Histochemical staining of the epiphysial growth plate revealed that free hyaluronan (i.e. available to the staining probe) was restricted to the zone of hypertrophy, where it was located in the pericellular space between the chondrocytes and the edge of the lacunae. Furthermore, the amount of hyaluronan staining was directly proportional to the size of the lacunae. Autoradiographic analysis of growth plates cultured with isotopically labeled glucosamine indicated that at least a portion of this hyaluronan was newly synthesized by the hypertrophic chondrocytes. Since hyaluronan can adsorb large amounts of water, it is possible that it exerted a hydrostatic pressure on the surrounding territorial matrix and thereby caused the expansion of hypertrophic lacunae. To assess this possibility, segments of the growth plate were placed in organ culture under different conditions. Under normal culture conditions, a band of hyaluronan staining migrated across the segments coinciding with the enlargement of lacunae in these regions, and the segments, as a whole, increased in size. In contrast, when the segments were cultured in the presence of hyaluronidase, which degraded the pericellular hyaluronan, the lacunae did not undergo enlargement and the overall size of the segments did not increase. These results suggest that the production of hyaluronan contributes to the enlargement of hypertrophic lacunae which is important for determining both the body's stature and proportions.

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