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Interaction of xenopus ectodermal cells with fibronectin and isolated fibronectin domains in vitro
$I I N T E R A C T I O N OF XENOPUS E C T O D E R M A L CELLS WITH F I B R U N E C T I N AND ISOLATED F I 8 R O N E C T I N DOPIAINS IN VITRO R.Winklbauer. M a x - P l a n c k - l n s t i t u t fur E n t w i c k l u n g s b i o l o g i e ,74 T U b i n g e n , F R G
THE EFFECTS OF CADMIUM ON SECRETORY AMELOGENESIS IN M I N E R A L I Z I N G T O O T H G E R M S IN VITRO. J. W ~ l t g e n s , A. E r o n c k e r e , D. L y a r u u I and Th. B e r v o e t s . Lab of Tooth Development, Dept. of Oral Cell Biology, ACTA, VriJe University, de Boelelaan 1115, 1081 EV A m s t e r d a m , The Netherlands. Cadmium (Cd) accumulates in the mineralizing matrices of developing tooth germs when animals are e~R~sed to this heavy metal. Radio autography with "v~Cd indicated that this metal can also be detected in tooth forming cells; however, its synthesis and mineralization are yet unknown. It was the aim of the present study to examine histologically and biochemically the effects of various Cd concentrations on these processes in vitro. First maxillary molar hamster tooth germs in secretory stage of amelogeneels were cultured for 24h in the presence of 0.1, ~ or I0 pM C ~ 1 2 and @~ther radiolebelled with ~N-Prollne, ~#C~ and ~'P or used for histological examination. For biosythetic experiments after labelling, the enamel organ was gently separated from the underlying enamel and both chemically extracted. Histologically I0 pM Cd resulted into a complete degeneration of the secretory ameloblaets along with inhibition of mineral transport, protein synthesis and protein secretion. In contrast at I pM Cd no histological differences were evident with control explants; however secretion of a protein was inhibited which was insoluble in ~ichloroacetic acid and contained 3H-Proline and P, as this fraction increased in cells of Cd treated enamel organs. Extracellularly, a lower amount of such compound was present in the enamel along with a smaller amount of ~5Ca compaired w i t h controls. At pM ~ n o clear effect was found on the molar ~5Ca/ozP ratio in the mineral of the forming enamel, We conclude that i pM Cd inhibits the secretion of a phosphorylated, proline containing enamel matrix protein along with a fall in mineral transport towards the forming enamel without changing the composition of the minerals formed in the enamel.
During a m p h i b i a n gastrulation, mesodermal cells migrate on f i b r o n e c t i n ( F N ) - c o n t a i n i n g fibrils which are attached to e c t o d e r m cells. We hava asked how e c t o d e r m a l cells interact with the isolated FN molecule. In a cell adhesion assay, the binding of these cells to F N - c o a t e d plastic surfaces was quantitated. A d h e s i o n and cell spreading was found only at unusually high concentrations of FN, but was specifically inhibited by an anti-serum against FN. For adhesion to occur, the intact FN molecule could De replaced by an isolated 120kD fragment containing the normal cell binding site, but also by a 30kD heparin-binding fragment. To characterize further the adhesion of ectoderm cells to these fragments, a synthetic peptide is used which is hcmui(JL}ous to the normal cell bindino site and prevents attachment of cells to this site by competitive inhibition.
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THE EFFECTS OF CADMIUM ON SECRETORY AMELOGENESIS IN M I N E R A L I Z I N G T O O T H G E R M S IN VITRO. J. W ~ l t g e n s , A. E r o n c k e r e , D. L y a r u u I and Th. B e r v o e t s . Lab of Tooth Development, Dept. of Oral Cell Biology, ACTA, VriJe University, de Boelelaan 1115, 1081 EV A m s t e r d a m , The Netherlands. Cadmium (Cd) accumulates in the mineralizing matrices of developing tooth germs when animals are e~R~sed to this heavy metal. Radio autography with "v~Cd indicated that this metal can also be detected in tooth forming cells; however, its synthesis and mineralization are yet unknown. It was the aim of the present study to examine histologically and biochemically the effects of various Cd concentrations on these processes in vitro. First maxillary molar hamster tooth germs in secretory stage of amelogeneels were cultured for 24h in the presence of 0.1, ~ or I0 pM C ~ 1 2 and @~ther radiolebelled with ~N-Prollne, ~#C~ and ~'P or used for histological examination. For biosythetic experiments after labelling, the enamel organ was gently separated from the underlying enamel and both chemically extracted. Histologically I0 pM Cd resulted into a complete degeneration of the secretory ameloblaets along with inhibition of mineral transport, protein synthesis and protein secretion. In contrast at I pM Cd no histological differences were evident with control explants; however secretion of a protein was inhibited which was insoluble in ~ichloroacetic acid and contained 3H-Proline and P, as this fraction increased in cells of Cd treated enamel organs. Extracellularly, a lower amount of such compound was present in the enamel along with a smaller amount of ~5Ca compaired w i t h controls. At pM ~ n o clear effect was found on the molar ~5Ca/ozP ratio in the mineral of the forming enamel, We conclude that i pM Cd inhibits the secretion of a phosphorylated, proline containing enamel matrix protein along with a fall in mineral transport towards the forming enamel without changing the composition of the minerals formed in the enamel.
During a m p h i b i a n gastrulation, mesodermal cells migrate on f i b r o n e c t i n ( F N ) - c o n t a i n i n g fibrils which are attached to e c t o d e r m cells. We hava asked how e c t o d e r m a l cells interact with the isolated FN molecule. In a cell adhesion assay, the binding of these cells to F N - c o a t e d plastic surfaces was quantitated. A d h e s i o n and cell spreading was found only at unusually high concentrations of FN, but was specifically inhibited by an anti-serum against FN. For adhesion to occur, the intact FN molecule could De replaced by an isolated 120kD fragment containing the normal cell binding site, but also by a 30kD heparin-binding fragment. To characterize further the adhesion of ectoderm cells to these fragments, a synthetic peptide is used which is hcmui(JL}ous to the normal cell bindino site and prevents attachment of cells to this site by competitive inhibition.
27S