitro and normal articular cartilage. Consequently, more investigations are required to use cartilage reformed in vitro as an autologous or allogeneic graft into the defect of articular cartilage.
D000818:Animals; D002358:Cartilage, Articular / Q000166:cytology* / Q000637:transplantation / Q000648:ultrastructure; D002449:Cell Aggregation; D002454:Cell Differentiation; D002478:Cells, Cultured; D003094:Collagen / Q000032:analysis; D011817:Rabbits
D003094:Collagen