It is proposed that the mode of interaction of hyaluronate (HA) with the cell surface changes during differentiation of embryonic limb mesodermal cells and that this modulation is important in the pathways of differentiation of the different cell populations within the limb. Specifically, it is proposed: i) that HA is bound to the early mesodermal cell by a non-receptor mediated mechanism; ii) that, at the time of prechondrogenic cell condensation in the limb core, HA receptors appear which now retain HA at the cell surface; iii) that these receptors mediate internalization of most of the mesodermal HA en route to its degradation by lysosomal hyaluronidase; iv) that the newly differentiating chondrocytes utilize the residual receptor-bound HA to form proteoglycan aggregates during assembly of new cartilage matrix; v) that myoblast fusion in the limb periphery also involves and requires removal of HA from the cell surface. Thus, the following experimental approaches will be taken. (a) The model of HA-cell interaction at critical stages of chick embryo limb development will be examined using cultured cells from these stages; specific morphological probes will also be used to examine the distribution of HA and cell surface receptor for HA in the developing limb. (b) The HA receptor present on the chondrocyte surface will be purified and characterized, and its potential role in mediating chondrocyte matrix assembly will be tested. (c) The mechanism of attachment of endogenous cell surface HA and the degree of receptor-mediated internalization and degradation of HA by cultured myoblasts and myotubes, as well as the role of these processes in fusion, will be investigated. (d) Monoclonal antibodies to the HA-receptor protein will be prepared and used for purification of the receptor protein and for morphological and experimental purposes. (e) The initiation of HA synthesis and interaction of nascent HA with membranes will be studied. Since aberrations in morphogenesis are basic to many congenital malformations, e.g. craniofacial anomalies and limb deformities, to inappropriate repair and regeneration processes, e.g. cirrhosis and atherosclerosis, and to the malignant behavior of tumor cells, an understanding of the influence of hyaluronate on cell behavior may provide important clues to the pathogenesis of these diseases.
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