Anti-atherogenic effect of statins: role of nitric oxide, peroxynitrite and haem oxygenase-1.

BACKGROUND AND PURPOSE: The pleiotropic effects of HMG-CoA inhibitors (statins), which include anti-inflammation, antioxidation and immunomodulation, are not yet fully understood. The present study was designed to elucidate the role of nitric oxide (NO), peroxynitrite (ONOO(-)) and haem oxygenase-1 (HO-1) in the anti-atherogenic effect of statins. EXPERIMENTAL APPROACH: Normal and atherosclerotic New Zealand rabbits were treated with atorvastatin or simvastatin in the presence or absence of inhibitors and promoters of endothelial nitric oxide synthase (eNOS) and HO-1. NO and ONOO(-) released from isolated aortae by calcium ionophore were measured with nanosensors placed 6 +/- 2 nm from aortic endothelium. Expression of eNOS and HO-1 protein, HO activity, plasma malondialdehyde (MDA) and vessel wall thickness were also measured. KEY RESULTS: Hypercholesterolaemia decreased eNOS expression by 31 +/- 3%, decreased NO (230 +/- 16 vs. 433 +/- 17 nmol x L(-1) control) and increased cytotoxic ONOO(-) (299 +/- 15 vs. 187 +/- 11 nmol x L(-1) control). The concentration ratio of [NO]/[ONOO(-)] decreased from 2.3 +/- 0.1 (normal) to 0.7 +/- 0.1 indicating an increase of nitroxidative stress in atherosclerotic endothelium. Expression of HO-1 protein increased by 20 +/- 8% in atherosclerosis and further increased (about 30%) after treatment with statins. Statins partially restored the [NO]/[ONOO(-)] balance (1.5 +/- 0.1 for atorvastatin and 1.4 +/- 0.1 simvastatin), decreased MDA and wall thickening. Promoters of eNOS and HO-1 (L-arginine and haemin) ameliorated the [NO]/[ONOO(-)] ratio while their inhibitors (L-NAME or tin-protoporphyrin) showed no improvement in these ratio. CONCLUSIONS AND IMPLICATIONS: Atherosclerosis induced an endothelial [NO]/[ONOO(-)] balance indicative of endothelial dysfunction. Statins showed anti-atherosclerotic effects mediated by HO-1/eNOS, restoring the [NO]/[ONOO(-)] imbalance and reducing lipid peroxidation.

In vivo endothelial interaction between ACE and COX inhibitors.

Here we studied the mechanism of thrombolytic response (THR) induced by angiotensin converting enzyme (ACE-I) in vivo in anaesthetised Wistar rats with extracorporeal circulation. Intravenous injections of ACE-Is, i.e. perindopril or quinapril at non-hypotensive doses of 3-30 microg kg(-1) produced a dose-dependent thrombolysis that was associated with a parallel rise in arterial blood levels of 6-keto-PGF(1 alpha), but not those of TXB(2) or PGE(2). L-NAME at a dose of 5 mg kg(-1) affected significantly neither ACE-I-induced thrombolysis nor prostacyclinemia; however, the pre-treatment with icatibant (0.1-0.5 mg kg(-1)) abolished both effects. The selective COX-1 inhibitor, SC 560 (100-300 microg kg(-1) i.v.), or a would be selective COX-3 inhibitor--paracetamol (acetaminophen, 1-3 mg kg(-1)), both agents induced a transient thrombolysis and slightly potentiated thrombolysis by ACE-Is. In contrast, selective COX-2 inhibitors (rofecoxib>>celecoxib>nimesulide>NS 398) were thrombogenic, and abolished THR and rise in 6-keto-PGF(1 alpha) induced by ACE-Is. Summing up, in our in vivo bioassay system ACE-Is such as quinapril, perindopril or captopril at non-hypotensive doses evoke THR that is mediated by endogenous bradykinin and prostacyclin derived from endothelial COX-2.

Endothelial action of thienopyridines and thienopyrimidinones in the isolated guinea pig heart.

Antiplatelet thienopyridines (ticlopidine, clopidogrel) and their thienopyrimidinone congeners, induce prostacyclin-dependent thrombolysis in vivo. Here we tested whether thienopyridines (ticlopidine, clopidogrel, and its enantiomer without antiplatelet properties) and structurally related thienopyrimidinones release NO from coronary endothelium in the isolated guinea pig heart, perfused according to Langendorff technique. The involvement of endothelium-derived NO in coronary vasodilation induced by these agents was assessed by effect of L-N(G)-nitro-arginine methyl ester (L-NAME). In addition, effect of thienopyridines or thienopyrimidinones on nitrite accumulation in cultured endothelium was assayed. Tienopyridines (10-100 micromol L(-1)) and thienopyrimidinones (10-30 micromol L(-1)) produced concentration-dependent increase in coronary flow comparable to that induced by acetylcholine (0.1 micromol L(-1)) or bradykinin (3 nmol L(-1)) which was inhibited by L-NAME (by 50-70%) but not by indomethacin. Furthermore, thienopyridines and thienopyrimidinones caused NO release from cultured endothelial cells. In conclusion, both thienopyridines independently from their antiplatelet action and their thienopyrimidinone congeners that are devoid of antiplatelet action stimulate coronary endothelium to release NO. Endothelial action of these compounds merits further investigation.

Clonidine-induced coronary vasodilatation in isolated guinea pig heart is not mediated by endothelial alpha2 adrenoceptors.

Functional role of endothelial alpha(2)-adrenoceptor in coronary circulation remains unclear. Clonidine, an agonist of alpha(2)-adrenoceptors, was reported to induce coronary vasodilatation via stimulation of endothelial alpha(2)-adrenoceptors or coronary vasoconstriction involving vascular smooth muscle alpha(2)-adrenoceptors. Moreover, H(2) receptor-dependent responses to clonidine were described. Here, we reassess the contribution of endothelial alpha(2)-adrenoceptor and H(2) receptors to coronary flow and contractility responses induced by clonidine in the isolated guinea pig heart. We found that clonidine (10(-9) - 10(-6) M) produced concentration-dependent coronary vasoconstriction without a significant change in contractility. This response was inhibited by the alpha(1)/alpha(2)-adrenoceptor antagonist - phentolamine (10(-5) M) and the selective alpha(2)-adrenoceptor antagonist yohimbine (10(-6) M), but it was not changed by the selective alpha(1)-adrenoceptor antagonist prazosin (10(-6) M). In the presence of nitric oxide synthase inhibitor, L-NAME (10(-4) M) the clonidine-induced vasoconstriction was potentiated. Clonidine at high concentrations of 10(-5) - 3 x 10(-5) M produced coronary vasodilatation, and an increase in myocardial contractility. These responses were abolished by a selective H(2)-receptor antagonist, ranitidine (10(-5) M), but not by phentolamine (10(-5) M). We conclude that in the isolated guinea pig heart, clonidine-induced vasoconstriction is mediated by activation of smooth muscle alpha(2)-adrenoceptors whereas clonidine-induced coronary vasodilatation is mediated by activation of vascular H(2) histaminergic receptors. Accordingly, endothelial alpha(2)-adrenoceptors does not seem to play a major role in coronary flow response induced by clonidine.

Eosinophil-epithelial cell interaction augments cysteinyl leukotrienes synthesis.

Eosinophils accumulation in the airways and sustained eosinophil-derived cysteinyl leukotrienes production represent key elements of the inflammatory response seen in asthma. However, it is not known whether activated epithelial cells influence cysteinyl leukotrienes production by eosinophils from healthy valunteers. The aim of the present study was therefore to analyse the effects of interactions between non-atopic eosinophils and epithelial cells on cysteinyl leukotrienes production in vitro. We measured cysteinyl leukotrienes released by phorbol 12-myristate 13-acetate (PMA) -activated human eosinophils or epithelial cells (human bronchial epithelial cell line -BEAS-2B) cultured alone or together. While activated BEAS-2B cells barely formed leukotrienes (1.39 pg/ml +/- 0.2) (n=32), activated eosinophils produced considerable amount of them (62.25 pg/ml +/- 10.29) (n=32). Interestingly, when activated eosinophils and epithelial cells were co-incubated, production of cysteinyl leukotrienes increased substantially (571.1 pg/ml +/- 80.9) (n=32). Thus, eosinophil-epithelial cell interactions, when occur, are associated with increased biosythesis of cysteinyl leukotrienes.

Mesoionic oxatriazoles (MOTA): NO-donating characteristics and pharmacology.

Biological role of nitric oxide (NO), functioning of isoforms of NO synthetases (NOS) and pharmacology of principle NO-donors were reviewed. NO donating characteristics and pharmacology of 23 mesoionic oxatriazoles (MOTA) were compared with those of 5-morpholinosydnonimine (SIN-1), S-nitroso-N-acetylpenicillamine (SNAP), sodium nitroprusside (NaNP) and glyceryl trinitrate (GTN). It is concluded that in vitro NO donating profile of MOTA hardly can be used as a predicting measure for their pharmacological activities either in vitro or in vivo. If anything, fast NO releasers seem to be stronger vasorelaxants than MOTA with slow NO releasing properties. Still, among representatives of this last category of MOTA one may find efficient antithrombotic and thrombolytic agents. For instance, MOTA 5-oxides were more potent thrombolytics than SIN-1, SNAP or NaNP. Also MOTA with potent anti-platelet action in vitro seem to be potent relaxants of tracheal strips. In summary, by manipulating the chemical structures of MOTA one may obtain relative selectivity towards vasorelaxant, anti-platelet, thrombolytic or tracheorelaxant properties. Thus different categories of MOTA might be designed with a hope of achieving hypotensive, antithrombotic, thrombolytic or anti-asthmatic drugs.

Flavonoids and nitric oxide synthase.

Induction of NOS-2 in macrophages and smooth muscles within vascular wall with concomittant suppression of endothelial NOS-3 activity is considered to be a hallmark of vascular inflammation that triggers atherogenesis. Accordingly, drugs designed to reverse these changes should not only support vaning function of NOS-3 but also suppress proinflammatory NO production by NOS-2. It means that using selective inhibitors of induction of NOS-2 (they spare ex definitione constitutive activity of NOS-3) is a more rational approach than using "selective" inhibitors of activity of previously induced NOS-2. First of all, those drugs are never sufficiently selective. In our work we tried to identify inhibitors of NOS-2 induction within the group of flavonoids, known stimulators of NOS-3 with putative antiatherogenic effects. Representatives of four main groups of flavonoids: flavonols (kaempferol, quercetin, rutin), flavones (apigenin, primuletin), flavanols (catechine) and flavanones (hesperetin, hesperidin, naringenin) were tried on NOS-2 induction and activity in the in vitro model of LPS-treated macrophages (cell line J774.2). While none of these compounds inhibited activity of NOS-2, all with unexpectedly scattered potencies inhibited induction of NOS-2 protein in LPS-treated J774.2 cells, as evidenced by Western blotting technique. Subsequently, RT-PCR and Northern blotting methods revealed that so far the most potent compounds, kaempferol and apigenin, at micromolar concentrations did inhibit NOS-2 induction at the level of NOS-2 gene transcription. We conclude that some of flavonoids are potent inhibitors of NOS-2 induction. At the same time they may increase endothelial NOS-3 activity. Could these flavonoids become natural parents of future drugs, which will be used for reversal of inflammatory component of atherothrombosis?

No-dependent vasodilation induced by nebivolol in coronary circulation is not mediated by beta-adrenoceptors or by 5 HT1A-receptors.

Nebivolol is a unique beta1-adrenoceptor antagonist which possesses peripheral vasodilator properties dependent on endothelial NO. Nebivolol-induced release of NO was attributed to its L stereoisomer and to its ability to stimulate endothelial beta2, beta3 adrenoceptors or 5-HT1A receptors. Here, in the isolated guinea pig heart we analysed coronary vasodilator potency of L- and D-nebivolol and a possible role of beta2, beta3 adrenoceptors and 5-HT1A receptors in nebivolol-induced vasodilation. Surprisingly, we found that not only L-nebivolol (3-30 x 10(-6) M) but also D-nebivolol (3-30 x 10(-6) M) induced coronary vasodilation, and both responses were inhibited by L-NAME (10(-4) M). In contrast with the stereoisomers of nebivolol, atenolol at the equimolar concentrations did induce slight vasoconstriction. The nonselective beta1/beta2-adrenoceptor antagonist--nadolol (10(-5) M), the selective beta3-adrenoceptor antagonist--L 748337 (10(-6) M) and the 5 HT1A receptor antagonist--NAN 190 (5 x 10(-6) M), none of them inhibited coronary vasodilation induced by D- and L-nebivolol. In summary, in the isolated guinea pig heart both D- and L-nebivolol act as coronary vasodilators. Coronary vasodilation induced by stereoisomers of nebivolol is mediated by endothelium-derived NO and does not depend on beta2, beta3 adrenoceptors or 5 HT1A receptors.

Synthesis and thrombolytic activity of new thienopyrimidinone derivatives.

It has been observed that ticlopidine and clopidogrel show, apart from their delayed antiplatelet properties, an immediate and transient thrombolytic action related to the ability of these thienopyridines to stimulate the secretory function of vascular endothelium. With the objective to construct new molecules with identical thrombolytic potency but at a higher level, we carried out different structural modifications in the thienopyridine chemical molecule to conclude that the presence of a second N atom in the pyridine cycle (yielding pyrimidine moiety) and the presence of an additional cycle fused to the thienyl ring would lead to enhanced thrombolytic effects. Here we report the six-step synthesis of a series of new benzothienopyrimidinone derivatives characterized by this searched for potent thrombolytic activity. The pharmacological assay used anaesthetised Wistar rats with extracorporal circulation in which arterial blood superfused thrombi adhering to a strip of collagen. Weight of thrombi was continuously monitored. Six compounds of the series were much more potent thrombolytic agents than their thienopyridine references: the effective thrombolytic dose that produced 30% of maximum thrombolysis (ED30) was at a range of 8 to 170 microg kg(-1) as compared with ED30 values of 16000 to 20000 microg kg(-1) for clopidogrel and ticlopidine respectively. Especially with the most active compound, this difference in the threshold thrombolytic dose, giving an intensity of action higher by three orders of magnitude, was accompanied by a lengthening of the response. Apart from that these compounds have shown to be synthetic thrombolytics, they certainly deserve further studying.

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.

Paradoxical augmentation of bradykinin-induced vasodilatation by xanthine/xanthine oxidase-derived free radicals in isolated guinea pig heart.

Increased generation of reactive oxygen species contribute to endothelial dysfunction in atherosclerosis, hypertension and heart failure. Recently, it was suggested that bursts of superoxide anions may inactivate endothelial surface-bound enzymes such as angiotensin converting enzyme (ACE). Here, we tested effects of xanthine/xanthine oxidase-derived superoxide anions on vascular responses and ACE activity in the isolated guinea pig heart. We analysed effects of intracoronary infusion of low concentration of xanthine oxidase (10 mU/ml) in the presence of xanthine (0,5 mM) (X/XO) on bradykinin, other endothelium-dependent and independent vasodilators (acetylcholine, ADP, SNAP), as well as vasoconstrictor responses to angiotensin I and angiotensin II. Surprisingly, X/XO significantly augmented coronary response to bradykinin without an effect on responses to ADP, acetylcholine, SNAP, angiotensin I and angiotensin II. In contrast, inhibition of ACE by perindoprilate (100 nM) resulted in augmentation of bradykinin-induced vasodilatation as well as diminution of angiotensin I-evoked vasoconstriction without an influence on other responses. In summary, in the isolated guinea pig heart, X/XO-derived free radicals selectively augmented coronary vasodilator response to bradykinin, which cannot be explained by X/XO-induced derangement of ACE. The mechanism of this paradoxical phenomenon, which might represent a defensive response of the coronary circulation to oxidative stress requires further investigations.

Significance of endothelial prostacyclin and nitric oxide in peripheral and pulmonary circulation.

BACKGROUND: Vasoprotective function of endothelial cells is associated, among others, with biosynthesis and release of nitric oxide (NO), prostacyclin (PGI2), prostaglandin E2 (PGE2), carbon monoxide (CO) and plasminogen activator (t-PA). These endothelial mediators calm down activated platelets and leukocytes, prevent the occurrence of parietal thrombotic events, promote thrombolysis, maintain tissue perfusion and protect vascular wall against acute damage and against chronic remodeling. Endothelial dysfunction in patients suffering from atherosclerosis or diabetes type 2 is associated not only with suppression in release of the above mediators but also with deleterious discharge of prostaglandin endoperoxides (PGH2, PGG2), superoxide anion (O2-, peroxynitrite (ONOO-), and plasminogen activator inhibitor (PAI-1). We looked for mechanisms of protective endothelial function, with a special respect to the differences between peripheral and pulmonary circulation. METHODS: Cultured endothelial cells of bovine aorta (BAEC) were used to study physiological and pharmacological mechanisms of increasing free cytoplasmic calcium [Ca2+]i. A porphyrinic sensor quantified the release of NO from BAEC. In cultured human umbilical vein endothelial cells (HUVEC) we looked for induction by bradykinin (Bk) of mRNAs for a number of enzymes. In blood perfused rat lungs we studied protective role of NO against injury inferred by lipopolysaccharide on pulmonary microcirculation that was accomplished by thromboxane A2 (TXA2), platelet activating factor (PAF), cysteinyl-leukotrienes (cyst-LTs) and the complement system. In vivo we analyzed the influence of Bk, perindopril and quinapril ('tissue type' angiotensin converting enzyme inhibitors, ACE-Is) on endothelial function in entire circulation of anaesthetized rats using a thrombolytic bioassay and EIA for 6-keto-PGF1 alpha and t-PA antigen. RESULTS: In BAEC Bk via kinin B2 receptors raised in a concentration-dependent manner (1 pM-10 nM) free cytoplasmic calcium ions [Ca2+]i, that triggered the release of NO from BAEC. Calcium ionophore (A23187, 1-100 nM) as well as receptor agonists such as adenosine diphosphate (ADP, 10 nM-1 microM), adrenaline (Adr, 1-10 microM) or acetylcholine (Ach, 10-100 microM) produced a similar rise in endothelial [Ca2+]i as did Bk at a nanomolar concentration. 'Tissue type' ACE-Is, e.g. quinapril or perindopril acted through accumulation of endogenous Bk. However, the potency of ACE-I to change endothelial function is by several orders of magnitude lower than that for exogenous Bk. In vivo the major difference between thrombolytic actions by quinapril or perindopril on one hand, and by exogenous Bk on the other was longevity of thrombolysis by ACE I and a distinct hypotensive action of exogenous Bk. Still, the long-lasting isolated thrombolytic effect of ACE I was mediated entirely by endogenous BK as evidenced by the preventive action of icatibant, a kinin B2 receptor antagonist. Moreover, in vivo the immediate thrombolysis by ACE-I was mediated by PGI2 rather than by NO or t-PA, as shown by pharmacological analysis, and by direct blood assays of 6-keto-PGF1 alpha and t-PA antigen. Bradykinin as a mediator of pleiotropic endothelial action of several cardiovascular drugs (e.g. ACE-I) may complete its mission not only through B2 receptor and [Ca2+]i--mediated release of PGI2 or NO. Here, we describe a new route of the Bk action. Bk mediated induction of the [Ca2+]i-independent, so called 'inducible', endothelial isoenzymes required for generation of CO, PGI2 and PGE2. After 4 hours of incubation of HUVEC with Bk (10 nM) it induced mRNAs for haemooxygenase 1 (HO-1), cyclooxygenase 2 (COX-2), prostaglandin E synthase (PGE-S) whereas mRNA for nitric oxide synthase 2 (NOS-2) was weakly affected. We proved also that unlike in peripheral circulation, in pulmonary circulation only NO but not PGI2 would play a protective role. In the blood-perfused lung, endotoxaemia liberates lipids, such as TXA2, PAF and cyst-LTs. These toxic lipids along with the activated complement mediate pulmonary damage. Pulmonary endothelial nitric oxide is the only local protector against lung injury evoked by the phagocytised bacterial lipopolysaccharide. SUMMARY: Summing up, in peripheral circulation endogenous Bk is the most efficient activator of protective endothelial function. For instance, thrombolytic action of 'tissue type' ACE-I depends on the Bk-released PGI2. Acting as an agonist of endothelial B2 kinin receptors Bk rises [Ca2+]i with a subsequent activation of constitutive COX 1 and NOS-3. This is followed by an immediate release of PGI2 and NO. Moreover, acting as 'microcytokine' Bk induces mRNAs for HO-1, COX-2 and PGE S, the isoenzymes responsible for a delayed endothelial biosynthesis of CO, PGI2 and PGE2. (ABSTRACT TRUNCATED)

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.

Residual pulmonary neutrophils are involved in vaso- and bronchoconstrictor responses to platelet activating factor in the isolated buffer-perfused rat lung.

The aim of the study was to quantify residual neutrophils sequestrated in the isolated rat lung, perfused at a constant flow with Krebs-Hanseleit solution, and to analyze their possible influence on functional responses of the isolated lung. For that purpose we assessed neutrophil content in the isolated lung as well as in the effluent from the lung using an assay of myeloperoxidase (MPO) activity. We showed that a considerable pool of neutrophils remained in the isolated lung even after a 20-min period of washout with buffer solution. Moreover, these residual neutrophils were responsible for platelet activating factor (PAF)-induced vaso- and bronchoconstriction but not for oedema formation. We conclude that when studying responses to pharmacological agents in isolated buffer-perfused lung, the presence of sequestrated neutrophils should be taken into account.

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.

Antiplatelet action of losartan involves TXA2 receptor antagonism but not TXA2 synthase inhibition.

Various AT1 receptor antagonists including losartan are known to inhibit human platelet activation by antagonising TXA2/PGH2 receptors (TP receptors). Presently, we check a hypothesis that losartan, an imidazole derivative in contrast with valsartan, a non-imidazole compound, may inhibit human platelet activation also through inhibition of TXA2 synthesis. Inhibitory action of losartan (2-n butyl-4-chloro-5-hydroxymethyl-1-beta(2'-(1H-tetrazol-5yl)biphenyl-4-yl)methyl] imidazole), its active metabolite EXP 3174 (2-n-butyl-4-chloro-1-beta (2-(1H-tetrazol-5-yl) biphenyl-4-yl) methyl]imidazole-5-carboxylic acid) and valsartan ((S)-N-valeryl-N-(beta2'-(1H-tetrazol-5-yl)biphenyl-4-yl]methyl]valine), on collagen-induced platelet aggregation and TXA2 generation was compared to effects achieved by each compound on U46619-induced aggregation in aspirinized platelets. Losartan and aspirin inhibited collagen-induced platelet aggregation with approximately the same potency, whereas EXP 3174 and valsartan showed much weaker antiplatelet effects. Interestingly, losartan, EXP 3174 and valsartan displayed similar potencies as inhibitors of U46619-induced aggregation in asprinized platelets as in collagen-induced aggregation in non-aspirinized platelets. None of the above three AT1 antagonists, up to a concentration of 300 microM, did influence collagen-induced TXA2 synthesis in human platelets. In conclusion, antiplatelet effects of AT1 antagonists, irrespective of the presence or absence of non-condensed imidazole in their chemical structure, involve antagonism of TP receptors but not inhibition of TXA2 synthesis in platelets.

[Historical outline of the Medical Department Museum of the Jagiellonian University]. / Zarys dziejów Muzeum Wydzialu Lekarskiego UJ.

The article presents the history of the Museum of History of the Medical Department of the Jagiellonian University in Cracow, which was founded in 1900. Initiated and created by Walery Jaworski, professor of Internal Diseases, the museum has gathered a rich and varied collection of artifacts. These comprise such exhibits as manuscripts, old texts, medical diplomas as well as photographs of doctors, medical and laboratory equipment and a collection of medals and mementoes of professors from Cracow University.

Role of thromboxane A2 and platelet activating factor in early haemodynamic response to lipopolysaccharide in rats.

The mechanism of early pulmonary and systemic haemodynamic response to intravenous infusion of LPS from Escherichia coli was investigated in anesthetised Wistar rats. 10 mg of LPS given at a rate of 4 mg/kg/min but not at a rate of 1 mg/kg/min induced an increase in pulmonary arterial pressure (PAP) and a fall in systemic arterial pressure (SAP). Pretreatment with a PAF receptor antagonist; WEB 2170 (5 and 25 mg/kg) inhibited both PAP and SAP responses to LPS (4 mg/kg/min) while an inhibitor of thromboxane synthesis; Camonagrel (10 and 20 mg/kg) abolished PAP response without a major effect on SAP response to LPS. In conclusion, both PAF and TXA2 mediate LPS induced rise in pulmonary arterial pressure while LPS-induced fall in systemic arterial pressure is mediated by PAF.

Transcellular biosynthesis of eicosanoids.

Transcellular biosynthesis of prostacyclin (platelets/endothelial cells), cysteinyl-leukotrienes (granulocytes/endothelial cells) or lipoxins (granulocytes/renal cells) constitute a new approach to the therapy of thrombosis, ischaemic heart disease and glomerulonephritis.