The Regulation of Blood Coagulation 
Thrombomodulin and Thrombin 
Thrombomodulin is a protein of the endothe- 
lial cell surface that binds thrombin, a blood clot- 
ting en2yme. Because of its effects on thrombin 
activity, thrombomodulin is an essential natural 
anticoagulant. Several inflammatory mediators 
decrease the expression of thrombomodulin by 
endothelial cells. Understanding this process 
may help us understand the abnormal blood coag- 
ulation that accompanies much human disease. 
We have cloned and expressed variants of recom- 
binant human thrombomodulin in a variety of 
cultured cell lines. These cells have been used to 
define the structural requirements for thrombo- 
modulin cofactor activity. 
Thrombin is a protease enzyme that is required 
to form blood clots from fibrinogen. Thrombin 
also can inhibit clotting by first binding to throm- 
bomodulin and then digesting certain other 
blood coagulation factors. Disruption of throm- 
bin's normal balance between promoting and in- 
hibiting clotting can, in principle, cause either 
bleeding or thrombosis. We have constructed 
mutant forms of thrombin w^ith predominantly an- 
ticoagulant or procoagulant activities. Such mu- 
tant thrombins help to define the structural basis 
for the different activities of thrombin. In addi- 
tion, they provide reagents to test the physiologi- 
cal importance of specific thrombin activities, 
and mutant thrombins may also be found to 
have therapeutic procoagulant or anticoagulant 
properties. 
Tissue Factor 
Tissue factor, a cell surface protein that is 
found on many cells that do not normally contact 
the blood, is the most important physiological 
initiator of blood coagulation. When blood ves- 
sels are damaged, tissue factor is exposed to and 
binds to blood coagulation factor VII. The factor 
Vll-tissue factor complex then initiates a cascade 
of reactions that cause blood to clot. Both mono- 
cytes and endothelial cells express tissue factor 
activity in response to many stimuli, and this con- 
tributes to the abnormal thrombosis that accom- 
panies systemic infections. 
We have isolated cDNA clones for human tissue 
factor and localized the gene to chromosome 1 . 
These clones have been used to study the regula- 
tion of tissue factor in endothelial cells. Tumor 
necrosis factor, a protein made during inflamma- 
tion, causes a dramatic but transient increase in 
tissue factor activity. This appears to be the result 
of activating the gene, which is normally silent in 
endothelium. A second level of control can be 
employed to amplify this response. Tissue factor 
mRNA normally is degraded very rapidly, but it is 
stabilized by some agents that stimulate endothe- 
lial cells, and this may contribute to the induc- 
tion of tissue factor activity during inflammation. 
We are currently studying the structural ba- 
sis for this regulation of tissue factor mRNA 
degradation. 
The systems we are investigating provide abun- 
dant opportunities to answer biological ques- 
tions concerning the regulation of blood coagula- 
tion and to approach fundamental questions 
related to signal transduction, gene expression, 
and protein structure-function relationships. 
These studies may illustrate how several proteins 
can be coordinately regulated to promote blood 
clotting reactions on the vascular endothelium 
during inflammation. We will continue to ex- 
plore the mechanisms by which vWF, thrombo- 
modulin, thrombin, and tissue factor are regu- 
lated and will extend this work to other 
endothelial cell proteins that can promote or in- 
hibit thrombosis. 
346 
