Plasmin system regulation in an ovalbumin-induced rat model of asthma.

BACKGROUND: So far studies showing the role of the plasmin system in airway remodelling have been conducted using in vitro models. The aim of the present study was to determine plasmin system regulation in an in vivo rat model of asthma. METHODS: Asthma in Wistar rats was induced by ovalbumin (OVA) sensitization followed by an OVA challenge (OVA/OVA, n = 6). Control groups were saline-sensitized challenged with OVA (VEH/OVA, n = 6) and OVA-sensitized challenged with saline (OVA/VEH, n = 6). Plasmin system components were determined in the plasma by ELISA. Plasminogen activator inhibitor-1 (PAI-1) was localized by an immunohistochemical reaction. RESULTS: Sensitization and challenge with OVA caused thickening of the airway wall, hypertrophy of smooth muscle cells, infiltration of inflammatory cells, subepithelial fibrosis, epithelial and endothelial lesions. Serum total IgE was significantly higher in OVA-sensitized rats as compared to VEH-sensitized control groups. Tissue plasminogen activator activity was significantly decreased in asthmatic animals (4.48 +/- 0.4 vs. 6.7 +/- 0.3 ng/ml for OVA/OVA and OVA/VEH; p < 0.05), and PAI-1 activity was statistically significantly higher in asthma rats (0.8 +/- 0.05 vs. 0.5 +/- 0.03 ng/ml for OVA/OVA vs. OVA/VEH; p < 0.05). alpha2-Antiplasmin was higher in rats receiving OVA sensitization than in those that were sham sensitized (p < 0.05). Immunohistochemical staining for PAI-1in the lungs of asthmatic animals showed very strong PAI-1 expression in lung inflammatory cells. CONCLUSIONS: We have demonstrated for the first time the existence of PAI-1 in lung inflammatory cells of rats with asthma. This finding was consistent with the superiority of plasmin system inhibition over activation in plasma.

Basal serum tryptase level correlates with severity of hymenoptera sting and age.

BACKGROUND: Increased serum tryptase has been linked to the severity of the reaction after Hymenoptera stings. The aim of the study was to measure basal tryptase levels in patients with Hymenoptera venom allergy and investigate the possible correlation between these levels and the severity of sting reaction. METHODS: One hundred nine patients were included in the study. Sixty-three were wasp venom-allergic and 46 were honey bee venom-allergic. Basal serum tryptase levels were measured by UniCAP. RESULTS: Basal serum tryptase levels were elevated in 12 (11%) of the 109 patients. Levels were 5.14 pg/L (3.62-5.84), 5.3 microg/L (2.94-6.54), 5.18 microg/L (3.71-6.25), and 6.98 microg/L (4.78-12.6), for patients with sting reactions of grade I, II, III and IV (as classified by Mueller), respectively. Basal serum tryptase levels correlated significantly with the sting reaction severity (r = 0.2752; P = .004) and with age (r = 0.2906; P = .002). Sting reaction severity also correlated with age (r = 0.3654; P = .001). CONCLUSIONS: Basal serum tryptase levels were found to be elevated in 11% of venom allergic patients and correlated significantly with both sting reaction severity and age.

The influence of inhaled corticosteroids on exhaled nitric oxide in stable chronic obstructive pulmonary disease.

The aim of the study was to evaluate the exhaled nitric oxide (F(ENO)) in clinically stable chronic obstructive pulmonary disease (COPD), its relationship to the severity of the disease, pulmonary function, smoking status, reversibility of airflow limitation, and ICS therapy. The study was conducted in 47 patients with COPD and 40 healthy controls. Flow/volume spirometry and F(ENO) measurement were performed before and after 2 months of ICS therapy. F(ENO) were significantly elevated in current smokers and ex-smoking COPD patients. In both groups of COPD patients inhaled corticosteroids (ICS) therapy caused a significant decrease in F(ENO) without significant changes in FEV1. A positive correlation between initial F(ENO) and postbronchodilator FEV1 (% predicted) was observed in the group of ex-smoking COPD patients, but not in the currently smoking COPD group. In both groups of COPD patients, the initial level of F(ENO) correlated with the reversibility of airway obstruction, the increase in postbronchodilator FEV1 and the decrease in F(ENO) following ICS treatment. F(ENO) increases in patients with stable COPD. ICS therapy decreased elevated F(ENO) levels in these patients without statistically significant changes in lung function. The selection of COPD patients with increased F(ENO) level and partial reversibility of airway obstruction should be helpful in terms of proposed ICS treatment.

Angiotensin-(1-7): an active member of the renin-angiotensin system.

Angiotensin-(1-7) [Ang-(1-7)] is an active member of renin-angiotensin system (RAS). It counterbalances vasoconstriction, mitogenic, arrhythmogenic and prothrombotic actions of Ang II. Inducing natiuresis and diuresis opposes also the water and sodium retention produced by Ang II. Till now the specific receptor side for Ang-(1-7) has been not cloned, but the current data strongly suggest that an interaction (cross-talk) between angiotensin receptors may play a role in the effects of Ang-(1-7).

Studies on the antithrombotic action of AT1 receptor antagonists.

BACKGROUND: In our previous experiments we showed that the prototype member of the AT1 receptor antagonists (AT1-As) family, losartan, prevented the development of arterial and venous thrombosis in rats. Recent studies have demonstrated that apart from blocking AT1 receptor, losartan is also a competitive antagonist to thromboxane A2/prostaglandin H2 receptor (TP receptor). Thus, we decided to assess if this feature could contribute to the antithrombotic action of losartan. MATERIAL AND METHODS: We compared the influence losartan, its active metabolite EXP3174 and valsartan on rat platelet adhesion to fibrillar collagen and platelet aggregation in response to thromboxane A2 analogue, U46619. We also assessed the efficacy of these drugs in platelet-dependent pulmonary thrombosis in mice as well as preventive and therapeutic models of venous thrombosis in rats. RESULTS: All the three compounds, given in a single dose, inhibited rat platelet adhesion to fibrillar collagen and platelet aggregation induced with U46619 in vitro and ex vivo, with the action of losartan being much more pronounced than that of EXP3174 or valsartan. Losartan also more effectively protected mice from death in response to the intravenous injection of collagen / epinephrine and it was the only compound which reduced mice mortality after the intravenous injection of U46619. In contrast, all the three AT1 receptor antagonists exerted a similar thrombolytic action and comparably decreased the thrombus weight in the therapeutic and preventive model of venous thrombosis, although in the latter case a high dose of losartan was slightly more effective than a corresponding dose of EXP3174 and valsartan. CONCLUSIONS: Since losartan is endowed with a relatively low affinity towards the AT1 receptor, we conclude that its superiority over EXP 3174 and valsartan in inhibiting thrombocyte function and platelet-dependent thrombosis could result from its stronger action on the TP receptor. This feature seems to be less important in the thrombolytic effect of AT1-As and in the inhibition of the venous thrombosis development, in which platelets play only a minor role.

Thiol repletion prevents venous thrombosis in rats by nitric oxide/prostacyclin-dependent mechanism: relation to the antithrombotic action of captopril.

Clinical and experimental data have recently accumulated for antithrombotic action of angiotensin-converting enzyme inhibitors (ACE-1s). We have shown previously that captopril (which contains a thiol group in the moiety) exerts more pronounced antithrombotic activity than does an equipotent dose of enalapril (the drug devoid of the thiol group). To clarify the relative importance of the presence of the thiol group in the molecule versus angiotensin-converting enzyme (ACE) inhibitory properties in the antithrombotic action of captopril, rats were treated with captopril (5 mg/kg twice daily; CAP), epicaptopril (stereoisomer of captopril devoid of ACE-inhibitory properties; 5 mg/kg twice daily; EPI), N-acetylcysteine (3.75 mg/kg twice daily; ACC), enalapril (3 mg/kg once daily; ENA), or distilled water (VEH) for 10 days, per os. After ligation of the vena cava, the incidence of the venous thrombosis and/or the thrombus weight decreased significantly in all but the ENA-treated groups when compared with control rats. The effect of CAP, EPI, and ACC was accompanied by a marked reduction of euglobulin clot lysis time and, with the exception of ACC, by an increase in prothrombin time in the blood collected from the site of the thrombus formation. Antithrombotic activity of EPI was completely abolished by nitric oxide synthase inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) or indomethacin, with the parallel reversal of fibrinolytic and coagulation parameters toward normal. Activated partial thromboplastin time, mean blood pressure, and bleeding time were not altered by either of the administered drugs. Thus, we demonstrated that thiol compounds exert antithrombotic activity by increasing fibrinolysis and/or suppression of the extrinsic pathway of the coagulation cascade in a nitric oxide/prostacyclin-dependent manner.

Angiotensin-(1-7). One step forward?

Angiotensin-(1-7) [Ang-(1-7)] is a paracrine hormone of the renin-angiotensin system (RAS). It counterbalances the negative actions of angiotensin II (Ang II) acting in the cardiovascular system, kidneys and central nervous system, and is responsible for blood pressure regulation and antiproliferative effects. Current data strongly suggest the existence of a specific receptor for this peptide. The concentration of Ang-(1-7) increases significantly during the administration of RAS blockers. One may suggest the involvement of this peptide in a beneficial effect of these drugs.

Hypotensive effect of angiotensin II after AT1-receptor blockade with losartan.

Recent data suggest that hypotensive effect of losartan may not be attributed solely to AT1-receptor blockade, but also to excessive AT2 or other receptors stimulation by elevated angiotensin II and its derivative peptides. Therefore in the present study we examined the effect of angiotensin II on mean blood pressure after AT -receptor blockade with losartan. Male Wistar rats were anaesthetised and received injection of either losartan (30 mg/kg, 1 ml/kg, i.v.) or saline (the same volume and route) followed by bolus injection of angiotensin II (100, 300 or 1,000 ng/kg; 1 ml/kg, i.v.) or 1-hour infusion of angiotensin II (200 ng/kg/min; 2.5 ml/kg/h, i.v.). Control animals received saline instead. Angiotensin II, given either as the injection or the infusion, caused an evident increase in mean blood pressure (p ranged from 0.05 to 0.001 depending on the experimental group). Losartan caused a rapid drop in mean blood pressure and blunted the hypertensive effect of angiotensin II (p < 0.01). Moreover, in the losartan-pretreated animals the hypotensive phase was enhanced by the infusion, but not single injection of angiotensin II, which was most evident from the 30 th minute of observation (p < 0.05 vs control). In conclusion, hypotensive effect of losartan may be amplified by simultaneous increase in angiotensin II level, the situation observed during chronic AT1-receptor blockade.

Losartan inhibits the adhesion of rat platelets to fibrillar collagen--a potential role of nitric oxide and prostanoids.

The aim of the study was to evaluate the effect of losartan on rat platelet adhesion to fibrillar collagen. Washed platelets were counted before and after 15 minutes incubation with collagen (50 microg/ml) and the percentage of adhering platelets was calculated as the index of their adhesion. When the platelets were incubated with collagen 40.8 +/- 0.3% of the platelets adhered. Losartan produced a dose dependent decrease in a number of adhering platelets both when the drug was administered to the animals ex vivo at doses of 3, 10 and 30 mg/kg (p < 0.01-0.001) or was added to the preparation of washed platelets in vitro in concentrations of 10(-8)-10(-5) M (p < 0.01-0.001). In the next step of the study we assessed the influence of L-NAME (10 mg/kg ex vivo, 30 microM in vitro) and indomethacin (2.5 mg/kg ex vivo, 30 microM in vitro) on the antiadhesive effect of losartan (10 mg/kg ex vivo, 10(-6) M in vitro). Blockade of nitric oxide synthase with L-NAME partially reversed the antiadhesive effect of losartan both ex vivo and in vitro. Indomethacin diminished the inhibitory effect of losartan on platelet adhesion when administered ex vivo, but it failed to modify this parameter when added to the suspension of platelets in vitro. In conclusion, losartan reduces platelet adhesion to fibrillar collagen in a dose-dependent manner. The observed action of losartan seems to be mediated mainly by endothelium- and platelet-derived nitric oxide.

The involvement of AT(2)-receptor in the antithrombotic effect of losartan in renal hypertensive rats.

In previous studies, we have shown that losartan possesses nitric oxide-dependent antithrombotic properties in various models of hypertension in rats. It was demonstrated that stimulation of AT2-receptors plays an important role in the pharmacological effects of AT1-receptor antagonists. Thus, in this study, we examine the participation of AT2-receptors in the antithrombotic action of losartan in renal hypertensive rats on venous thrombosis induced by a two-hour ligation of the vena cava. Losartan administration(30 mg/kg, p.o.) resulted in a marked decrease in thrombus weight (by 85%, p<0.001). PD123319, an AT2-receptor antagonist (10 mg/kg, i.v.), administered concomitantly with losartan, abolished its antithrombotic effect, whilst it had no influence on thrombus weight when given alone. A significant decrease in systolic blood pressure was observed in animals given losartan. PD123319 administration didnot abolish this action of losartan and did not alter blood pressure when given alone. No changes in prothrombin time, activated partial thromboplastin time, or euglobulin clot lysis time were observed in animals administered losartan and/or PD123319.Similarly, primary haemostatics evaluated by bleeding time and platelet count did not change in any group of rats. In conclusion, we have shown that AT2-receptor stimulation is involved in the antithrombotic action of losartan in renal hypertensive rats.

The antithrombotic effect of angiotensin-(1-7) closely resembles that of losartan.

Angiotensin-(1-7) [Ang-(1-7)] is the bioactive peptide which may be responsible for some of the pharmacological effects of losartan. Our previous study has demonstrated the antithrombotic action of losartan in a model of experimental thrombosis. In the present study, we compared the antithrombotic action of losartan and Ang-(1-7). Acute (10 mg/kg, p.o.) and chronic (10 mg/kg, p.o., three weeks) losartan administration to spontaneously hypertensive rats (SHR) induced a decrease in thrombus weight (1.6 +/- 0.6 mg and 1.2 +/- 0.3 mg respectively vs. control 2.9 +/- 0.8 mg; p<0.05, p<0.05). A similar reduction was observed in two-kidney, one-clip hypertensive rats (2K-IC)receiving acute losartan administration (1.39 +/- 0.29 mg vs. 3.25 +/- 0.62 mg; p<0.01). Infusion of Ang-(1-7) to2K-lC rats also reduced the thrombus weight(1.01 +/- 0.34 mg, 1.23 +/- 0.38 mg and 2.17 +/- 0.36 mg for 1, 10, 100 pmol/kg/min, respectively vs. 3.58 +/- 0.6 mg control; p<0.01, p<0.01, p<0.05). Losartan produced a decrease in systolic blood pressure (BP) in SHR as well as in 2K-1C rats, while Ang-(1-7) infusion had no effect on BP. Acute losartan dosing to 2K-1C rats decreased platelet adhesion to fibrillar collagen(24.9 +/- 1.0% vs. control 31.5 +/- 1.1%, p<0.001). The incubation of platelet samples with Ang-(1-7) (10-6 and 10 5 M) also reduced adhesion to fibrillar collagen(38.4 +/- 0.1% and 33.8 +/- 0.8% respectively vs. control 40.0 +/- 0.6%; p<0.05, p<0.001). There were no apparent changes in prothrombin time, activated partial thromboplastin time and euglobulin clot lysis time in losartan and Ang-(1-7)-treated groups. We conclude that, like losartan, Ang-(1-7) is able to act as an antithrombotic agent.

The role of endothelium in antithrombotic effect of the renin-angiotensin system blockade.

Drugs blocking the renin - angiotensin system, angiotensin converting enzyme inhibitors and AT1 receptor antagonists, among many pharmacological effects may exert an antithrombotic action. The mechanisms, which mediate their antithrombotic activity are associated with enhanced nitric oxide and prostacyclin release or with attenuation of angiotensin II action (Fig. 1, 2). Nevertheless, endothelium plays an important role in this process linking the renin-angiotensin and fibrinolysis / coagulation systems.