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Biomedical subjects

E Kaczmarek

Publications and source records attributed to E Kaczmarek.

At least 55 records · Page 3Linked to original sources
PubMed1996

Identification and characterization of CD39/vascular ATP diphosphohydrolase.

Vascular ATP diphosphohydrolase (ATPDase) is a plasma membrane-bound enzyme that hydrolyses extracellular ATP and ADP to AMP. Analysis of amino acid sequences available from various mammalian and avian ATPDases revealed their close homology with CD39, a putative B-cell activation marker. We, therefore, isolated CD39 cDNA from human endothelial cells and expressed this in COS-7 cells. CD39 was found to have both immunological identity to, and functional characteristics of, the vascular ATPDase. We also demonstrated that ATPDase could inhibit platelet aggregation in response to ADP, collagen, and thrombin, and that this activity in transfected COS-7 cells was lost following exposure to oxidative stress. ATPDase mRNA was present in human placenta, lung, skeletal muscle, kidney, and heart and was not detected in brain. Multiple RNA bands were detected with the CD39 cDNA probe that most probably represent different splicing products. Finally, we identified an unique conserved motif, DLGGASTQ, that could be crucial for nucleotide binding, activity, and/or structure of ATPDase. Because ATPDase activity is lost with endothelial cell activation, overexpression of the functional enzyme, or a truncated mutant thereof, may prevent platelet activation associated with vascular inflammation.

Adenosine Triphosphatases↗
PubMed1995

Biosynthesis of plasma factor XIII: evidence for transcription and translation in hepatoma cells.

Factor XIIIa belongs to a family of ubiquitous transglutaminases, which catalyze formation of covalent bonds between the epsilon-amino group of specific lysines and the gamma-carboxyl group of glutamines. Factor XIII is synthesized as a zymogen and after activation, it participates in both the coagulation and fibrinolytic mechanisms. Most transglutaminases are intracellular, but factor XIII is both intracellular and extracellular. the biosynthesis of extracellular (plasma) factor XIII, with the structure of a noncovalent heterotetramer, A2B2, is complex. Here, evidence is presented from PCR analysis and Northern blotting that mRNAs for both A and B subunits are present in the liver. The distribution of mRNA, specific for factor XIII subunits, in various human tissues was also analyzed. Among the tissues examined, the only signal for B subunit was found in the liver. For subunit A, the signal was observed in placenta, liver, kidney, lung, skeletal muscle and heart with varying intensities; in brain or pancreas there was no signal. With an immunoperoxidase method, factor XIII A subunit was identified in the PLC/PRF/5 cell line. By ELISA and reverse immunoblotting, with antibodies specific for the A-B complex, it was also shown that these cells produce and secrete factor XIII. From all of these results, we conclude that the liver is a source of plasma factor XIII, and that the complex A2B2 is secreted from these cells.

Base Sequence↗
PubMed1994

Morphometric analysis in the diagnosis and differentiation of certain glomerulopathies (MCD, GNMes, FSG)

Morphometric analysis was used to evaluate mesangial components in MCD (minimal changes disease), GNMes (mesangial glomerulonephritis), FSG (focal segmental glomerulosclerosis). In GNMes the increase of matrix was found to be generally proportional to the amount of mesangial cells. There are clear statistically significant differences in the ratio of matrix volume to cell component volume and of matrix volume to the whole mesangial area in MCD and GNMes as compared with FSG. In the case of GNMes where morphometric results resemble those in FSG cautious prognosis is recommended as there is a possibility of FSG. This has been confirmed both by the course of the disease and the results of repeated biopsy.

Adult↗
PubMed1993

Physical characterization of recombinant tissue plasminogen activator.

Electron microscopic and physical-chemical properties of one- and two-chain tissue plasminogen activator (t-PA) were studied. The molecular weight of one-chain t-PA obtained by both sedimentation equilibrium and SDS-PAGE was estimated to be about 65,000, while both chains in the reduced two-chain form were in the range of 35,000-40,000. Sedimentation coefficients were identical for both forms of t-PA (S(0)20,w = 4.12). The two forms of t-PA were indistinguishable by electron microscopic analysis, which confirmed the sedimentation results, and showed that they were ellipsoidal and relatively compact. The major and minor axes were approx. 13 nm and approx. 10 nm and f/f0 was 1.36. The individual domains of t-PA are relatively small and are folded within the molecule, so that the overall appearance is globular.

Centrifugation↗
PubMed1993

Initial interaction between fibrin and tissue plasminogen activator (t-PA). The Gly-Pro-Arg-Pro binding site on fibrin(ogen) is important for t-PA activity.

Gly-Pro-Arg-Pro (GPRP) is a potent inhibitor of fibrin polymerization. It also causes a concentration-dependent inhibition of the activation of plasminogen by t-PA when soluble desAABB-fibrinogen (fibrin II) or fibrinogen are used as promoters of t-PA. Fibrinogen has a much weaker promoter activity than does fibrin II. Experiments were undertaken to explain the mechanism of action of GPRP on plasminogen activation. Kinetic data indicated that when GPRP was present in the assay system, the fibrin(ogen)-GPRP complex was ineffective as a t-PA promoter. Only the free forms of fibrin II or fibrinogen were promoters of t-PA. GPRP specifically eluted t-PA, which was previously bound to a fibrin-Sepharose column, and also inhibited t-PA binding to fibrin-Sepharose in a concentration-dependent manner. These experiments were compared to those with other tetrapeptides: GHRP, GPGG, and GRGD. Only GHRP, which is known to bind more weakly to fibrin(ogen) than GPRP, had any effect. These results suggest that the GPRP binding site on the fibrin(ogen) molecule is important for t-PA activation and is a binding site for t-PA in the initial interaction between t-PA and fibrin. We propose that at or near the GPRP binding site on fibrin there is an initial binding site for t-PA. We hypothesize that kringle 1, which contains the sequence G128RRP, may be involved in the initial binding of t-PA to fibrin.

Amino Acid Sequence↗
PubMed1993

Hypertrophy or hyperplasia in cardiac muscle. Post-mortem human morphometric study.

In 103 hearts with various forms of cardiac muscle hypertrophy the following parameters were estimated: diameter, length, volume, density and number of myocytes, and density of myocyte nuclei. The values of all histometric parameters correlated well with left ventricular (LV) weight up to 350 g. In heavier hearts these parameters remained approximately of the same magnitude. The number of myocytes was significantly higher in hearts with LV weight above 250 g. The influence on LV weight of age, coronary artery diameters, degree of atherosclerosis, weight and percent of fibrous tissue was also evaluated. On the basis of a linear discriminant function, hearts were divided into three classes: (1) LV weight < or = 250 g (absence of hyperplasia, hypertrophy only); (2) LV weight 251-350 g (hypertrophy+signs of hyperplasia); (3) LV weight > 350 g (marked signs of hyperplasia). The percent of fibrosis increased proportionally to LV weight. Where LV weight was above 250 g there was a subsequent increase in the mean percent of fibrosis (approx. 26%). This phenomenon (plateau of percent fibrosis) is the result of an increased number of myocytes (myocyte hyperplasia). We suggest that, independent of aetiology, in all hearts above 350 g (patients with congestive heart failure) hyperplasia phenomenon exists.

Adult↗
PubMed1993

Performance characteristics of fibrinogen assays. Results of the College of American Pathologists Proficiency Testing Program 1988-1991.

Plasma fibrinogen concentration appears to be an important risk factor for the development of atherosclerotic cardiovascular disease of a similar magnitude to cholesterol. The quality control of plasma fibrinogen assays has taken on new importance as a consequence of this potential role as an atherosclerotic risk factor. This article reviews the performance characteristics of 40,000 fibrinogen assays comprising the College of American Pathologists Proficiency Testing Program from 1988 through 1991. Instrument and reagent variables both play roles in the poor interlaboratory reproducibility documented by this study. The absence of either a national or international standard for plasma fibrinogen assays has been a major source of reagent variability. The validation and calibration of the College of American Pathologists lyophilized plasma reference preparation for fibrinogen determination is also reported in this study. The availability of this validated reference plasma should markedly improve interlaboratory reproducibility.

Analysis of Variance↗
PubMed1992

[The role of the nucleus in the mechanism of transformation of muscle after infection by Trichinella spiralis larvae. III. Morphometric analysis of the functional transformation of the muscle cell nucleus after infection].

Morphometric investigations (using the convergence analysis method) of geometric parameters of nuclei and nucleoli of transformed muscle cells were carried in mice 15 days, 30 days and 6 month after infection with T. spiralis larvae. The analysis showed the largest increase of investigated the parameters of 15th day after the infection. The results of morphometric analysis are in agreement with the morphologic, ultrastructural and histochemical observations (published in I and II part) as for as functional changes of muscle cells in various stages of trichinellosis are concerned.

Animals↗
PubMed1992

[Hypertrophy or hyperplasia of myocytes in heart hypertrophy?].

One of the most controversial problem in cardiac muscle pathology is the existence of myocyte hyperplasia. The term hypertrophy indicates an increase in size of the individual muscle cells without changing their total number, whereas in hyperplasia there occurs proliferation of the myocyte. This fundamental question of the character of cardiac growth remains unresolved in spite of the wide attention it has received. Contemporary views concerning the cardiac muscle hyperplasia are presented. From clinical point of view the problem is significant for two reasons. The loss of the ability of muscle cells to proliferate is responsible for the irreversible myocardial destruction after injury. From another point of view, if the increase of the heart muscle is maintained, although a complete remission of cardiac hypertrophy becomes impossible. In 103 hearts with various forms of cardiac muscle hypertrophy the following parameters were estimated: diameter, length, volume, density and number of myocytes, as well as the density of nuclei of myocytes. The values of all histometric parameters correlated well with the LV weight up to 350 g. In heavier hearts these parameters were approximately at the same magnitude. The number of myocytes was significantly higher in hearts with LV weight above 250 g than in hearts below 250 g: 5.53 x 10(9) vs 4.31 x 10(9), p < 0.001. The influence of coronary artery diameters, degree of atherosclerosis, weight and percent of fibrous tissue and age on LV weight were evaluated as well. From these parameters only coronary artery diameters significantly influenced on LV weight.(ABSTRACT TRUNCATED AT 250 WORDS)

Cardiomegaly↗
PubMed1991

Fibrinogen Ledyard (A alpha Arg16----Cys): biochemical and physiologic characterization.

Fibrinogen Ledyard was discovered in a 10-year-old boy with a mild bleeding history. His father had the same defect and a bleeding history after surgery. Both patients were heterozygous. The plasma fibrinogen concentration was normal immunologically (335 mg/dL) and very low functionally (52 mg/dL). Purified fibrinogen Ledyard had a prolonged polymerization, which was somewhat corrected by addition of Ca2+ ions. High performance liquid chromatography (HPLC) analyses of the fibrinopeptides released by thrombin showed 1 mol of fibrinopeptide A (FPA) and 2 mol of fibrinopeptide B (FPB) released per mole of fibrinogen Ledyard. Steady-state kinetic parameters were evaluated for release of FPA by thrombin. When the concentration of fibrinogen Ledyard was corrected to 50% of total protein, because only 50% of fibrinogen Ledyard can release FPA, the kinetic constants were similar to those of control fibrinogen (Km = 7.5 mumol/L for A alpha chain, kcat = 54 s-1). This finding indicates that the cleavage site of the A alpha chain in these abnormal molecules may not interact with the catalytic site of thrombin. The three chains of fibrinogen Ledyard were isolated on reverse-phase C4-HPLC. The sequence of the amino terminus of A alpha chain showed that Arg in position 16 was replaced by Cys in the abnormal molecules. Approximately half of fibrinogen Ledyard (52%) was clotted by reptilase, suggesting that fibrinogen Ledyard may consist of 50% normal homodimers (A alpha Arg16 . A alpha Arg16) and 50% abnormal homodimers (A alpha Cys16 . A alpha Cys16). Abnormal molecules could form disulfide bond between the A alpha Cys16 residues. Thus, the abnormal molecules have a different structure that does not bind to thrombin. Probably the abnormality of polymerization of fibrinogen Ledyard results from the interaction of the abnormal molecules with normal fibrin monomers, so that the growth of fibrin protofibrils is inhibited. This abnormal fibrinogen supports adenosine diphosphate-induced platelet aggregation in a normal manner.

Amino Acid Sequence↗
PubMed1990

[Myocardial structure in various forms of hypertrophy. I. Myocyte hypertrophy or hyperplasia? Pathophysiological aspects and the methods of study].

One of most controversial problems in cardiac muscle pathology is the existence of myocyte hyperplasia. The term hypertrophy indicates an increase in size of the individual muscle cells without changing their total number, whereas in hyperplasia proliferation of the myocyte occurs. This fundamental question of the character of cardiac growth remains unresolved in spite of the wide attention in has received. Contemporary views concerning the cardiac muscle hyperplasia are presented. From clinical point of view the problem is significant for two reasons. The loss of the ability of muscle cells to proliferate is responsible for the irreversible myocardial destruction after injury. From another point of view, if the increase of the heart muscle is maintained, although a complete remission of cardiac hypertrophy becomes impossible. In the light of these controversies we undertook the attempt to solve this complicated problem. Morphometric evaluation of the cardiac myocyte is described in detail.

Anthropometry↗
PubMed1990

[Myocardial structure in various forms of hypertrophy. II. Myocyte hypertrophy or hyperplasia? Results of the study].

In 103 hearts with various forms of cardiac muscle hypertrophy the following parameters were estimated: diameter, length, volume, density and number of myocytes, as well as the density of nuclei of myocytes. The values of all histometric parameters correlated well with the LV weight up to 350 g. In heavier hearts these parameters were approximately at the same magnitude. The number of myocytes was significantly higher in hearts with LV weight above 250 g than in hearts below 250 g: 5.53 x 10(9) vs 4.31 x 10(9), p less than 0.001. The influence of coronary artery diameters, degree of atherosclerosis, weight and percent of fibrous tissue and age on LV weight were evaluated as well. Only coronary artery diameters significantly influenced on LV weight. On the basis of linear discriminant function, three classes of hearts were separated: 1) LV weight 250 g - absence of hyperplasia, only hypertrophy 2) LV weight 251-350 g - hypertrophy + signs of hyperplasia 3) LV weight 350 g - marked signs of hyperplasia Among 18 patients with the LV weight above 350 g (all patients with congestive heart failure), 11 suffered from valvular disease, 3 were postinfarction patients, 2 suffered from primary hypertension and 2 from primary congestive cardiomyopathy. It indicates that, irrespective to the etiologic factor, hyperplasia is a simple result of the cardiac muscle mass increase.

Adult↗
PubMed1988

Thrombin binding to the A alpha-, B beta-, and gamma-chains of fibrinogen and to their remnants contained in fragment E.

In order to study thrombin interaction with fibrinogen, thrombin binding to fragments D and E (prepared by plasmin digestion of fibrinogen) and to intact S-carboxymethylated chains of fibrinogen (A alpha, B beta, and gamma) was analyzed by autoradiography, immunoblotting, and affinity chromatography. Complex formation was observed between late fragment E and thrombin but not with fragment D. The three reduced chain remnants of fragment E all formed complexes with thrombin. Also, thrombin bound to the intact, separated A alpha, B beta, and gamma chains of fibrinogen as well as to the alpha and beta chains of fibrin. In these experiments the extended substrate-binding site, but not the catalytic-binding site, was being examined because fragment E had as its amino-terminal amino acids Val20 in the alpha chain, Lys54 in the beta chain, and Tyr1 in the gamma chain. Also, thrombin inhibited in its active center by D-phenyl-alanyl-L-prolyl-L-arginine-chloromethyl ketone bound to fragment E and to the separated chains in the same manner as unmodified thrombin. A lysine residue to thrombin was essential for its binding to fibrinogen. Thrombin attached to CNBr-activated Sepharose through its amino groups did not bind to fragment E, but when thrombin was attached through its carboxyl groups, it bound fragment E.

Amino Acid Sequence↗
PubMed1988

Alloantibodies, autoantibodies, and immune complexes in patients with lung cancer.

The sera of patients with lung cancer, nonmalignant lung disease, and blood donors were subjected to various immunologic assays. Nine assays, based on immunoradiometric (IRMA) and immunoenzymatic (ELISA) principles, included 3 types of fetal cell antibodies, 2 established lung cancer cell antibodies, anti-DNA, anti-IgG autoantibodies, and immune complex assays based on C1q binding and anti-C3 activity. Antitumor cell antibody level was significantly lower in patients with lung cancer compared to blood donors. In the remaining 7 assays, the lung cancer patients tended towards higher median values compared to both control patients and blood donors, but without statistical significance, with the exception of anti-DNA antibodies. Statistical analysis of all 9 assays taken together has shown significant differences between the 3 groups. When only 5 assays were used to assess 3 types of fetal cell antibodies, anti-DNA antibodies, and immune complexes by means of ELISA anti-C3, the margins between groups increased. A range of values for the selected assays was established that may discriminate 70% of tested individuals of the 3 groups. These results suggest the existence of a characteristic profile of deranged humoral immunity in lung cancer patients.

Adenocarcinoma↗
PubMed1987

Fibrinogen-sepharose interaction with prothrombin, prethrombin 1, prethrombin 2 and thrombin.

Binding of prothrombin, prethrombin 1, prethrombin 2 and thrombin to fibrinogen-Sepharose was studied. Thrombin and prethrombin 2 bound to fibrinogen-Sepharose, while prethrombin 1 and prothrombin did not. Bound thrombin and prethrombin 2 were recovered from the column by eluting with 0.1 M NaCl/0.05 M Tris-HCl buffer (pH 7.4). The affinity of thrombin and prethrombin 2 to fibrinogen-Sepharose depended on ionic strength and reached a maximum at 50 mm concentration. Prethrombin 2 interacts with fibrinogen as well as thrombin; and prothrombin fragment 1.2 is not important in the formation of this complex. Thus, prethrombin 2, which is a precursor of thrombin without measurable enzymatic activity and which lacks the single cleavage at Arg-322-Ile-323 present in thrombin, has the same or very similar structural conformation as thrombin and has the same macromolecular substrate recognition site. These results confirm the earlier results that active center is not necessary in fibrinogen-thrombin interaction.

Amino Acid Sequence↗
PubMed1985

Distribution of different cell types within the rat thymus in the neonatal period of life.

This study attempted to define reciprocal positions of cell types within the thymus. Random or non-random contacts between specific cell types were analyzed by means of graph theory. For analysis, thymus blocks were sectioned serially and, then, thymus cells were categorized into types, based on morphological criteria. The distribution of individual cell types within the cortex, cortico-medullary zone and medulla was presented in form of a map. In the analysis, three types of epithelial cell, characteristic of each thymus zone, macrophages, Langerhans-like cells and lymphocytes were found in non-random relations to one another. Moreover, characteristic groups of cells associated with one another were also demonstrated.

Animals↗