Effect of kininogen on the activity of the kallikreinogen-kallikrein system of human blood serum.

In experiments with kallikreinogen, kallikrein and kininogen preparations from human blood serum partially purified on QAE-Sephadex A-50, CM-Sephadex and Sephadex G-200 it was established that N-benzoyl-L-arginine ethyl ester (BAEE)-esterase activity of kallikrein and the process of autoactivation of kallikreinogen are inhibited by kininogen, especially by its high-molecular form.

Isolation of a cationic fragment of high molecular mass kininogen and effect of this fragment on human plasma kallikrein activity.

The effect of a cationic fragment of high molecular mass (HMM) kininogen on kallikrein activity and the autocatalytic process of prekallikrein-to-kallikrein conversion were studied. Prekallikrein and kallikrein were obtained by ion exchange and affinity chromatography; the purification of HMM kininogen was achieved using QAE- and DEAE-Sephadex, and separation of the cationic fragment was carried out by gel filtration. It was shown that the fragment of HMM kininogen (mol. mass. 7000, high lysine content) suppressed the activity of a labile form of kallikrein and prevented activation of prekallikrein. A model of the prekallikrein-kallikrein system is proposed. The model is based on the recently established fact of occurrence of two kallikreins and two kallikrein precursors. It involves formation from kininogen, besides kinins, of a kallikrein inhibitor.

Preparation and some properties of highly purified human serum kallikrein.

A 3000--6000-fold purified kallikrein was obtained from human serum in 10--25% yield by chromatography on QAE-Sephadex A-50, Molselect CM-50 and on soybean trypsin inhibitor (SBTI)-AH-Sepharose 4-B. The enzyme had a specific activity of 14--23 U, as measured by BAEE hydrolysis. Some properties of highly purified kallikrein are described.

Preparation and some properties of human serum kallikrein immobilized on ARM-Sepharose.

A highly purified human serum kallikrein immobilized on CH-Sepharose 4-B was obtained. KM values for N-benzoyl-L-arginine ethyl ester and N-tosyl-L-arginine methyl ester hydrolysis of this preparation were 1.10 x 10(-3) M and 3.6 x 10(-4) M, respectively; pH optimum of hydrolysis of these esters were found to be 8.2 and 8.5, respectively. The immobilized kallikrein possessed kininogenase activity and was capable of activating prekallikrein.

[Comparative study of the oncornavirus A and D proteins of human J-96 cells by the methods of isoelectric focusing and polyacrylamide gel electrophoresis]. / Sravnitel'noe izuchenie belkov onkornavirusov A i D v perevivaemykh kletok cheloveka J-96 metodami izoélektricheskogo fokusirovaniia i élektroforeza v poliakrilamidnom gele.

Three peaks of 14C-radioactivity with buoyant densities of 1.23--1.24, 1.26 and 1.29 g/ml were detected in a cytoplasmic extract of J-96 cells upon equilibrium centrifugation in sucrose gradient. Electron microscopy of the 1.23--1.24 g/ml buoyant density fraction revealed particles 60--80 nm in diameter showing morphology characteristic of oncornavirus A. Isoelectric focusing in polyacrylamide gel showed polypeptides of extracellular D virus and oncornavirus A to differ in isofocusing points (pI). Proteins of extracellular D virus were localized in zones with pH 3.7, 4.0, 4.4, 4.7, 5.6, 6.5, 8.1, 9.45, and 10.0; polypeptide of intracytoplasmic oncornavirus A had the following isofocusing points: 4.0, 4.9, 6.7, 7.3, 9.0, 9.45 and over 10.0. Electrophoresis of polypeptides of D virus and intracellular oncornavirus A revealed differences in the molecular weights of the components. No proteins with molecular weights of 10,000, 12,000, 15,000, and 27,000 dalton characteristic of the extracellular D virus were found in oncornavirus A virions. The analysis of protein patterns obtained in parallel experiments of isoelectric focusing and polyacrylamide gel electrophoresis suggests that oncornaviruses A and D of J-96 cells differ in the characteristics (pI and molecular weight) of the structural polypeptide components.

[Detection of human leukocyte elastase from a plasma alpha-1-proteinase inhibitor complex by its enzymatic activity with a synthetic substrate]. / Vyiavlenie leikotsitarnoi élastazy cheloveka iz kompleksa s plazmennym alph1-proteinaznym ingibitorom po ee énzimaticheskoi aktivnosti s sinteticheskim substratom.

A spectrophotometric procedure was developed for estimation of elastase activity in human leukocytes in the form of complex with blood serum alpha 1 proteinase inhibitor after loosening of the complex and detection of the enzymatic activity with N-tert-butyl-hydroxy-carbonyl-Ala-p-nitrophenyl ester as a substrate in presence of acetone or acetonitrile. The optimal conditions were described, which were required for estimation of the enzyme 100% activity followed by addition of the leukocyte elastase standard preparation into blood serum as well as for measurement of the enzyme activity in the complex with alpha 1 proteinase inhibitor. The procedure took 6-10 min to evaluate the elastase activity in the complex with the inhibitor using simple buffer containing organic solvents at 30 degrees and the available two-beam spectrophotometer.

[Effect of various infusion media and solutions on the kallikrein-kinin system of human plasma]. / Deistvie nekotorykh infuzionnykh sred i rastvorov na kallikrein-kininovuiu sistemu plazmy krovi cheloveka.

Activities of prekallikrein and Hageman factor (factor XII) were studied in some natural and artificial infusion media: polyglukine, rheoglumane, gemodes, polypher, gelatinol, saline; in solutions for parenteral nutrition--hydrolysate of casein, aminopeptide, aminokrovine, glucose solutions as well as in 10% NaCl and novocain. All the solutions studied did not affect the purified preparations of prekallikrein, but some of them activated partially purified preparations of Hageman factor. However, all these solutions, except of Ringer solution, activated Hageman factor and prekallikrein in human blood serum. Activation of prekallikrein appears to occur via the active form of Hageman factor. Among the solutions studied casein hydrolysate, 5% glucose, gelatinol and novocain exhibited the highest stimulating effect on Hageman factor and prekallikrein. Possible mechanisms of the activation are discussed.

[Pseudoexfoliative syndrome. Possible role of C-reactive protein and autoantibodies in the pathogenesis of the disease]. / Psevdoéksfoliativnyi sindrom. Vozmozhnoe uchastie C-reaktivnogo belka i autoantitel v patogeneze zabolevaniia.

Participation of acute inflammatory proteins (C-reactive protein, alpha-I-inhibitor of proteinases, C1q component of complement, IgG and IgM) in pathogenesis of pseudoexfoliative syndrome (PES) and, particularly of senile cataract was studied. In the course of the syndrome development blood vessel permeability was increased in the anterior chamber of the eye. Content of proteins in aqueous humor was increased 10-fold and even more as compared with normal state. High amount of proteins, main fractions of which were located at zones of 68,000, 37,000 and 20,500 D, were detected after polyacrylamide gel electrophoresis in presence of sodium dodecylsulfate. The zone of a protein with molecular mass of 20,500 D was found in presence of a reducing agent; in absence of the latter the zone was only slightly visible. The enzyme-labelled assay, used for detection of antigens in lens capsule, enabled to find such antigens as C-reactive proteins, IgG, IgM and less distinctly C1q component of complement.

[Comparative analysis of methods of determination of kallikrein and prekallikrein in the human plasma (review of the literature)]. / Sravnitel'nyi analiz metodov opredeleniia kallikreina i prekallikreina v plazme krovi cheloveka (obzor).

Main procedures developed for estimation of kallikrein activity and of content of its precursor are reviewed; critical comparative analysis of their principles is presented. Specificity, advantages and limitations of these procedures are discussed. General rules of enzymatic kinetics are considered in connection with development and application of these procedures. Some cautions are discussed considering wrong conclusions and recommendations, based on the data obtained by means of the inadequate methods.

[The use of a tripeptide chromogenic substrate for simultaneous determination of 4 indices of kallikrein-kinin system]. / Ispol'zovanie tripeptidnogo khromogennogo substrata dlia odnovremennoi otsenki chetyrekh pokazatelei aktivnosti kallikrein-kininovoi sistemy.

Evaluation of the kallikrein-kinine system activity in blood plasma is of importance due to participation of the components of this system in regulation of blood plasma proteolysis. Using D-Pro-Phe-Arg-pNA as a substrate, a procedure for simultaneous estimation of four patterns of the kallikrein-kinine system activity was developed: I. maintenance of contact phase, 2. content of prekallikrein (1.49 un/ml in normal state), 3. activity of kallikrein (0.104 un/ml), 4. antikallikrein potential (0.700 IU/ml).

[Participation of neutrophils in the pathogenesis of allergic inflammation: inhibitory-protease index in assessing and predicting atopic illnesses]. / Uchastie neitrofilov v patogeneze allergicheskogo vospaleniia: ingibitorno-proteaznyi indeks v otsenke i prognozirovanii atopicheskikh zabolevanii.

Total trypsin-like (BAEE esterase), elastase-like (BOC esterase) and antitryptic activities were studied in blood serum of patients with atopic diseases. The elastase-like activity was increased in blood serum of all the patients examined during the acute period of the disease; the enzyme activation depended on the pathology severity and clinical picture manifestations. The increase in blood plasma total proteolytic activity correlated with reverse alteration in blood antitryptic potential. The data obtained suggest that activation of neutrophils occurred in atopic diseases, thus the rate of elastase-like activity in blood might be used as an objective pattern in examination of patients and in checking of treatment course. The developed inhibitory-protease index may serve also as a criterion in evaluation of the pathological state severity.

[Disruption of regulatory possibilities of the plasma kallikrein-kinin system in chronic kidney failure and during hemodialysis]. / Narushenie reguliatornykh vozmozhnostei kallikrein-kininovoi sistemy plazmy krovi pri khonicheskoi pochechnoi nedostatochnosti i vo vremia gemodializa.

Activation of the kallikrein-kinin system was detected in chronic renal failure treated by routine therapy. In blood plasma of the patients with chronic renal failure, kallikrein was activated from 34.2 to 63.2 mU/ml, while activity of prekallikrein was decreased from 382.0 to 117.8 mU/ml; BAEE-esterase and elastase-like activities simultaneously increased from 340 +/- 20 to 464 +/- 43 mU/ml and from 150 +/- 20 to 340 +/- 18 mU/ml, respectively. In the patients on hemodialysis during the interdialysis period, there were reductions of BAEE-esterase and kallikrein activities to 265.8 +/- 18.5 and 25.9 mI/ml, respectively; and the content of prekallikrein remained decreased. During hemodialysis the elastase-like activity was markedly increased up to 501 mU/ml due to leukocyte activation. The findings suggest that during hemodialysis, due to leukocytic elastase, there is a possible occurrence of thromboses as a result of a potential depletion of the kallikrein-kinin system and decreased blood antiproteolytic potential, including antithrombin III.

[Effectiveness of commercial polyvalent proteinase inhibitors. Comparative analysis of their effect on kallikreins from different sources and trypsin]. / Effektivnost' deistviia kommercheskikh polivalentnykh ingibitorov proteinaz. Sravnitel'nyi analiz deistviia na kallikreiny razlichnogo proiskhozhdeniia i tripsin.

Polyvalent peptide-protein inhibitors of proteinases trasilol, contrical, hordox and pantrypine were studied using purified preparations of kallikreins from human blood plasma and hog pancreas as well as of trypsin. All the trade grade preparations of the inhibitors studied exhibited more distinct affinity to trypsin and tissue kallikrein as compared with blood plasma kallikrein. Trasilol inhibited most effectively the activity of trypsin, contrical--of pancreatic kallikrein. Contrical, trasilol and hordox inactivated blood plasma kallikrein at similar rates, pantrypine was several times less effective.

[Effectiveness of commercial polyvalent proteinase inhibitors. Use of gordox in vascular surgery]. / Effektivnost' deistviia kommercheskikh polivalentnykh ingibitorov proteinaz. Primenenie gordoksa pri khirurgicheskom vmeshatel'stve po povodu patologii sosudov.

Effect of hordox on activity of the kallikrein-prekallikrein system was studied in blood plasma of patients with stenosis of abdominal aorta during the reconstructive operation and in patients with septic complications after the open-heart operation. These impairments were accompanied by a distinct increase in kallikrein activity with simultaneous decrease in prekallikrein content, suggesting activation of the kininogenase system; the kallikrein activity was normalized in patients treated with the inhibitor. The highest efficiency of the inhibitor was observed in patients with stenosis of abdominal aorta; in patients with septic complications it was 4-fold less effective. The data obtained suggest that inhibition of the blood plasma kininogenase system correlates with restoration in these patients. The results, obtained in study of the inhibitor efficiency in vitro and in vivo, may be used for calculation of therapeutic doses of the proteinase inhibitors.

[Activity of the prekallikrein-kallikrein system and characteristics of its regulation in various allergies]. / Aktivnost' sistemy prekallikrein-kallikrein i osobennosti ee regulatsii pri nekotorykh allergicheskikh zabolevaniiakh.

Activity of kallikrein and content of prekallikrein were estimated in blood serum of 34 patients with atopy and of 17 patients with urticaria by means of the chromatographic procedure. In these patients activity of alpha 1-proteolytic inhibitor (alpha 1-PI) and alpha 2-macroglobulin (alpha 2-MG) was studied. At the acute period of pollinosis activation of the kallikrein-kinin system was found, which correlated with the disease aggravation. During specific immunotherapy of the patients with atopy activation of the kallikrein-kinin system occurred, which depended on the total concentration of allergen administered. At the same time, activation of the kallikrein-kinin system, observed under conditions of urticaria, was most distinct in the patients with chronic relapsing urticaria and was related to the degree of the disease aggravation. Preparations of proteinase inhibitors analogous to contrical were only short-term effective in chronic relapsing urticaria. In the patients with distinct aggravation of pollinosis inhibitory activity of alpha 2-MG was markedly increased which occurred apparently as a result of blood pachyemia simultaneously with activation of the kallikrein-kinin system. Distinct increase in the alpha 1-PI activity was not found in the patients with pollinosis and urticaria even at the step of pronounced aggravation. Phenotyping of the inhibitor in 10 patients with a marked decrease in its activity enabled to find 6 persons exhibiting the heterozygous genotype with a defect allele.

[The effect of leukocyte elastase on high molecular weight kininogen from human plasma in the presence of alpha-1 protease inhibitor. Analysis of proteolytic degradation]. / Deistvie leikotsitarnoi élastazy na vysokomolekuliarnyi kininogen plazmy krovi cheloveka v prisutstvii al'fa-1-proteinaznogo ingibitora. Analiz proteoliticheskoi degradatsii.

Degranulation of polymorphonuclear leukocytes (neutrophils) and releasing of leukocyte elastase during inflammation occur not only in injured tissue but in plasma in the presence of considerable excess of alpha-1 proteinase inhibitor (alpha-1PI). However, in spite of the absence of free elastase in patients' plasma, even in such severe inflammation as peritonitis and septicaemia, degradation of the connective tissue structures and plasma proteins may be determined. However the reasons of such destructive action are not yet determined. In this paper the action of leukocyte elastase on human plasma high molecular weight kininogen (HMWK) was studied in the absence or in the presence of different concentrations of alpha-1PI. The results showed that degradation of the intact molecules of HMWK occurred under the action of elastase during 1-2 hours of combined incubation even if the concentration of alpha-1PI in the mixture in 3-5 fold exceeds the molar elastase concentration. The rate of elastase inhibition by alpha-1PI in the presence of HMWK did not depend on an order of enzyme and inhibitor addition to the incubation medium. HMWK degradation by elastase in the presence of alpha-1PI was accompanied by impairments in its adhesion function although high tolerance of HMWK inhibitory activity with respect to SH-proteinases preserved. Thus, total inhibition of leukocyte elastase by alpha-1PI, in the presence of high molecular weight kininogen develops during relatively long time interval. The pronounced destruction of intact HMWK molecules takes place during this period of gradual elastase inhibition. This fact seems to be very important in pathogenesis of thrombo-haemorrhage syndrome as a complication of severe inflammation.

[Possible participation of angiotensin-converting enzyme and leukocyte elastase in the pathogenesis of insulin-independent diabetes mellitus]. / O vozmozhnom uchastii angiotenzin-prevrashchaiushchego fermenta i leikotsitarnoi élastazy v patogeneze insulin-nezavisimogo sakharnogo diabeta.

It is commonly accepted that the tolerance to insulin and hyperglycemia of the patients with non-insulin dependent diabetes mellitus (NIDDM) is due to some defect of insulin receptors or disturbances in the signaling pathway of the cell. This disease is often accompanied by hypertension. In this paper the high activity of plasma kallikrein-kinin system (KKS) (kallikrein activity was 6-8 times higher than normal), of angiotensin-converting enzyme (ACE) (4 times greater than normal), and of leukocyte elastase (2.7 times higher than normal) were demonstrated in plasma of patients with NIDDM. Increasing of KKS activity was coincident with rising of ACE activity, which may be the cause of the fast bradykinin inactivation and arising of hypertension. The treatment with ACE inhibitor during 3 months (4 mg of Perindopril per day) decreased ACE activity in patients' plasma which was accompanied with decreasing of the arterial pressure and some restoration of the carbohydrate metabolism indicators. The hyperinsulinemic euglycaemic clamping of 7 patients with NIDDM and essential hypertension showed that ACE-inhibitor (Perindopril, 4 mg) prevented bradykinin from destruction and increased the glucose consumption by tissues. The high activity of polymorphonuclear leukocytes and secretion of the elastase in NIDDM patients' plasma and/or instability of plasmatic and granular membranes of leukocyte in conditions of hyperglycaemic plasma are probably the cause of endothelial irritation and high ACE secretion. Secondly, the leukocyte may be the cause of injuring and decreasing of susceptibility of the cell receptors for insulin and bradykinin.

[Overall trypsin-like, elastase-like and antitryptic activity in patients with eye contusion]. / Obshchaia tripsinopodobnaia élastazopodobnaia i antitripticheskaia aktivnost' u patsientov s kontuziei glaza.

The investigations of tear fluid of eye after contusion injury revealed on increase of trypsin-like activity in 1-3 day and 10-19 day after trauma, the progressively elevation of elastase-like activity and lowering of inhibitory potential along 2-3 week. It was detected indirect correlation between the elastase-like activity and severity of the contusion injury of the eye, the grade of corneal edema, corneal erosion and conjunctival wound, hyphema. This results was shown the participation and the role of proteolytic and inflammatory processes in pathogenesis of eye blunt trauma. It was establish that the during of local inflammation is 2 week end more and is necessary antiinflammatory therapy with use the proteases inhibitors.

[Mechanism of activation of the kallikrein-kinin system in human plasma in stomach and duodenal ulcer]. / O mekhanizme aktivirovaniia kallikrein-kininovoi sistemy plazmy krovi cheloveka pri iazvennoi bolezni zheludka i dvenadtsatiperstnoi kishki.

Activity of kallikrein and content of prekallikrein were studied in blood serum of patients with gastroduodenal ulcer accompanied (or not accompanied) by hemorrhage. The rate of the kallikrein-kinin system activation was higher under conditions of the disease complicated by hemorrhage. Extracts of bioptic samples obtained from ulcerous zones as well as the extracts of leukocytes were shown to activate prekallikrein and Hageman factor in human blood plasma. At the same time, the activating effect of these extracts was distinctly lowered in the ulcer accompanied by hemorrhage. The phenomenon observed might occur due to high intensity of polymorphonuclear leukocytes degranulation during hemorrhage in the ulcer area.

[Proteolytic enzymes of the lacrimal fluid as pathogenesis factors of chronic corneal ulcer]. / Proteoliticheskie fermenty sleznoi zhidkosti kak faktory patogeneza khronicheskikh iazv rogovoi obolochki glaza.

Proteolytic enzymes, particularly collagenase, are involved in development of eye cornea chronic ulcers. Analysis of lacrimal fluid obtained from the patients enabled to find not only the collagenase activity but and serine enzymes exhibiting BAEE esterase activity. Plasmin, blood plasma and tissue kallikreins regulated permeability of capillaries in conjunctiva and lacrimal gland as well as they activated latent collagenase. Studies of BAEE esterase activity in lacrimal fluid of the patients are required for prescription of adequate pathogenetic treatment.