Nonenzymatic glycation of fibronectin impairs adhesive and proliferative properties of human vascular smooth muscle cells.
Nonenzymatic glycation of proteins is involved in the pathogenesis of diabetes vascular complications. Extracellular matrix proteins are a prominent target for nonenzymatic glycation because of their slow turnover rates. The aim of this study was to investigate the influence of human fibronectin (F) nonenzymatic glycation on adhesion and proliferation of cultured human vascular smooth muscle cells (hVSMC). Incubation of human F with 500 mmol/L D-glucose at 37 degrees C induced a time-dependent increase in fluorescence detectable at 440 nm after excitation at 363 nm. Nonenzymatic glycation did not affect binding of F itself to the plates. Adhesion of hVSMC to F increased with the increase of incubation time of the cells on the protein from 30 minutes up to 120 minutes and remained stable thereafter. Adhesion to glycated fibronectin (GF) was reduced in comparison to control F at all the different adhesion times. Adhesion of hVSMC to GF was reduced when F was exposed to glucose for 4, 9, or 28 days (P=.0417 to .0025), but not when F was exposed for 1 day. Adhesion of hVSMC to GF was reduced compared with adhesion to nonglycated F at all coating concentrations from 0.2 to 10 micrograms/mL (P=.05 to .014). Thus, nonenzymatic glycation of F impairs adhesion of hVSMC in vitro. Proliferation of hVSMC on F increased with increasing concentrations of the protein as coating agent (ANOVA:P<.0001 for both nonglycated F and GF). Proliferation with F glycated for 4, 9, and 28 days was reduced at concentrations of 1, 3, and 10 micrograms/mL as compared with proliferation with nonglycated F (P=.0253 to .0001). Proliferation on F glycated for only 1 day was not significantly reduced. When the number of hVSMC plated on control F was reduced by 25% to take into account the reduced adhesion, the number of cells that proliferated on F was still reduced. In conclusion, nonenzymatic glycation of F impairs adhesive and proliferative properties of hVSMC.
Cavalot
F
F
Department of Clinical and Biological Sciences, University of Turin, San Luigi Gonzaga Hospital-Orbassano, Italy.
Anfossi
G
G
Russo
I
I
Mularoni
E
E
Massucco
P
P
Mattiello
L
L
Burzacca
S
S
Hahn
A W
AW
Trovati
M
M
D002448:Cell Adhesion; D002455:Cell Division; D002478:Cells, Cultured; D005353:Fibronectins / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D009131:Muscle, Smooth, Vascular / Q000166:cytology*
D005353:Fibronectins