Binding of thrombin to cultured human fibroblasts: evidence for receptor modulation.

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Abstract
Previous work has indicated that thrombin 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 was studied. Fibroblasts receiving enzyme at subculture bound about 2 times more [125I]thrombin than those processed in the same way several hours later. The apparent dissociation constant for both groups was .apprx. 1.5 .times. 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. The ability of these fibroblasts to specifically bind [125I]thrombin decreased progressively over a 2-h period after subculture and remained constant for at least 24 h. The binding of [125I]thrombin in both experimental groups seemed inversely dependent upon the culture density. The biological effects of elevated thrombin binding in cells receiving enzyme at subculture were examined. Inhibited DNA synthesis and altered cellular morphology were directly related to this parameter. Fibroblasts may possess cryptic thrombin receptors that become exposed during subculture of after injury in vivo. These possibilities and the relationship of cell shape to the availability of thrombin receptors are discussed.