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W Uracz

Publications and source records attributed to W Uracz.

At least 19 recordsLinked to original sources
PubMed2002

Interleukin 1beta induces functional prostaglandin E synthase in cultured human umbilical vein endothelial cells.

Prostaglandin endoperoxide H2 (PGH2) is generated from arachidonic acid by either constitutive (COX-1) or inducible (COX-2) cyclooxygenases. In arterial wall PGH2 is converted by PGI2 synthase (PGI-S) to prostacyclin (PGI2), and in platelets by thromboxane synthase (TX-S) to thromboxane (TXA2). Other prostanoids as PGD2, PGF2alpha, or PGE2 were believed to arise non-enzymatically from PGH2. Only recently, human prostaglandin E synthase (PGE-S) has been identified and cloned as a membrane bound, microsomal, glutathione-dependent inducible enzyme. Here we demonstrated that interleukin 1beta (IL-1beta) is an inducer of COX-2 and PGE-S in human umbilical vein endothelial cells (HUVEC). Functional expression of PGE-S was measured at the level of specific mRNA by semi-quantitative RT-PCR, PGE-S protein was detected by Western blot in HUVEC, while PGE2 was measured by immunoassay in the supernatant. Actinomycin D, a classical transcription inhibitor, was used to prove that indeed IL-1beta induced the functional PGE-S enzyme. PGE2 generation in HUVEC was inhibited by indomethacin, acetaminophen and dexamethasone. In conclusion, we found that in cultured endothelial cells IL-1beta induced as evidenced by the appearance of its transcript and its functional enzyme. The induction of endothelial PGE-S and COX-2 appeared to be and their transcripts were induced as fast as one might expect from immediate early genes. It means that IL-1beta-triggered-PGE2 biosynthesis in endothelial cells is probably regulated by induction of both COX-2 and PGE-S. This is way we hypothesise the existence of at least two distinct pools of COX-2: the first selectively coupled to PGE-S and the second one that is coupled to PGI-S yielding the main endothelial product--PGI2.

Blotting, Western↗
PubMed2001

Comparison of endothelial pleiotropic actions of angiotensin converting enzyme inhibitors and statins.

Two in vitro and one in vivo assay were performed to study the endothelial pleiotropic actions of "tissue type" angiotensin converting enzyme inhibitors (ACE-Is) such as perindopril and quinapril, their active forms, that is, quinaprilat and peridoprilat, or of statins belonging to natural (lovastatin), semisynthetic (simvastatin), and synthetic enantiomeric (atorvastatin, cerivastatin) classes. Cytoplasmic [Ca2+]i levels in cultured bovine aortic endothelial cells and endothelium-dependent nitric oxide-mediated coronary vasodilatation in the Langendorff preparation of guinea pig heart constituted our in vitro assays. The in vivo assay consisted of study of PGI2-mediated thrombolytic response in arterial blood of rats after intravenous administration of drugs. In this last assay, perindopril and quinapril proved to be, by two orders of magnitude, more potent PGI2-dependent thrombolytics than the most potent statin (atorvastatin). However, in both in vitro assays we found a higher endothelial efficacy of statins as compared to ACE-Is. In particular, those statins that contain the lactone ring in their molecules (lovastatin, simvastatin) were the most potent coronary vasodilators. In summary, the in vivo profile of action of ACE-Is and statins contrasted with their reversed order of potency in vitro. We hypothesize that the endocrine-like function of the pulmonary circulation [28-31] may be responsible for the in vivo bradykinin-triggered, PGI2-mediated thrombolysis by ACE-Is, whereas the pleiotropic action of statins, possibly involving inhibition of prenylation [14-19], is diffused throughout many vascular beds.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed2000

Thrombolysis by thienopyridines and their congeners.

We propose that anti-platelet thienopyridines, such as ticlopidine or clopidogrel, are thrombolytic owing to endothelial release of prostacyclin (PGI2) and tissue plasminogen activator (t-PA). In this study we used anaesthetised Wistar rats with extracorporal circulation in which arterial blood superfused thrombi which adhered to a strip of collagen. Weight of thrombi was continuously monitored. When administered intravenously, clopidogrel or its R enantiomer deprived of anti-platelet action, both at doses of 3 mg x kg(-1), produced lost in weight of thrombi by 14.1 +/- 1.3% or 16.0 +/- 1.4% (n = 9), and at doses 10 mg x kg(-1) by 28.3 +/- 2.3% or 30.4 +/- 1.9% (n = 8), respectively. Maximum of thrombolysis occurred 30-45 min following the drug administration. Ticlopidine at a dose of 30 mg x kg(-1) reduced weight of thrombi by 33.7 +/- 1.7% (n = 32). Thrombolytic action of ticlopidine was accompanied by a rise in 6!keto-PGF1alpha blood levels from 0.42 +/- 0.10 to 1.58 +/- 0.29 ng x ml(-1) and t-PA antigen plasma levels from 4.70 +/- 1.00 to 12.90 +/- 1.15 ng x ml(-1) (n = 7). Five out of eleven tested thienopyridine congeners with pyrimidine or pyrimidinone instead of pyridine rings had thrombolytic potencies similar to that of clopidogrel (ED30s at a range of 6.2-11.4 mg x kg(-1)). A substantial increase in thrombolytic potency (ED30s at a range of 0.3-2.1 mg x kg(-1)) was observed for congeners in which thienyl ring was condensed with an additional cyclopentyl, cyclohexyl or cycloheptyl structures or in which thienopyridine complex was replaced for a pyridopyrimidine one. We claim that thienopyridines, independently of their delayed anti-platelet action, do produce immediate thrombolysis in vivo. This new activity emulates capacity of their native, non-metabolised molecules to release prostacyclin and tissue plasminogen activator. We have also shown that structural changes in molecules of thienopyridines may intensify their thrombolytic potency.

Animals↗
PubMed2000

The protective role of nitric oxide in the brain ischemia.

A role of nitric oxide in ischemia/reperfusion (I/R) injury of brain in normotensive (Sprague-Dowley rats, SD) and stroke-prone spontaneously hypertensive rats (SHR-SP) was studied. Cerebral ischemia was produced in rats by occlusion of the middle cerebral artery (MCA). NO and O2- releases in the brain in response to MCA occlusion followed by reperfusion were simultaneously monitored (2h) using electrochemical microsensors. The size of infarct was evaluated in the course of I/R from images of brain slices stained with 2,3,5-triphenyltetrazolium chloride. Similar patterns of NO and O2- releases were exhibited for SD and SHR-SP rats in the entire course of the experiments. However, the concentration of NO release was significantly lower during I/R in SHR-SP than in SD rats (the maximal NO concentration was 2.61 +/- 0.22 micromol/L for SD and 1.51 +/- 0.16 micromol/L for SHR-SP rats; *P < 0.01). In contrast, the concentration of O2- release during cerebral ischemia was significantly higher in SHR-SP than SD rats (the maximal increase was 122 +/- 24 nmol/L for SD and 220 +/- 44 nmol/L for SHR-SP rats; *P<0.01). The infarct sizes revealed in the course of I/R were larger in SHR-SP than SD rats (1.8 +/- 0.4% vs. 1.1 +/- 0.4% at 30 min., 2.84 +/- 0.8% vs. 2.21 +/- 0.6% at 100 min. and 9.20 +/- 1.1% vs. 5.8 +/- 0.6% at 180 min. ofthe brain weights, respectively; *P < 0.01 for each time-point). These studies indicate that nitric oxide plays a protective role during I/R and deficiency of NO in SHR-SP rats is due to excess of O2- production. The deficiency in NO concentration correlates positively with the increase of cerebral I/R injury.

Animals↗
PubMed1999

Effect of ticlopidine on streptokinase-induced thrombolysis in rats.

Using our original assay system we found that ticlopidine (TP, 30 mg/kg i.v.) had produced a prompt thrombolysis of preformed clots in extracorporeal circulation of anaesthetized rats. In contrast with ticlopidine streptokinase (SK, 30,000 U/kg i.v.) proved thrombogenic for an initial period of 10-30 min, followed only later by the expected thrombolytic action. In ticlopidine pretreated rats the thrombogenic effect of streptokinase was eliminated, and its thrombolytic potency intensified. This is the first experimental support for a conception that a combined therapy with ticlopidine and streptokinase may restrict the "early hazard" associated with streptokinase therapy.

Animals↗
PubMed1999

Thienopyridines: effects on cultured endothelial cells.

In cultured endothelial cells harvested from human umbilical vein (HUVEC) or bovine aorta (BAEC) the 30 min incubation with calcium ionophore A 23187 (1 microM) or ticlopidine (100 microM) caused an increase in nitrite generation in HUVEC from basal 227 +/- 37 to 372 +/- 60 or to 325 +/- 33 pmoles per 10(6) cells, respectively, and in BAEC from basal 182 +/- 17 to 378 +/- 18 or to 423 +/- 66 pmoles per 106 cells (n = 6), respectively. Calcium ionophore A 23187 (1 microM) or ticlopidine (100 microM) next to 30 min incubation with BAEC increased release of 6-keto-PGF 1alpha from basal level of 9.4 +/- 1.8 to 96.2 +/- 5.1 or to 99.5 +/- 10.2 pmoles per 10(6) cells, respectively. The pretreatment with aspirin (300 microM) cut down this rise to 4.2 +/- 0.1 pmoles per 10(6) cells (n = 8). Basal cytoplasmic calcium levels, [Ca2+]i, in immortalised HUVEC cell line - ECV304, HUVEC and BAEC were 47.7 +/- 3.3 nM (n = 53), 68.3 +/- 5.0 nM (n = 30) and 53.1 +/- 3.0 nM (n = 15), respectively. In these cultured endothelial cells calcium ionophore A 23187 (0.1 microM) produced net maximum rise in [Ca2+]i by 157 +/-27 nM (n = 16)[ ECV304], by 107 +/- 58 nM (n=4) [HUVEC], and by 231.0 +/- 41.3 nM (n = 8) [BAEC], respectively, while ticlopidine (30 microM) produced net maximum rise in [Ca2+]i by 30.0 +/- 3.2 nM (n=9)[ECV304], 48.8 +/- 15.6 nM (n = 4)[HUVEC] and 28.4 +/- 5.4 nM (n = 8)[BAEC], respectively. Effect of ticlopidine on [Ca2+]i was not only weaker than that of calcium A 23187 but also its maximum appeared after a lag period that was 2 3 times longer than that for A23187. In ECV304 clopidogrel at concentrations of 10, 30 and 100 microM produced maximum increment of [Ca2+]i by 16.5 +/- 3.8 nM (n = 7), 47.0 +/- 6.9 nM (n = 8) and 67.2 +/- 8.3 nM (n = 8), respectively. Incubation of BAEC with A23187 (microM), ticlopidine or clopidogrel (100 microM) for 2 h did not influence viability of cultured endothelial cells. We claim that thienopyridines, independently of their delayed anti-platelet properties ex vivo do release NO and PGI2 from cultured endothelial cells in vitro. The above endothelial action of thienopyridines might be mediated by a rise in [Ca2+]i, however, this possibility has not been proved.

6-Ketoprostaglandin F1 alpha↗
PubMed1998

Protective role of pulmonary nitric oxide in the acute phase of endotoxemia in rats.

We present for the first time direct continuous assay of NO concentration (porphyrinic sensor) in the lung parenchyma of Sprague-Dawley rats in vivo during endotoxemia. Intravenous infusion of lipopolysaccharide (LPS, 2 mg x kg(-1) x min(-1) for 10 minutes) stimulated an acute burst of NO from constitutive NO synthase (NOS) that peaked 10 to 15 minutes after the start of LPS infusion, mirroring a coincident peak drop in arterial pressure. NO concentration declined over the next hour to twice above pre-LPS infusion NO levels, where it remained until the rats died, 5 to 6 hours after LPS infusion. The chronic drop in arterial pressure observed from 70 minutes to 6 hours after the start of LPS infusion was not convincingly mirrored by a chronic increase in NO concentration, even though indirect NO assay (Griess method, assaying NO decay products NO2-/NO3-) showed that NO production was increasing as a result of continuous NO release by inducible NOS. A NOS inhibitor, N(omega)-nitro-L-arginine (L-NNA, 10 mg/kg i.v.) injected 45 minutes before LPS infusion, resulted in sudden death accompanied by macroscopically/microscopically diagnosed symptoms similar to acute respiratory distress syndrome <25 minutes after the start of LPS infusion. Pharmacological analysis of this L-NNA+LPS model by replacing L-NNA with 1-amino-2-hydroxy-guanidine (selective inhibitor of inducible NOS) or by pretreatment with S-nitroso-N-acetyl-penicillamine (NO donor), camonagrel (thromboxane synthase inhibitor), or WEB2170 (platelet-activating factor receptor antagonist) indicated that in the early acute phase of endotoxemia, LPS stimulated the production of cytoprotective NO, cytotoxic thromboxane A2, and platelet-activating factor.

Acute Disease↗
PubMed1993

Detection, by in situ hybridization using sulphonated cDNA probe, the specific mRNA for HLA-DR alpha induced in monocyte cell lines by recombinant interferon gamma.

We adopted the nonradioactive method used for blot hybridization for the detection of inducible mRNA for HLA-DR alpha by the in situ hybridization. Unstimulated and interferon gamma stimulated MonoMac6 and U937 human monocytic cell lines were used as target cells. Sulphonation of plasmid pBR322 with HLA-DR alpha cDNA insert (2 x 700 bp, in Pstl restriction site) was performed according to the manufacturer's procedure (SulfoProbe Kit, Sigma). The hybridization signals were detected with mouse monoclonal, anti-sulphonated DNA antibody, followed by immunovisualization with anti-mouse IgG-alkaline phosphatase conjugates. Unstimulated MonoMac6 and U937 cells showed few granular reaction products only in small percentage of cells (1-5%), while in IFN gamma stimulated cells the fine granular immunoenzymatic reaction was observed in the cytoplasm of majority of cells (> 80%). This method seems to be easy and rapid to perform, making it applicable for routine diagnostic purposes in tissue sections and biopsies.

Blotting, Southern↗
PubMed1992

Augmentation of monocyte-mediated cytocidal activity by a low dose tumour necrosis factor measured by the kinetic colorimetric microplate assay.

This paper describes a simple kinetic colorimetric assay for the quantitation of human peripheral blood monocyte-mediated cytotoxic activity against tumour cells. Isolated effector monocytes were cultured overnight with an increasing number of target cells in 96-well microplates. Cytotoxic activity of monocytes was determined by modified nitroblue tetrazolium (MTT) dye assay using standard ELISA reader offering possible automation. The test was performed with three different effector/target cell ratios using a fixed number of monocytes. This allowed the expression of cytotoxic activity of monocytes in cytotoxic activity units. The assay was found to be a simple method to demonstrate that low doses of TNF (1 U/ml) enhanced monocyte-mediated cytotoxicity.

Colorimetry↗
PubMed1992

Effect of peritoneal macrophages from intermittent peritoneal dialysis patients on lymphocytes in culture.

To investigate the biological activity of peritoneal macrophages, cells isolated from dialysate of 30 patients with end-stage kidney disease treated by intermittent peritoneal dialysis and from ascites of 6 patients with cardiac insufficiency (relative control group) were added to autologous, phytohemagglutinin (PHA)-stimulated lymphocyte cultures. Macrophages of dialyzed patients induced a dose-dependent increase in autologous lymphocyte proliferation, whereas macrophages obtained from control subjects exerted a suppressive effect on those cultures. The enhanced lymphocyte proliferation by macrophages from dialyzed patients was corroborated by the increased metabolic activity of macrophages as evaluated by the increased nitro blue tetrazolium (NBT) reduction test and increased functional expression of Fc receptors (FcR). The subpopulation of macrophages from patients with HLA-DR antigens as determined by HB55 monoclonal antibody, inhibited lymphoproliferation in vitro. We conclude that peritoneal macrophages from dialyzed patients represent a heterogenous population of cells with different phenotypic and functional characteristics.

Adult↗
PubMed1991

Functional characteristics of peritoneal macrophages of renal failure patients on peritoneal dialysis.

Functional activity of peritoneal macrophages of 50 patients with end-stage renal failure on intermittent peritoneal dialysis (IPD) and of 30 control subjects with normal renal function was determined. Phagocytosis of latex particles by macrophages of dialyzed patients was significantly lower as compared with the controls. Further depression of the phagocytic activity was observed during bacterial peritonitis. Macrophages from the dialyzed patients also showed nonsignificantly decreased functional expression of Fc receptors (FcR) and increased spontaneous nitro blue tetrazolium (NBT) reduction.

Adolescent↗
PubMed1989

The T cell receptor in the mouse.

In the eighties the genes of the T lymphocyte antigen receptor (TcR) have been cloned. The search for the elusive and controversial TcR is over. It is clear that genes for TcR are distinct from those of Ig genes although these genes belong to the immunoglobulin supergene family. The aim of this article is to scan briefly a history of the searching for TcR, than to update our knowledge about TcR. With the search for TcR behind, a new set of question and goals lies ahead. How does TcR, which is similar to Ig, recognize foreign antigen in association with major histocompatibility complex (MHC) gene products? What is the relationship between TcR and Iat molecules, including misterious I-J product?

Animals↗
PubMed1988

The altered expression of MHC-class II determinants on monocytes of cancer patients.

The expression of MHC class II determinants Ia.7 (detected by cross reactive mouse anti-Iak antibody) and HLA-DR on monocytes (MO) of gastric and colorectal cancer patients was examined. An increased proportion of MO bearing the Ia.7 determinant was found, while the number of MO expressing DR was not elevated. In gastric cancer patients the increased expression of the Ia.7 determinant was most pronounced in advanced cancer (stage IVA and IVB). The increased expression of this determinant was related to the presence of the tumour as the number of MO expressing Ia.7 decreased 6 months following surgical resection of the tumour. Further, the increased expression of Ia.7 on MO correlated with the tumour infiltration of the serosa. The Ia.7 determinants were mainly expressed on MO which also expressed the receptor for the Fc part of immunoglobulin. Immunostaining in cellular infiltrates surrounding the tumour revealed that Ia.7+ macrophages (MO) were more numerous than in normal gastric mucosa and severe chronic gastritis and were mostly present in close proximity to tumour cells, while DR+MO were mainly localized within the stromal tissue of the tumour and their number was not increased in cancer infiltrates. These observations indicate that the Ia.7+ subpopulation of MO may be involved in the anti-tumour response of the host.

Epitopes↗
PubMed1988

Immunoregulation of lymphoproliferation in vitro by monocytes and their subpopulations. I. The induction of suppressor T cells in long term cultures and the role of MHC class II determinants and preliminary characteristics.

Peripheral blood monocyte (MO) subpopulations isolated on a basis of functional expression or a lack of Fc receptor (FcR+ and FcR- MO) were used to study the regulation of the antigen (PPD)-driven lymphoproliferation. Long-term cultures of T lymphocytes with FcR+ MO, but not FcR- MO, led to the induction of T suppressor (Ts) cells that inhibited antigen-driven lymphoproliferation in the test cultures. These Ts cells were resistant to mitomycin C, belonged to afferent-acting category of Ts cells, expressed CD8 and HLA-DR determinants, and showed no antigenic specificity nor genetic restriction in action. The expression of MHC class II molecules (HLA-DR and HLA-DP but not HLA-DQ determinants) on MO which were used for antigen presentation was critical for Ts cell induction. It was concluded that specialized MO subpopulations may regulate the lymphoproliferation by inducing Ts cells (or Ts cell circuit) that in turn inhibit antigen-driven immune response.

Cells, Cultured↗
PubMed1988

Juvenile rheumatoid arthritis. Monocyte dysfunction in selected patients.

The in vitro parameters of cell-mediated immunity were studied in 20 children with an established diagnosis of Juvenile rheumatoid arthritis (JRA) (age range 4-15 years) and 23 age- and sex-matched healthy children. (No attempt was made to correlate the observed changes with clinical course or treatment). We are not certain, at this time, of clinical relevancy or the generalizability of the findings. The normal level of T-lymphocytes (CD3+) and normal proportions of CD4+ and CD8+ lymphocytes were seen in children with JRA. The in vitro response of lymphocytes to T-cell mitogen phytohemagglutinin (PHA) also was normal. The suppressor activity of JRA monocytes was essentially the same as controls. In contrast, monocytes from patients with JRA showed the following: decreased expression of receptors for Fc part of IgG immunoglobulin (FcR), diminished nitro blue tetrazolium (NBT) reduction activity, and depressed expression of Ia.7 major histocompatibility complex (MHC) class II determinants. This indicates that certain monocyte functions in selected patients with a variety of manifestations of JRA are depressed.

Adolescent↗