Effects of extracellular matrix on the morphology and behaviour of rabbit auricular chondrocytes in culture.

Isolated chondrocytes dedifferentiate to a fibroblast-like shape on plastic substrata and proliferate extensively, but rarely form nodules. However, when dissociation is not complete and some cartilage remnants are included in the culture, proliferation decreases and cells grow in a reticular pattern with numerous nodules, which occasionally form small cartilage-like fragments. In an attempt to reproduce this stable chondrogenic state, we added a cartilage protein extract, a sugar extract, and hyaluronan to the medium of previously dedifferentiated chondrocytes. When protein extract was added, many cartilaginous nodules appeared. Hyaluronan produced changes in cell phenotype and behaviour, but not nodule formation. Protein extract has positive effects on the differentiation of previously proliferated chondrocytes and permits nodule formation and the extensive production of type-II collagen. A comparison with incompletely dissociated chondrocyte cultures suggests that the presence of some living cells anchored to their natural extracellular matrix provides some important additional factors for the phenotypical stability of chondrocytes on plastic surfaces. In order to elucidate if it is possible that the incidence of apoptosis is related to the results, we also characterized the molecular traits of apoptosis.

Differential Behavior Between Isolated and Aggregated Rabbit Auricular Chondrocytes on Plastic Surfaces.

A knowledge of the behavior of chondrocytes in culture is relevant for tissue engineering. Chondrocytes dedifferentiate to a fibroblast-like phenotype on plastic surfaces. Dedifferentiation is reversible if these cells are then cultured in suspension. In this report a description is given of how when chondrocyte aggregates formed in suspension are next seeded on plastic, most of them attach as round or polygonal cells. This morphological differentiation, with synthesis of type II collagen, is stable for long culture periods. This simple method can be of use as a model for studies of chondrocyte behavior on plastic. The results indicate that in addition to culture conditions, such as cell isolation method or cell density, chondrocyte behavior on plastic depends on the presence of aggregates.

Delamination of neuroepithelium and nonneural ectoderm and its relation to the convergence step in chick neurulation.

We have analysed the characteristics of the neuroectoderm-nonneural ectoderm meeting point at several axial levels in relation to the mechanics of neurulation in each level. The results show wide differences at cephalic and somitic levels. At cephalic levels, where convergence plays an important role, the delamination process appears at the beginning of the convergence step. This phenomenon produces a major isolation of the basal lamina, forming a space between this structure and the epithelial sheet in whose basal surface a new basal lamina begins to form. This cavity contains abundant extracellular matrix stained with ruthenium red (RR) and tannic acid (TA), and its increase in volume correlates with the progressive convergence of neural folds. At somitic levels, where the convergence is not important, delamination involves the progressive formation of a half-moon-shaped cavity. This structure appears between a dorsal attachment point, in the tip of neuroectodermal wall, and a ventral attachment point which coincides with the point of bending that determines the bilateral furrow, if it exists. In this small cavity, delamination is not related to an isolation of basal lamina. The RR-staining of the extracellular matrix in this cavity is scarce and the volume increase is smaller than in the cephalic region. These results are discussed in terms of neural fold convergence and neural tube closure.

Intracellular distribution of yolk droplets, lipid bodies and Golgi apparatus in the chick neuroepithelial cells during neurulation.

Vitelline and lipidic inclusions which are present in the neuroepithelial cells during chick embryo neurulation show a typical intracellular localization in the apical zone of the cell. In the same cellular zone the Golgi apparatus can be seen during the successive stages of neurulation. These patterns of inclusion and organelle polarity during chick embryo neurulation may be related to active consumption of the reserves contained in inclusions during this morphogenetic process. Such an active consumption would imply a close relationship between the vitelline and lipidic inclusions and the Golgi apparatus. On the other hand, the apical position of the Golgi apparatus in the neuroepithelial cells reveals the remarkable apicobasal polarity of these cells which remains unchanged during chick embryo neurulation.