Summary
Endothelial cells and their extracellular matrix formed in vitro are often used as
a model for subendothelium in studies on platelet-vessel wall interaction. We have
characterized the influence of culture conditions of endothelial cells on the formation
of extracellular matrix and on the interaction of the matrix with platelets. Passage
number, time of confluence, serum concentration and the addition of heparin, growth
factors and antibiotics to the culture medium were varied and the extracellular matrices
were isolated. The amount of fibronectin and von Willebrand factor present in the
matrix were measured and the number of platelets adhering to these matrices after
perfusion with citrated whole blood at a shear rate of 1000 s-l was determined. A three times increase of the amount of von Willebrand factor in
the matrix was found when the serum concentration was increased from 2.5% to 30%.
When the passage number of the cells was increased or the period during which the
cells were at confluence was extended, the amount of von Willebrand factor in the
matrix was decreased up to 50%. Addition of heparin or ECGS (endothelial cell growth
supplement) decreased the von Willebrand factor content in the matrix. Addition of
penicillin or streptomycin to the culture medium had no influence on the amount of
von Willebrand factor deposited in the matrix or secreted into the medium, however,
other antibiotics such as gentamycin and neomycin decrease the amount of von Willebrand
factor in the matrix. No influence on the amount of fibronectin in the matrix was
found under all conditions tested.
There was a strong correlation between the amount of von Willebrand factor in the
matrix and the number of platelets adhering to the matrix. A decrease or increase
of the amount of von Willebrand factor was always correlated with a decrease or increase
of the number of platelets adhering to the matrix. These results indicate that the
synthesis and deposition of von Willebrand factor in the extracellular matrix by cultured
endothelial cells is very sensitive to variations in culture conditions and that the
amount of von Willebrand factor in the matrix predominantly determines the reactivity
of the matrix for platelets.
