Increased solubility of newly synthesized collagen in retinal capillary pericyte cultures by nonenzymatic glycosylation.

Abstract

The effect of increasing glucose concentration on the solubility of newly synthesized collagen was studied in bovine retinal capillary pericyte (BRCP) cultures. Synchronized pericytes were labeled with [3H]-proline in media containing either 5, 10, 20 or 40 mM glucose, or 5 mM glucose and 100 micrograms/ml beta-aminopropionitrile fumarate (BAPN). The relative rate of collagen synthesis in the medium (soluble collagen) was significantly increased, and the rate in the cell layer (insoluble collagen) was decreased, by an elevated glucose concentration. The increase in collagen solubility with 20 and 40 mM glucose (over normal, 5 mM glucose) was 14 and 42%, respectively, of that obtained with BAPN. BAPN and 4-deoxypyridoxine inhibited BRCP lysyl oxidase activity, but elevated glucose concentrations did not. Increased nonenzymatic glycosylation of newly synthesized collagen was demonstrated in BRCP cultures co-labeled with L-[14C]-glucose and L-[3H]-proline. The increase in collagen solubility with elevated glucose concentrations therefore appears to be the result of the inhibition of lysyl-derived cross-link formation because of an increased nonenzymatic glycosylation of the epsilon-amino groups of lysyl residues of newly synthesized collagen in culture.