Does plasma urokinase antigen reflect tumor urokinase in prostatic carcinoma?
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Bone Neoplasms↗
Biomedical subjects
B R Binder
Publications and source records attributed to B R Binder.
Plasminogen activation in diabetes mellitus: normalization of blood sugar levels improves impaired enzyme kinetics in vitro.
We have demonstrated previously that fibrin enhanced plasmin formation by the vascular plasminogen activator was significantly impaired, when components isolated from the plasma of three uncontrolled diabetic patients (type I) were used to study plasminogen activation in vitro. In the present study it can be demonstrated that functional properties of the vascular plasminogen activators as well as of the plasminogens from the same three diabetic patients are significantly improved after normalization of blood sugar levels and improvement of HbA1c values. Most pronounced the Km of diabetic vascular plasminogen activator in the presence of fibrin returned to normal values, and for diabetic plasminogen the prolonged lag period until maximal plasmin formation occurred was shortened to almost control values. From these data we conclude that the observed abnormalities of in vitro fibrinolysis are not primarily associated with the diabetic disease, but might be secondary to metabolic disorders caused by diabetes.
Adult↗
Kinetic analysis of plasminogen activation by purified plasma kallikrein.
Kinetics of plasminogen activation by purified activated plasma kallikrein have been studied in a purified system using Glu-plasminogen as a substrate. A synthetic paranitroanilide substrate was used for quantification of the formed plasmin. In that system kallikrein cleaved plasminogen with a Km value of 0.56 microM, a kcat of 1.6 X 10(-4) s-1 and a catalytic efficiency kcat/Km of 2.7 X 10(-4) s-1 microM-1. Addition of CNBr fibrinogen fragments resulted in an increase of Km to 1.18 microM, an increase of kcat to 5.1 X 10(-4) s-1 and an increase in the catalytic rate constant kcat/Km to 4.3 X 10(-4) s-1 microM-1. Addition of purified high molecular weight kininogen had no effect on the kinetics of plasminogen activation whether or not stimulating fibrinogen fragments were present. A stimulating effect of fibrinogen fragments could also be shown for the cleavage of the low molecular weight paranitroanilide substrate H-D-Pro-Phe-Arg-pNA by kallikrein; in that system the kcat for substrate cleavage by kallikrein increased from 200 s-1 to 280 s-1, while the Km value remained unchanged. From these data it can be concluded that based on enzyme kinetic studies plasminogen activator activity of purified plasma kallikrein is about 1/1000 of that of high molecular weight urokinase and is only slightly influenced by addition of stimulating fibrinogen fragments. Addition of high molecular weight kininogen does not affect plasminogen activator activity of purified plasma kallikrein.
Fibrin↗
Increased urokinase activity to antigen ratio in human renal-cell carcinoma.
Eighteen patients with renal-cell carcinoma have been investigated in an attempt to elucidate the ratio of active urokinase enzyme to urokinase antigen in the tumor and adjacent normal kidney. The tumor itself exhibited a significantly increased total fibrinolytic activity, an increase in the relative contribution of anti-urokinase IgG-inhibitable plasminogen activator activity and increased levels of urokinase antigen when compared to normal renal tissue. In tumor-adjacent tissue total fibrinolytic activity was also, but not significantly, increased, this increase being completely due to tissue-type plasminogen activator. Correlation of active urokinase-type plasminogen activator with urokinase antigen revealed that in tumor tissue the enzyme was present to more than 70% in its active form whereas in tumor-adjacent tissue and normal renal tissue only half of the enzyme appeared to be active. No correlation was obtained between urokinase antigen present in one of the 3 tissues investigated and plasma urokinase antigen.
Adult↗
[Use of monoclonal antibodies in the diagnosis of fibrinolysis].
Monoclonal antibodies to tissue type plasminogen activator (t-PA), urokinase (u-PA) and alpha-2-antiplasmin were obtained by immunizing Balb C mice with the respective purified antigens and fusion of spleen cells with mouse myeloma cells (NSO). The selected monoclonal antibodies reacted exclusively with the respective antigens used for immunization. 3 out of 7 monoclonal antibodies directed against t-PA inhibited plasminogen activation by t-PA and 6 of 7 interfered with the fibrin potentiating effect. Only one out of 5 monoclonal antibodies directed against u-PA inhibited plasminogen activation by u-PA. None of the monoclonal antibodies directed against t-PA or u-PA inhibited the cleavage of low molecular weight paranitroanilide substrates by the respective plasminogen activators. Likewise, the 3 monoclonal antibodies to alpha-2-antiplasmin did not functionally inhibit alpha-2-antiplasmin. The selected monoclonal antibodies were used to develop sensitive test systems for the respective antigens. The lower detection limit for determination of t-PA was found to be 0.5 ng/ml, for determination of u-PA 0.05 ng/ml, and for determination of alpha-2-antiplasmin 5 ng/ml.
Animals↗
Changes in plasminogen activator activity and plasmin inhibition in the pig during experimental hypovolaemia.
A markedly decreased plasminogen activator activity (PAA) was seen in lung, heart and kidney after experimental hypovolemia in the pig. In 2 of 12 cases, an increased PAA was found in liver and spleen. In skeletal muscle, skin, sciatic nerve and in 10 of 12 cases in liver and spleen PAA was unchanged. A slightly increased plasmin inhibition was noted only in the lung. These changes in tissue PAA were reflected by an increase in the PAA of euglobulin fractions concomitant with a decrease in antiplasmin activity in the blood samples obtained during the bleeding experiments.
Animals↗
Effect of hypophysectomy on the tissue and blood plasminogen activator activity in the rat.
The response of the fibrinolytic system to hypophysectomy in the rat was studied. 4 and 6 weeks after hypophysectomy the tissue plasminogen activator activity (PAA) was significantly increased in the intima of the aorta, slightly but significantly increased in the renal medulla, and significantly decreased in the vessels of the testis. No change in the PAA of the caudal vena cava, heart, lung and renal cortex was noted. Also, the tissue plasmin inhibition remained unchanged. No effect on the PAA in plasma euglobulin fractions was observed, whereas F VIII was slightly but not significantly decreased 6 weeks after hypophysectomy.
Animals↗
Development and clinical use of an oral heat-inactivated whole cell pertussis vaccine.
An orally administered, killed whole cell pertussis vaccine was developed which proved to be safe and potent in animal experiments. This vaccine was studied in limited field trials in Austria. In these field trials 1 X 10(12) bacterial cells were given orally on days 2, 3, 4, and 5 after birth with an oral booster of 1 X 10(12) cells given at six weeks. This vaccine was tolerated without side effects in more than 20,000 newborns, did induce a specific immune response including anti-pertussis IgG in serum, anti-pertussis IgA in saliva and specific pertussis mitogen stimulation. This immune response occurred significantly earlier in the orally but not in the parenterally vaccinated group. In addition, anti-pertussis antibodies in saliva were not induced in the parenterally vaccinated group. Pertussis morbidity in 11,192 orally vaccinated infants was compared to 3496 infants who did not receive oral vaccination and was found to be significantly lower for up to one year. There was no overall difference between orally and non-orally vaccinated infants as far as pertussis hospitalization and verified pertussis infection after one year was concerned.
Administration, Oral↗
Tissue plasminogen activator inhibitor in human plasma: development of a functional assay system and demonstration of a correlating Mr = 50,000 antiactivator.
An assay system for determination of the "fast"-acting inhibitor (antiactivator, AA) of tissue-type plasminogen activator (tPA) in human plasma was developed. The system is based on incubation of plasma samples with various amounts of vessel wall-derived tPA for 8 minutes at 25 degrees C, followed by acidification and determination of the residual tPA activity by an indirect spectrophotometric assay. One unit of AA was defined as the amount inhibiting 1 U of tPA. AA levels in normal controls (n = 26) were 1.4 to 17.4 U/ml (median 3.0 U/ml) and 0.9 to 17.5 U/ml (median 3.0 U/ml) in patients with a history of deep venous thrombosis (n = 26). When plasma was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by reverse fibrin autography, AA activity appeared as an inhibitory band corresponding to a relative molecular mass of 50,000. In six samples the inhibitory activity of this band was directly correlated to the functional AA activity of the plasma samples.
Electrophoresis, Polyacrylamide Gel↗
Plasminogen activator activity in bone metastases of prostatic carcinomas as compared to primary tumors.
The plasminogen activator content of extracts of 14 prostatic carcinomas and the respective bone metastases was determined and found to be at an average 1.5 times higher in the extracts from bone metastases than in the primary tumors. Furthermore, the relative contribution of the two known types of plasminogen activators, urokinase-type (u-PA) and tissue-type (t-PA), was evaluated using specific antibodies. About 70% of the plasminogen activator activity in the primary tumors was inhibited by anti-urokinase IgG, whereas the same antibody nearly completely inhibited the plasminogen activator activity in extracts from bone metastases. Using antibodies against t-PA about 30% of the plasminogen activator activity could be quenched in extracts of primary tumors but less than 10% in extracts of bone metastases. Further studies revealed that the increased amount of u-PA in extracts of bone metastases is not caused by different extractability but is also reflected by a relative increase in the amount of u-PA demonstrable by immune histochemical techniques using anti-urokinase IgG. Upon purification, the predominant plasminogen activator from extracts of bone metastases could also be identified physicochemically as urokinase.
Adenocarcinoma↗
Urokinase antigen in plasma: age and sex dependent variations.
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Adolescent↗
[Tissue plasminogen activator activity in early prostatic cancer and in bone metastases of prostatic cancer].
Most malignant cells exhibit increased plasminogen activator activity which, in turn, leads to the formation of the fibrinolytically active enzyme, plasmin. Since solid tumours in man are surrounded by a fibrin network, the fibrinolytic activity of the tumour may influence tumour growth and metastasis. In the present study plasminogen activator activity, as assessed in purified extracts, was compared in benign hyperplasia of the prostate (group A, n = 6), non-metastasizing+ prostatic carcinoma (group B, n = 26), and in prostatic carcinoma with bone metastasis (group C, n = 10). Plasminogen activator activity was significantly higher in prostatic carcinoma than in hyperplasia, but there was no significant difference in plasminogen activator activity between prostate carcinoma with or without bone metastasis. However, plasminogen activator activity in the bone metastasis cells was significantly higher than in the primary tumour. If a positive correlation between fibrinolytic activity of the tumour and the metastasizing capacity were postulated, particular importance could be attached to bone metastasis in prostatic cancer.
Bone Neoplasms↗
Comparison between vascular plasminogen activator activity and changes in blood flow in the renal cortex in pigs.
Patterns of vascular plasminogen activator activity (PAA) and of blood flow were compared in the renal cortex of pigs. The comparison was made in pigs with or without induction of hemorrhagic hypotension and continuous infusion of indomethacin (prostaglandin synthetase inhibitor) or saralasin (competitive inhibitor of angiotensin II). Blood flow was measured by radiolabelled microspheres. A significant decrease in the vascular PAA was observed only in saralasin treated animals (15% in not hemorrhaged pigs and 30% in the inner and 45% in the outer half of the renal cortex in hemorrhaged pigs). Changes in plasmin inhibitor activity in the renal cortex were not noted. No correlation could be seen between vascular PAA and changes in blood flow in the renal cortex or changes in the mean arterial blood pressure. The vascular fibrinolytic response to saralasin should be due to an effect of saralasin independent of the changes in blood flow induced by this agent.
Angiotensin II↗
Plasminogen activation in diabetes mellitus. Kinetic analysis of plasmin formation using components isolated from the plasma of diabetic donors.
Two components of the fibrinolytic system, plasminogen and the vascular plasminogen activator, have been isolated to apparent homogeneity from the post-venous occlusion plasma of three diabetic patients (hemoglobin A1C greater than 7%) and of one nondiabetic control person. Plasminogen activation was studied for each person separately in the absence and presence of CNBr fragments of fibrinogen. Activation of diabetic plasminogen by urokinase was not significantly altered as compared to the activation of control plasminogen. The same was found when diabetic plasminogen was activated by control vascular plasminogen activator in the presence of fibrinogen fragments but only at plasminogen concentrations below 10-30 nM; at higher substrate concentrations, however, plasminogen activation was impaired in a pattern resembling substrate inhibition. Activation of control plasminogen by diabetic vascular plasminogen activator was completely impaired in the absence of fibrinogen fragments. Addition of fibrinogen fragments stimulated plasmin formation by diabetic vascular plasminogen activator resulting in kinetic constants which were similar to the activation of control plasminogen by control vascular plasminogen activator in the absence of fibrinogen fragments (Km = 7.5 microM, kcat = 0.05 S-1). Addition of fibrinogen fragments in controls decreased Km values to less than 0.1 microM. Despite addition of fibrinogen fragments the rate of plasmin formation from diabetic plasminogen by diabetic vascular plasminogen activator isolated from the same diabetic donor was so small that kinetic constants could not be calculated.
Diabetes Mellitus↗
Isolation and characterization of plasminogen activators from hyperplastic and malignant prostate tissue.
Plasminogen activators from prostate tissue were purified to apparent homogeneity by a procedure involving reverse ammonium sulfate gradient solubilization, chromatography on gelatine-Sepharose, gel filtration on Sephadex G-150, and chromatography on Con A-Sepharose as a final step. Two activators were obtained. The predominant one exhibited physicochemical, immunochemical and functional properties indistinguishable from human urinary high molecular weight urokinase. The other one, which amounted to about 20% was immunochemically related to tissue type plasminogen activator and its plasminogen activator activity was enhanced by addition of CNBr-fibrinogen fragments in a similar pattern as for the vascular plasminogen activator. The molecular weight, however, and enzymatic activities on the synthetic low molecular weight paranitroanilide substrates pyro-Glu-Gly-Arg-pNA and H-D-Ile-Pro-Arg-pNA were different to vascular plasminogen activator but similar to high molecular weight urinary urokinase.
Humans↗
Tissue plasminogen activator activity in prostatic cancer.
19 patients with carcinoma of the prostate and 6 controls with hyperplasia of the prostate, have been investigated to elucidate the relationship between tumor fibrinolytic activity, the degree of malignancy, tumor stage and metastatic spread. Significantly (p less than 0.001) higher tissue plasminogen activator activities were found in metastatic carcinoma compared to nonmetastatic carcinoma or hyperplasia of the prostate. Antibodies against human urokinase caused inhibition of urokinase and plasminogen activator activity from human prostatic carcinoma or hyperplasia. Antibody inhibition studies as well as physicochemical characteristics of the plasminogen activator produced by prostatic hyperplasia and carcinoma tissue indicate that this plasminogen activator is identical or similar to urokinase.
Antibodies, Neoplasm↗
Characterization of a specific anti-human urokinase antibody: development of a sensitive competition radioimmunoassay for urokinase antigen.
Specific inhibiting IgG antibodies were raised in a rabbit using purified human high molecular weight urokinase as antigen. The antibodies reacted with both high molecular weight and low molecular weight human urokinase using an Ouchterlony double-immunodiffusion technique in such a way that one line of complete identity was obtained. Neither precipitation nor functional inhibition was observed for the tissue-type plasminogen activator. Kinetic studies with plasminogen as a natural high molecular weight substrate or the synthetic low molecular weight p-nitroanilide substrate pyro-Glu-Gly-Arg-pNA revealed, for both substrates, mainly a competitive type of inhibition for the Fab fragments of the specific anti-urokinase antibodies. This characterized anti-urokinase IgG was employed to develop a competitive radioimmunoassay, for human urokinase with 125I-labeled urokinase as tracer. In this radioimmunoassay, the lower detection limit for urokinase antigen was 10 pg/ml sample; the intrassay variation was 2.8%, and the interassay variation was 5.3%. Applying this radioimmunoassay to plasma samples obtained from healthy young volunteers, urokinase antigen could be measured in a concentration of 7.82 +/- 3.97 ng/ml for mean and 6.66 +/- 2.39 ng/ml for women (mean +/- SD).
Adult↗
Localization and immunological characterization of plasminogen activators in human prostate tissue.
In tissue sections of human hyperplastic as well as carcinomatous prostate glands fibrinolytic activity could be attributed to collecting ducts and vessel walls. Inhibition studies with antibodies directed against human urokinase (u-PA) or tissue plasminogen activator (t-PA) revealed the presence of a plasminogen activator completely inhibitable by anti-t-PA IgG in the vessel walls, while in collecting ducts the plasminogen activator activity was inhibited by anti-u-PA IgG to only 80% and by anti-t-PA IgG to about 20%. A mixture of anti-u-PA and anti-t-PA IgG was able to inhibit fibrinolytic activity in collecting ducts completely. In carcinomatous prostate glands total fibrinolytic activity was significantly higher, but localization and relative contribution of anti-u-PA-inhibitable and anti-t-PA-inhibitable plasminogen activators remained the same.
Humans↗