We report on the covalent immobilization of bone morphogenetic protein 6 (BMP-6) and its co-presentation with integrin ligands on a nanopatterned platform to study cell adhesion and signaling responses which regulate the transdifferentiation of myoblasts into osteogenic cells. To immobilize BMP-6, the heterobifunctional linker MU-NHS is coupled to amine residues of the growth factor; this prevents its internalization while ensuring that its biological activity is maintained. Additionally, to allow cells to adhere to such platform and study signaling events arising from the contact to the surface, we used click-chemistry to immobilize cyclic-RGD carrying an azido group reacting with PEG-alkyne spacers via copper-catalyzed 1,3-dipolar cycloaddition. We show that the copresentation of BMP-6 and RGD favors focal adhesion formation and promotes Smad 1/5/8 phosphorylation. When presented in low amounts, BMP-6 added to culture media of cells adhering to the RGD ligands is less effective than BMP-6 immobilized on the surfaces in inducing Smad complex activation and in inhibiting myotube formation. Our results suggest that a local control of ligand density and cell signaling is crucial for modulating cell response.
A radical mechanism is proposed for the formation of the 1,2-polymer 2 in the "spontaneous" polymerization of 4-vinylpyridinium salts 4-VP. HX (X = N O 3, C104, HS04, I) in aqueous solution on the basis of strong inhibition by oxygen and other radical inhibitors.Quantitative investigation of the influence of oxygen on the formation of the 1,2-polymer allowed the derivation of a formal kinetic scheme for the initiation and polymerization steps which is consistent with a free radical mechanism. Dependence of the average molecular weight of 2 on monomer concentration is in agreement with this model. Dependence of the inhibition time on the oxygen concentration as well as an acceleration of the final polymerization rate by prolonged treatment of the monomer solution with oxygen suggests the transitory involvement of polymeric peroxides. ZUSAMMENFASSUNG :Fur die Bildung des 1,2-Polymerisats 2 bei der ,,spontanen" Polymerisation von 4-Vinylpyridiniumsalzen 4-VP.HX ( X = N 0 3 , C104, HS04, I) in waI3riger Losung wird auf Grund der Inhibitorwirkung von Sauerstoff und anderen Radikalfangern ein radikalischer Mechanismus vorgeschlagen.Die quantitative Untersuchung des 0,-Einflusses auf die 1,2-Polymerisation fiihrte zu einer formalen Kinetik fur die Initiierungs-und Wachstumsreaktion auf der Basis einer radikalischen Polymerisation. Die Abhangigkeit des mittleren Molekulargewichts von 2 von der Monomerkonzentration ist in Ubereinstimmung mit diesem Modell. Die Abhangigkeit der Inhibitionszeit von der Sauerstoflkonzentration und eine Beschleunigung der Polymerisation nach langerer 0 ,-Behandlung der Monomer-Losung deuten auf eine Bildung polymerer Peroxide hin.
ZUSAMMENFASSUNG :Kritische Mizellkonzentrationen (cmc) aus Oberfllchenspannungsmessungen und durch Farbstoffsolubilisierung werden von 4-Vinylpyridinium-bzw. 4-Athylpyridinium-Salzlosungen bestimmt. Erhohung der Gegenionen-Konzentration erniedrigt die cmc; gleichzeitig damit andert sich auch die Zusammensetzung der Polymerisationsprodukte der spontanen Polymerisation in Wasser. Mogliche Erklarungen hierfur werden diskutiert. SUMMARY:Critical micell concentrations (cmc) of 4-vinylpyridinium or 4-ethylpyridinium salt solutions are determined from surface tension measurements and solubilisatio,n of a dyestuff. Increase of the concentration of the gegenions decreases the cmc; at the same time, the composition of the polymerization product of the spontaneous polymerization in water also changes. Possible explanations for this are discussed. EinlcitungDie Eigenschaften von 4-Vinylpyridiniumsalzen (1) -insbesondere deren Reaktionsverhalten bei der spontanen Polymerisation in waI3riger Losung wurden in letzter Zeit intensiv untersucht -'). ,,Spontane Polymerisation" bedeutet hierbei, daI3 die waI3rigen Losungen der Monomersalze ohne weiteren Zusatz zu polymerisieren beginnen. Es zeigte sich, daI3 sowohl Reaktionsordnung als auch Polymerstruktur in charakteristischer Weise von der Monomerkonzentration abhiingen'), wie es in G1. (i) schematisch dargestellt ist. *I 5. Mitteilung: E. Perplies. H. Ringsdorf u. J. H. Wendorff, Ber. Bunsenges. Phys.Chem. 78,921 (1974).
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