The liver has been proposed as a site of synthesis of blood-clotting factors, in part because of the demonstrated dominant role of the liver in synthesis of all plasma protein fractions except the y-globulins (1, 2). In addition, observations after hepatectomy (3, 4) and hepatic injury (5, 6) and in hepatic disease (7,8) have provided a body of evidence consistent with hepatic synthesis of fibrinogen, prothrombin, and Factors V, VII, IX, and X. This evidence, however, has not been considered definitive, mainly because it is based on indirect observations in vivo in humans and in animals suffering from variable, often profound, metabolic abnormalities induced by hepatic dysfunction or extirpation.This report describes the use of the isolated perfused rat liver in a more direct experimental evaluation of the role of the liver in the production of blood-clotting factors. The use of the isolated perfused liver was encouraged by Lupton's observations of shortened prothrombin time during perfusions of 3 to 4 hours (9), and by Pool and Robinson's studies of changes in clotting factor activity during in vitro incubation of rat liver slices (10). Presented here is not only more detailed direct evidence of hepatic synthesis of prothrombin and Factor VII, but also the first direct evidence for hepatic synthesis of Factors V and X.

The enzymatic properties of purified preparations of chicken liver and chicken skeletal muscle fructose bisphosphatases (D-fructose-1,6-bisphosphate 1-phosphohydrolase, EC 3.1.3.11) were compared. Both enzymes have an absolute requirement for Mg2+ or Mn2+. The apparent Km for MgCl2 at pH 7.5 was 0.5 mM for the muscle enzyme and 5 mM for the liver enzyme. Fructose bisphosphate inhibited both enzymes. At pH 7.5, the inhibitor constants (Ki) were 0.18 and 1.3 mM for muscle and liver fructose bisphosphatases, respectively. The muscle enzyme was considerably more sensitive to AMP inhibition than the liver enzyme. At pH 7.5 and in the presence of 1 mM MgCl2, 50% inhibition of muscle and liver fructose bisphosphatases occurred at AMP concentrations of 7 X 10(-9) and 1 X 10(-6) M, respectively. EDTA activated both enzymes. The degree of activation was time and concentration dependent. The degree of EDTA activation of both enzymes decreased with increasing MgCl2 concentration. Ca2+ was a potent inhibitor of both liver (Ki, 1 X 10(-4) M) and muscle (Ki, 1 X 10(-5) M) fructose bisphosphatase. This inhibition was reversed by the presence of EDTA. Ca2+ appears to be a competitive inhibitor with regard to Mg2+. There is, however, a positive homeotropic interaction among Mg2+ sites of both enzymes in the presence of Ca2+.

No abstract

No abstract

The effects at various pH levels of fructose 1,6-diphosphate (FDP), AMP, CaCl2, MnCl2, and/or MgCl2 on the thermal stability of pure forms of avian liver and muscle fructose 1,6-diphosphatases were investigated. Both enzymes were more stable when stored at 20 °C than at 2 °C. Maximum stabilities of the two enzymes when incubated at various temperatures in a Tris buffer occurred at approximately pH 6. All of the above compounds when added to a pH 7.5 Tris buffer markedly increased the thermal stabilities of both enzymes. Individually, FDP provided the highest degree of protection for the muscle enzyme whereas MnCl2 was most effective with the liver enzyme; in all cases the degree of protection was concentration dependent. At pH 7.5 combinations of MgCl2, FDP, and AMP were most effective. In the presence of these compounds initial inactivation of the muscle and liver enzymes did not occur until respective temperatures of 74 and 78 °C (15 min) were reached; these compare with initial inactivation temperatures in the absence of these compounds of 55 and 59 °C. The interaction of MgCl2 and FDP with the two enzymes at varying pH levels showed markedly different patterns. At pH7.5 and 8.8 increasing levels of MgCl2 provided increasing levels of protection whereas at pH 6.2 increasing MgCl2 concentrations resulted in an enhanced degree of inactivation. In the presence of FDP an opposite pattern was observed at the various pH levels. The presence of phosphate in the buffer tended to yield results similar to those of FDP.