Kinetics and stoichiometry of inactivation of some pancreatic and leucocyte serine proteinases by rabbit alpha 1-proteinase inhibitors F and S.

Purified rabbit alpha 1-proteinase inhibitors F and S were incubated with bovine trypsin, chymotrypsin or horse leucocyte neutral proteinases in order to determine the stoichiometry of inhibition, inactivation rates of the enzymes and dissociation constants of the complexes. Trypsin reacted with the two forms of alpha 1-PI with different velocities but in the molar ratio of 1:1 and yielding stable complexes. Chymotrypsin reacted very fast with the two forms of alpha 1-PI but required a three-fold molar excess of alpha 1-PI-S for almost complete inhibition, and again the dissociation constants were low.

Synthesis and secretion of some plasma proteins by tissue slices of Morris hepatoma 7777 and liver from control and turpentine-injected rats.

Slices of Morris hepatoma 7777 or rat liver isolated from control or turpentine-injected rats were incubated for 2 h with 14C-leucine. Radioactivities incorporated into albumin, alpha-fetoprotein, fibrinogen, alpha 1-AP-globulin, haptoglobin and alpha 1-acid glycoprotein were determined after the proteins had been isolated from the incubation medium or tissue homogenate by immunoprecipitation with monospecific antisera. It was found that hepatoma synthesizes fibrinogen, alpha 1-AP-globulin and alpha 1-acid glycoprotein in the amounts comparable to rat liver, whereas formation of albumin and haptoglobin is reduced 5- to 10-fold.

Changes in the blood level and affinity to concanavalin A of rat plasma glycoproteins during acute inflammation and hepatoma growth.

Plasma concentrations of ten individual proteins were measured by electroimmunoassay in young male Buffalo rats following injection of turpentine oil or implantation of Morris hepatoma 7777. The highest relative responses to inflammation and tumour growth were found for alpha 2-macroglobulin, alpha 1-acute-phase globulin and alpha 1-acid glycoprotein. As shown by crossed immuno-affinoelectrophoresis the concanavalin A-reactive fractions of the latter two glycoproteins were predominantly increased in plasma from injured and tumour-bearing rats.

The interaction between chymotrypsin and horse leucocyte neutral proteinases inhibitor.

The inhibition of alpha-chymotrypsin by horse leucocyte neutral proteinases inhibitor was time-dependent with synthetic substrate N-benzoyl-L-tyrosine ethyl ester but not with azo-casein. This time dependence could be used to calculate the rate constant kass for the association of the inhibitor with bovine alpha-chymotrypsin (kass = 0.3 X 10(6)M-1 S-1).

Regulation of vitamin K-dependent carboxylation.

Vitamin K-dependent carboxylation activity measured with pentapeptide substrate (Phe-Leu-Glu-Glu-Leu) gradually decreases upon in vivo injection of vitamin K to vitamin K-deficient rats. A decrease in pentapeptide carboxylation can also be observed by the in vitro addition of antibodies against prothrombin and other vitamin K-dependent proteins to the soluble system derived from vitamin K-deficient rat liver microsomes. In both cases, adding back in vitro partially decarboxylated vitamin K-dependent proteins or purified hepatic prothrombin precursor restores the level of pentapeptide carboxylation.

Th activities of perfused livers of control and streptozotocin diabetic rats in the synthesis of some plasma proteins and peptides.

By measuring the incorporation of 14C-DL-leucine a decreased capacity of isolated perfused steptozotocin diabetic rat liver to synthetize plasma total proteins, albumin and the seromucoid fraction was found as compared with a control. The relative rate of synthesis of the seromucoid proteins calculated as proportional to the rate of synthesis of albumin or the total plasma proteins was approximately twice as great as in control liver. 14C-DL-leucine was also incorporated in perfused liver into the nonprotein but non-dialysable sugar-peptide fraction but the synthesis of the glycopeptide components of this fraction was markedly reduced in diabetic rat liver.

The carcinoembryonic antigen test in urologic cancer.

The diagnostic value of the CEA test was evaluated in 2,029 blood and urine samples from 308 patients with urologic cancer, 13 with nonurologic cancer, 20 urologic patients with nonmalignant disease, and in 30 controls. The blood CEA test was positive in 50% of the patients with active urologic cancer and in 35% of those with inactive cancer. The urine CEA test was elevated only in patients with active bladder cancer, but most of these had concurrently infected urines.

The effect of D-galactosamine on plasma protein synthesis by the perfused rat liver from turpentine-stimulated donors.

D-galactosamine (100 mg) was added to the reconstituted blood during 4h perfusion of livers isolated either from control rats or those injected with turpentine 20 h or 5 h earlier. This dose of galactosamine administered 30 min before [3H]lysine significantly inhibited the incorporation of the label into liver proteins, and even more into plasma proteins, but albumin and acute-phase reactants (fibrinogen, seromucoid fraction, Concanavalin A-adsorbed glycoproteins) were all similarly affected. When galactosamine was administered in vivo simultaneously with turpentine, and the liver was isolated 5 h later, trauma-induced fibrinogen synthesis was selectively inhibited.

A polyvalent proteinase inhibitor from horse-blood-leucocyte cytosol. Isolation, purification and some molecular parameters.

Cytosol of horse blood polymorphonuclear leucocytes contains an inhibitor active against neutral proteinases from the granules of these cells and against chymotrypsin, elastases I and II from pig pancreas, but not against trypsin. A method has been elaborated to isolate and purify this inhibitor by means of salting out with ammonium sulphate (45---70% saturation), followed by chromatography and rechromatography on a column of DEAE-Sephadex A-50. The preparation obtained is homogeneous during polyacrylamide gel electrophoresis in the presence of 0.

Inhibition of the liver and plasma protein acute-phase response in mice by D-galactosamine.

Local inflammation evoked in Swiss albino mice by subcutaneous injection of Celite resulted in a rise of liver tyrosine aminotransferase activity and plasma level of fibrinogen and seromucoid, while liver alanine aminotransferase activity and plasma level of fibrinogen and seromucoid, while liver alanine aminotransferase activity and the plasma level of albumin and total protein remained unaltered. By measuring the incorporation of [14C] leucine, stimulation of liver and plasms protein synthesis by Celite injection was demonstrated. Administration of D-galactosamine (2-5 mg/10 g body weight) inhibited the enhanced synthesis of liver proteins, and especially of trauma-induced synthesis of plasma fibrinogen and seromucoid.

Substrate specificity and modifications of the active centre of elastase-like neutral proteinases from horse blood leucocytes.

Two proteinases (2A and 2B) purified from the granular fraction of horse blood leucocytes degrade casein (Km values 12.8 and 6mg/ml respectively) with maximum activity at pH 7.4 and in the presence of 2m-urea.

Isolation and some molecular parameters of elastase-like normal proteinases from horse blood leucocytes.

Cytoplasmic granules were isolated from horse blood polymorphonuclear leucocytes by the heparin method and extracted with 0.9% NaCl by repeated freezing. Soluble proteins were separated on a column of Sephadex G-75 followed by chromatography on a column of CM-Sephadex with a NaCl gradient.

Comparison of protein components of the plasma membrane fraction from rat liver and Morris hepatomas 5123D and 7777.

Isolated plasma membrane fractions from rat liver and Morris hepatoma 5123D and 7777 were labelled with radioiodine 125I by a chemical or enzymatic procedure and then were solubilized in 2 per cent solution of sodium dodecyl sulphate containing 1 per cent 2-mercaptoethanol. Solubilized proteins were separated into 20--22 zones staining with Coomassie Brilliant Blue R-250 after disc gel electrophoresis (7.5 per cent polyacrylamide gel).

Carcinoembryonic antigen test in renal cell carcinoma.

CEA (carcinoembryonic antigen) determinations were performed on 203 blood and urine samples from 23 patients with renal cell carcinoma. Neither the blood nor the urine CEA test was able to confirm the diagnosis or predict the status of the disease in more than one half of these patients. As presently constituted, the CEA test is not accurate for the diagnosis or prognosis of renal cell carcinoma.

The CEA test in urologic cancer: an evaluation and a review.

The diagnostic value of the carcinoembryonic antigen test was evaluated in 2,029 blood and urine samples from 308 urologic cancer patients, 13 patients with nonurologic cancer, 20 urologic patients with nonmalignant disease, and in 30 controls. The blood CEA test was positive in 50% of patients with active urologic malignancy and in 35% of patients with inactive malignancy. The urine CEA test was elevated only in patients with active bladder cancer but most of these patients had concurrently infected urines.