FACTORS REGULATING THE 
PRODUCTION OF COAGULATION ACTIVITIES 
IN PERFUSED OF^GANS 
W. J. Dodds* and L. W. Hoyef 
Isolated rabbit livers and spleens were perfused first 
for 1 hr to release stored coagTilation activity and then 
for another 4 hr as previously described.^ The perfus- 
ate (80 ml) was devoid of coagulation activity and con- 
sisted of washed rabbit blood cells, human albumin in 
Tyrode's solution and trisodium citrate. At 30 min of 
the second perfusion period, 5 ml of plasma from nor- 
mal individuals or from those with congenital or ac- 
quired defects of coagulation were added to perfusates. 
Subsequent production of coagulation factors was in- 
versely proportional to perfusate coagulation activity, 
an observation which suggests regulation by a negative 
feedback process. The effect of splenic perfusion with 
factor VIII deficient plasma from patients with 
CRM ( — ) and (-f) hemophilia A and von Wille- 
brand's disease was compared; plasma was designated 
as CRM (-f-) if it contained material which cross-re- 
acted with human anti-factor VIII. As would be ex- 
pected for a system regulated by negative feedback, 
factor VIII production by the spleen continued while 
factor IX production ceased. In addition, a marked 
enhancement of splenic factor VIII activity was ob- 
served in the presence of CRM(-|-) hemophilia A 
plasma but not with CRM( — ) or von Willebrand's 
plasmas (p =r < 0.01). Experiments with factor IX 
deficient plasma from patients with CRM( — ) and 
( + ) hemophilia B demonstrated a similar but less 
striking enhancement of factor IX activity in the pres- 
ence of CRM( + ) hemophilia B plasma as compared 
with CRM(-) plasma (p = < 0.04). CRM(-t-) 
plasma contained material which cross-reacted with 
human anti-factor IX. These observations suggested 
that antigenically detectable but nonfunctional mole- 
cules accumulate in CRM ( + ) plasmas and can be con- 
verted rapidly by the spleen to active coagulation factors. 
Alternatively, inactive precursors of factor VIII and 
IX in CRM(-f ) plasma could stimulate splenic factor 
VIII and IX production, respectively. 
INTRODUCTION 
The liver is known to play a major role in the 
synthesis of plasma proteins, including the co- 
* Division of Laboratories and Research, New York State De- 
partment of Health, Albany, New York 12201. 
** Department of Medicine, University of Connecticut School of 
Medicine, Hartford, Connecticut 06105. 
*** Supported in part by U.S. Public Health Service Grant 
HL-09902 and by a Veterans Administration research grant. 
agulation factors, fibrinogen, prothrombin, and 
factors V, VII, IX, and X.^ Recent studies, how- 
ever, have implicated the liver, spleen and kid- 
ney as sites of synthesis and/or storage of 
factor VIII.^-^ Other sites for factor VII and IX 
synthesis have also been proposed.^-^ In pre- 
vious organ perfusion experiments, the appear- 
ance of coagulation factor VII, VIII, and IX 
activities gradually increased with time and was 
prevented by inhibitors of protein synthesis.^ 
The experimental approach was different from 
that of other investigators because an initial 
hour of perfusion was included to permit re- 
lease into the perfusate of tissue-stored coagu- 
lation factors.^ This was followed by a second 
perfusion period of 4 hr with a fresh aliquot of 
perfusate. The current studies were designed to 
investigate the possibility that a negative feed- 
back process regulated the appearance of coagu- 
lation activities during both the initial hour of 
perfusion (release period) and the final 4 hr of 
perfusion (synthesis period). 
MATERIALS AND METHODS 
Rabbit livers and spleens were isolated, pre- 
pared, and perfused as described previously.^'^ 
The perfusate was devoid of coagulation activ- 
ity and consisted of 80 ml of the following mix- 
ture : 25 ml of washed rabbit blood cells, 47 ml 
of 6 % human albumin in Tyrode's solution, and 
8 ml of 3.8% trisodium citrate. Final citrate 
concentration was maintained at 0.38% by ad- 
dition of 1-2 ml of 3.8% citrate hourly; without 
this additive platelet-fibrinous masses formed 
and became trapped in the perfusion filter. Ini- 
tial experiments with heparin were discontin- 
ued because heparinized perfusates produced an 
increased peripheral vascular resistance and ar- 
terial perfusion pressure, and a tendency to- 
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