The Journal of Cell Biology, Vol 87, 601-610, Copyright © 1980 by The Rockefeller University Press
Binding of thrombin to cultured human fibroblasts: evidence for receptor modulation
WM Hall and P Ganguly
Previous work from this laboratory has indicated that thrombin's influence
on cell growth can be negative as well as positive. Addition of enzyme to
actively growing or confluent cultures of human skin fibroblasts produced
growth stimulation, whereas cultures receiving thrombin at the time of
subculture displayed inhibited DNA synthesis and mitosis. The specific
binding of [125I]thrombin to cells under stimulatory and inhibitory
conditions has been studied. Fibroblasts receiving enzyme at subculture
bound about two times more [125I]thrombin than those processed in the same
way several hours later. The apparent dissociation constant for both groups
was approximately 1.5 x 10(-8) M. In each case binding was saturable,
although cells receiving enzyme at subculture showed a much higher rate of
binding. Experiments were conducted in which enzyme was added to cells at
various times after subculture. It was found that the ability of these
fibroblasts to specifically bind [125I]thrombin decreased progressively
over a 2-h period after subculture and then remained constant for at least
24 h. Evidence is also presented indicating that the binding of
[125I]thrombin in both experimental groups was inversely dependent upon the
culture density. The biological effects of elevated thrombin binding in
cells receiving enzyme at subculture were examined. It was found that
inhibited DNA synthesis and altered cellular morphology were directly to
this parameter. This study suggests that fibroblasts may possess cryptic
thrombin receptors that become exposed during subculture or after injury in
vivo. These possibilities and the relationship of cell shape to the
availability of thrombin receptors are discussed.
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