Chewing lice from high-altitude and migrating birds in Yunnan, China, with descriptions of two new species of Guimaraesiella.

In total, 366 birds representing 55 species in 24 families and eight orders, were examined for chewing lice (Phthiraptera: Amblycera, Ischnocera) in two high-altitude localities in Yunnan Province, China. In Ailaoshan, almost all of the birds examined were resident passeriforms, of which 36% were parasitized by chewing lice. In Jinshanyakou, most birds were on migration, and included both passerine and non-passerine birds. Of the passerine birds caught in Jinshanyakou, only one bird (0.7%) was parasitized by chewing lice. The prevalence of Myrsidea and Brueelia-complex lice on birds caught in Ailaoshan was higher than in previous reports. Of the chewing lice identifiable to species level, three represent new records for China: Actornithophilus hoplopteri (Mjöberg, 1910), Maculinirmus ljosalfar Gustafsson & Bush, 2017 and Quadraceps sinensis Timmermann, 1954. In total, 17 new host records are included, of which we describe two as new species in the Brueelia-complex: Guimaraesiella (Cicchinella) ailaoshanensis sp. nov. ex Schoeniparus dubius dubius (Hume, 1874) and G. (C.) montisodalis sp. nov. ex Fulvetta manipurensis tonkinensis Delacour & Jabouille, 1930. This published work has been registered in ZooBank, http://zoobank.org/urn:lsid:zoobank.org:pub:9FC3D8EE-2CED-4DBE-A1DB-471B71260D27.

Whole genome microarray expression analysis in blood identifies pathways linked to signs and symptoms of a patient with hypercalprotectinaemia and hyperzincaemia.

A child, 2 years with the 'hypercalprotectinaemia with hyperzincaemia' clinical syndrome, presented with atypical symptoms and signs, notably persistent fever of approximately 38°C, thrombocythaemia of > 700 × 109 /l and a predominance of persistent intestinal symptoms. In an effort to find a cure by identifying the dysregulated pathways we analysed whole-genome mRNA expression by the Affymetrix HG U133 Plus 2·0 array in blood on three occasions 3-5 months apart. Major up-regulation was demonstrated for the Janus kinase/signal transducer and activators of transcription (JAK/STAT) pathway including, in particular, CD177, S100A8, S100A9 and S100A12, accounting for the thrombocytosis; a large number of interleukins, their receptors and activators, accounting for the febrile apathic state; and the high mobility group box 1 (HMBG1) gene, possibly accounting for part of the intestinal symptoms. These results show that gene expression array technology may assist the clinician in the diagnostic work-up of individual patients with suspected syndromal states of unknown origin, and the expression data can guide the selection of optimal treatment directed at the identified target pathways.

Patterns of cryptic host specificity in duck lice based on molecular data.

Documenting patterns of host specificity in parasites relies on the adequate definition of parasite species. In many cases, parasites have simplified morphology, making species delimitation based on traditional morphological characters difficult. Molecular data can help in assessing whether widespread parasites harbour cryptic species and, alternatively, in guiding further taxonomic revision in cases in which there is morphological variation. The duck louse genus Anaticola (Phthiraptera: Philopteridae), based on current taxonomy, contains both host-specific and widespread species. Mitochondrial and nuclear DNA sequences of samples from this genus were used to document patterns of host specificity. The comparison of these patterns with morphological variations in Anaticola revealed a general correspondence between the groups identified by DNA sequences and morphology, respectively. These results suggest that a more thorough taxonomic review of this genus is needed. In general, the groups identified on the basis of molecular data were associated with particular groups of waterfowl (e.g. dabbling ducks, sea ducks, geese) or specific biogeographic regions (e.g. North America, South America, Australia, Eurasia).

A major susceptibility locus for atopic dermatitis maps to chromosome 3q21.

Atopic dermatitis (eczema) is a chronic inflammatory skin disease with onset mainly in early childhood It is commonly the initial clinical manifestation of allergic disease, often preceding the onset of respiratory allergies. Along with asthma and allergic rhinitis, atopic dermatitis is an important manifestation of atopy that is characterized by the formation of allergy antibodies (IgE) to environmental allergens. In the developed countries, the prevalence of atopic dermatitis is approximately 15%, with a steady increase over the past decades. Genetic and environmental factors interact to determine disease susceptibility and expression, and twin studies indicate that the genetic contribution is substantial. To identify susceptibility loci for atopic dermatitis, we ascertained 199 families with at least two affected siblings based on established diagnostic criteria. A genome-wide linkage study revealed highly significant evidence for linkage on chromosome 3q21 (Zall=4.31, P= 8.42 10(-6)). Moreover, this locus provided significant evidence for linkage of allergic sensitization under the assumption of paternal imprinting (hlod=3.71,alpha=44%), further supporting the presence of an atopy gene in this region. Our findings indicate that distinct genetic factors contribute to susceptibility to atopic dermatitis and that the study of this disease opens new avenues to dissect the genetics of atopy.

Soft nanostructuring of YBCO Josephson junctions by phase separation.

We have developed a new method to fabricate biepitaxial YBa2 Cu3 O7-δ (YBCO) Josephson junctions at the nanoscale, allowing junctions widths down to 100 nm and simultaneously avoiding the typical damage in grain boundary interfaces due to conventional patterning procedures. By using the competition between the superconducting YBCO and the insulating Y2 BaCuO5 phases during film growth, we formed nanometer sized grain boundary junctions in the insulating Y2 BaCuO5 matrix as confirmed by high-resolution transmission electron microscopy. Electrical transport measurements give clear indications that we are close to probing the intrinsic properties of the grain boundaries.

Effects of recombinant human prothrombin on thrombin generation in plasma from patients with hemophilia A and B.

BACKGROUND: The present study was carried out to investigate the impact of FII levels, and their increase, on the hemostatic potential in plasma from hemophilia A and B patients with and without inhibitors. METHOD: Recombinant human factor (F) II (rhFII) was added ex vivo to plasma from 68 patients with hemophilia A and B, with or without inhibitors. The hemostatic potential as measured by thrombin generation (calibrated automated thrombogram [CAT]) was focused on the endogenous thrombin potential (ETP) as it has been shown to correlate with the clinical phenotype of bleeding in hemophilia patients and has also been used to guide bypassing therapy in hemophilia patients with inhibitors before elective surgery. The factor eight inhibitor bypassing agent (FEIBA(®) ) was used as a reference to the clinical situation. RESULTS: The study shows that rhFII concentration-dependently increased ETP by a similar magnitude in hemophilia A and B, both with and without inhibitors. Compared with FEIBA, rhFII showed a shallower concentration-response curve. In both types of hemophilia 100 mg L(-1) of rhFII roughly doubled the ETP. A corresponding response was obtained by 0.5 U mL(-1) of FEIBA. CONCLUSION: These data support the theory that FII is one of the major components responsible for the efficacy of FEIBA. The data also indicate that rhFII may be useful, alone or in combination with other coagulation factors, in some of the conditions for which FEIBA is used today, although more data are needed to substantiate this.

Exploration of efficacy and bleeding with combined phosphoinositide 3-kinase ß inhibition and aspirin in man.

BACKGROUND: Based on animal and human data, phosphoinositide 3-kinase (PI3K)ß is a promising antithrombotic target. However, the relation between efficacy and bleeding when combined with current antiplatelet therapies is unclear. OBJECTIVE: To strengthen the PI3Kß target validation using the short-acting inhibitor AZD6482 alone and in different combinations with P2Y12 and cyclooxygenase (COX)-1 inhibition in vitro (human platelets), in vivo (dog), and in healthy subjects. METHODS AND RESULTS: Evaluation of complete target inhibition of PI3Kß (by AZD6482), P2Y12 (by ticagrelor), and COX-1 (by aspirin) alone and in the different combinations vs. concentration responses for a panel of platelet agonists in vitro (adenosine diphosphate, collagen, thrombin receptor activating peptide) indicates that the rank order of antiplatelet efficacy is P2Y12  > PI3Kß > COX-1 as monotherapy and P2Y12 plus PI3Kß > P2Y12 plus COX-1 > PI3Kß plus COX-1 as dual therapy, with little additional effect with triple therapy. Use of a conscious dog model to assess ex vivo antiplatelet effect in parallel with bleeding time prolongation (standard incision in the ear) confirms the wide separation of efficacy vs. bleeding for PI3Kß inhibition and that this separation is reduced when combined with aspirin and more reduced when combined with clopidogrel. In healthy subjects, AZD6482, in combination with aspirin, shows a potential for greater antiplatelet potency but less bleeding potential compared with clopidogrel plus aspirin. CONCLUSIONS: PI3Kß inhibition, in comparison with P2Y12 and COX-1, delivers medium antiplatelet effect but with minimal bleeding. PI3Kß inhibition, in combination with aspirin, in healthy subjects, provides a potential for greater overall antiplatelet effect compared with clopidogrel plus aspirin, but with significantly less bleeding potential.

A randomized, controlled, dose-guiding study of the oral direct thrombin inhibitor ximelagatran compared with standard therapy for the treatment of acute deep vein thrombosis: THRIVE I.

This randomized, controlled, multicentre study evaluated the efficacy and tolerability of the oral direct thrombin inhibitor ximelagatran, compared with a low-molecular-weight heparin (dalteparin) followed by warfarin, in the treatment of deep vein thrombosis (DVT) of the lower extremity. Patients with acute DVT received oral ximelagatran (24, 36, 48 or 60 mg twice daily) or dalteparin and warfarin for 2 weeks. Evaluation of paired venograms from 295 of 350 patients showed regression of the thrombus in 69% of patients treated with ximelagatran and 69% of patients treated with dalteparin and warfarin. Progression was observed in 8% and 3% of patients, respectively. Changes in thrombus size according to the Marder score were similar in all groups. Treatment discontinuation due to bleeding occurred in two patients receiving ximelagatran (24- and 36-mg groups) and in two patients receiving dalteparin and warfarin. Reduction in pain, edema and circumference of the affected leg was similar in all groups. Oral ximelagatran appears to be a promising alternative to current anticoagulant therapy to limit the progression of acute DVT, and it seems to possess a wide therapeutic window.

Acute antithrombotic effects of ximelagatran, an oral direct thrombin inhibitor, and r-hirudin in a human ex vivo model of arterial thrombosis.

BACKGROUND: Thrombin plays a major role in thrombus formation through activation of platelets and conversion of fibrinogen to fibrin. OBJECTIVES: To investigate the antithrombotic effects of the oral direct thrombin inhibitor (DTI) ximelagatran and the parenteral DTI r-hirudin in humans. SUBJECTS AND METHODS: Healthy male volunteers randomized into four parallel groups each with 15 subjects received either ximelagatran (20, 40 or 80 mg orally) or r-hirudin (0.4 mg kg-1 intravenous bolus + infusion of 0.15 mg kg-1 h-1 for 2 h and 0.075 mg kg-1 h-1 for 3 h). Antithrombotic effects were assessed as changes in total thrombus area (TTA) and total fibrin area (TFA) from baseline, using the Badimon perfusion chamber model at baseline and 2 h and 5 h after drug administration. RESULTS: Two hours postdosing, ximelagatran showed antithrombotic effects at both high and low shear rates (TTA% of mean baseline value +/- SEM was 76 +/- 13% and 71 +/- 17% [both P < 0.05] for the 20-mg dose, 85 +/- 11% [P > 0.05] and 62 +/- 15% [P < 0.05] for the 40-mg dose and 60 +/- 11% and 26 +/- 7% [both P < 0.05] for the 80-mg dose, respectively). r-Hirudin also showed a significant antithrombotic effect at high and low shear rates (76 +/- 11% [P = 0.05] and 57 +/- 17% [P < 0.05] of baseline values, 2 h postdosing, respectively). The inhibitory effects on TFA were similar to those on TTA. CONCLUSIONS: The oral DTI ximelagatran shows antithrombotic effects under both high and low shear conditions. The antithrombotic effect of 40-80 mg ximelagatran appeared comparable to that of parenterally administered r-hirudin, which has been previously demonstrated to be clinically effective in acute coronary syndromes.

Regional haemodynamic effects of endothelin-1 in rat and man: unexpected adverse reaction.

Endothelin-1 was infused into the non-dominant brachial artery in two male subjects. We then monitored intra-arterial mean blood pressure, right atrial pressure, the heart rate and forearm blood flow (by plethysmography). Endothelin-1 at a dose of 5 x 10(-14) to 5 x 10(-9) mol, infused over 5 min periods, elicited no major changes in mean arterial pressure, heart rate and right atrial pressure. We observed an initial increase in forearm blood flow, followed by dose-dependent decreases of 25, 34 and 42% at 5 x 10(-11) to 5 x 10(-9) mol. A higher dose of endothelin-1, 5 x 10(-8) mol, given to only one of the subjects, elicited sweating and vomiting. In this subject, mean arterial pressure, right atrial pressure and the heart rate did not change, while forearm blood flow increased transiently. A deep muscular pain developed in the forearm receiving the endothelin-1 infusion after 30 min (maximum 2 h, duration 10 h), and this pain was intensified by touch and muscle contractions. The force of muscle contractions in the forearm was markedly reduced and a visible oedema developed. In order to investigate the mechanisms of oedema formation, endothelin-1 (10(-10) to 5 x 10(-8) mol/l) was given intra-arterially in a rat hindquarter preparation which was perfused at a constant flow rate. In the rat, endothelin-1 increased both pre- and postcapillary resistance, leading to an increase in capillary hydrostatic pressure and a marked net transcapillary fluid transfer from the perfusate to tissue. There was no sign of increased vascular permeability.(ABSTRACT TRUNCATED AT 250 WORDS)

The importance of enzyme inhibition kinetics for the effect of thrombin inhibitors in a rat model of arterial thrombosis.

The relation between the antithrombotic effect in vivo, and the inhibition constant (Ki) and the association rate constant (k(on)) in vitro was investigated for eight different thrombin inhibitors. The carotid arteries of anaesthetized rats were exposed to FeCl3 for 1 h, and the thrombus size was determined from the amount of incorporated 125I-fibrinogen. The thrombin inhibitors were given intravenously, and complete concentration- and/or dose-response curves were constructed. Despite a 50,000-fold difference between the Ki-values comparable plasma concentrations of hirudin and melagatran were needed (0.14 and 0.12 micromol l(-1), respectively) to obtain a 50% antithrombotic effect (IC50) in vivo. In contrast, there was a comparable in vitro (Ki-value) and in vivo (IC50) potency ratio for melagatran and inogatran, respectively. These results can be explained by the concentration of thrombin in the thrombus and improved inhibition by the low-molecular-weight compounds. For all eight thrombin inhibitors tested, there was an inverse relationship between k(on)-values in vitro and the slope of the dose response curves in vivo. Inhibitors with k(on)-values of < 1 x 10(7) M(-1) s(-1) gave steep dose response curves with a Hill coefficient > 1. The association time for inhibition of thrombin for slow-binding inhibitors will be too long to give effective antithrombotic effects at low plasma concentrations, but at increasing concentrations the association time will decrease, resulting in a steeper dose-response curve and thereby a more narrow therapeutic interval.

Factor V Leiden (G1691A) and prothrombin gene G20210A mutations as potential risk factors for venous thromboembolism after total hip or total knee replacement surgery.

Patients (n = 1600) from 12 European countries, scheduled for elective orthopaedic hip or knee surgery, were screened for Factor V Leiden and prothrombin gene G20210A mutations, found in 5.5% and 2.9% of the populations, respectively. All patients underwent prophylactic treatment with one of four doses of melagatran and ximelagatran or dalteparin, starting pre-operatively. Bilateral ascending venography was performed on study day 8-11. The patients were subsequently treated according to local routines and followed for 4-6 weeks postoperatively. The composite endpoint of screened deep vein thrombosis (DVT) and symptomatic pulmonary embolism (PE) during prophylaxis did not differ significantly between patients with or without these mutations. Symptomatic venous thromboembolism (VTE) during prophylaxis and follow-up (1.9%) was significantly over-represented among patients with the prothrombin gene G20210A mutation (p = 0.0002). A tendency towards increased risk of VTE was found with the Factor V Leiden mutation (p = 0.09). PE were few, but significantly over-represented in both the Factor V Leiden and prothrombin gene G20210A mutated patients (p = 0.03 and p = 0.05, respectively). However, since 90% of the patients with these genetic risk factors will not suffer a VTE event, a general pre-operative genotyping is, in our opinion, of questionable value.

Effects of melagatran, a new low-molecular-weight thrombin inhibitor, on thrombin and fibrinolytic enzymes.

Melagatran, a new, competitive and rapid inhibitor of thrombin with a molecular mass of 429 Da is described. Melagatran is well tolerated when administered in very high doses, and the oral bioavailability in the dog is relatively high. The aim of the study was to determine, in the preclinical setting, the degree of selectivity against the fibrinolytic system required for entering the clinical development phase. Melagatran was compared with two structurally similar thrombin inhibitors, inogatran and H 317/86. The potent inhibition of thrombin by melagatran was demonstrated by a low inhibition constant (Ki) for thrombin (0.002 micromol/l) and prolongation of clotting time to twice the control value in coagulation assays at low concentrations (0.010, 0.59 and 2.2 micromol/l for thrombin time, activated partial thromboplastin time and prothrombin time, respectively). Furthermore, thrombin-induced platelet aggregation was inhibited at the same concentration (IC50-value 0.002 micromol/l) as the Ki-value for thrombin. In two assays of global fibrinolysis, inhibition was observed at a concentration of 1.1 micromol/l in a euglobulin plasma fraction model, while no inhibition was observed at a concentration of < or = 10 micromol/l in a plasma model. In an in vivo model of endogenous fibrinolysis in the rat, inhibition of fibrinolysis was observed at > or = 1.0 micromol/l. In all assays, except the Ki-ratio determinations, the compounds could be graded with regard to selectivity against the fibrinolytic system: inogatran > melagatran > H 317/86. For melagatran, inhibition of fibrinolysis was not observed at concentrations below the upper limit of the proposed therapeutic plasma concentration interval (< 0.5 micromol/l). Thus, melagatran seems to have a sufficient selectivity against the fibrinolytic system, while H 317/86 was considered to be insufficient for clinical development.

Effects of thiopental on resistance vessels in cat skeletal muscle.

Barbiturates are used clinically as anaesthetics and to reduce raised intracranial pressure. One side effect is hypotension, usually ascribed to a depression of cardiac contractility, while their effects on the resistance vessels are more controversial: both vasodilation and vasoconstriction have been described. This study analyzes the effects of thiopental on basal vascular tone in the cat skeletal muscle. We found that total resistance increased by almost 20% at low (50 mumol/l) and decreased down to about 50% of control at high (350 mumol/l) plasma concentrations of thiopental. The vasoconstriction dominated in the large arterioles (i.d. greater than 25 microns) and the vasodilation in the small arterioles (i.d. less than 25 microns). A dose-dependent inhibition of myogenic vascular reactivity (here defined as the maximum resistance increase to a transient rise in transmural pressure) coincided with the vasodilation. Autoregulation of blood flow was depressed by thiopental. During vasoconstriction there was a net transcapillary fluid absorption and during vasodilation a net fluid filtration. The fluid movements could be ascribed to variations in capillary hydrostatic pressure. If applicable to the cerebral circulation these results suggest that thiopental at high plasma concentrations might induce, instead of reduce, interstitial brain oedema.

Antithrombotic effects and bleeding time of thrombin inhibitors and warfarin in the rat.

Warfarin limits the synthesis of y-glutamyl carboxylated forms of coagulation factors, factor II, factor VII, factor IX, and factor X, protein C, and protein S and as a result impairs the function of these proteins. In contrast, direct inhibitors of thrombin only affect one enzyme in the coagulation cascade. The aim of this study was to investigate the antithrombotic effect and the slope of the dose-response curves of the multifactorial coagulation inhibitor warfarin in comparison with the single factor low-molecular-weight thrombin inhibitors melagatran and inogatran. An arterial thrombosis model in rats was used, and vessel damage was induced by topical application of ferric chloride to the carotid artery. The slopes of the dose-response curves were 3.6, 1.8, 1.1, and 1.2, for warfarin, heparin, inogatran, and melagatran, respectively. For warfarin the antithrombotic effect increased from 23% to 81% when the dose was doubled. In contrast, 10-fold increases in the doses of inogatran and melagatran were necessary to obtain a similar increase in antithrombotic effect. The doses needed to obtain 80% antithrombotic effect for heparin, warfarin, and melagatran were investigated in a tail transection bleeding model. For heparin, this dose significantly prolonged the bleeding time and the blood loss; for warfarin, only the total bleeding time was increased while for melagatran there was no increase in bleeding. We conclude that, thrombin inhibitors affecting only one enzyme in the coagulation cascade seem preferable to inhibitors affecting multiple enzymes, such as warfarin, due to shallower dose-response curves and a wider therapeutic interval.

Effect of activated prothrombin complex concentrate or recombinant factor VIIa on the bleeding time and thrombus formation during anticoagulation with a direct thrombin inhibitor.

UNLABELLED: Melagatran is the active form of the oral, direct thrombin inhibitor H 376/95. In several animal models of thrombosis, the antithrombotic properties of melagatran have been demonstrated, without any increase in experimental bleeding. However, as with all anticoagulants, in emergency situations, reversal of the anticoagulation may be necessary. In this study, increasing doses of activated prothrombin complex concentrate (APCC, Feiba) or recombinant factor VIIa (r-F VIIa, NovoSeven) were superimposed on high doses of melagatran, or saline, in anaesthetised rats. The haemostatic effect was evaluated in two bleeding time models and a potential prothrombotic effect was evaluated in an arterial thrombosis model. Compared with melagatran alone (0.5 micromol/kg/h), Feiba in doses of > or =25 U/kg significantly shortened the prolonged bleeding time and reduced blood loss. In addition, Feiba > or =50 U/kg when added to melagatran (2 micromol/kg/h), significantly reduced bleeding time. No potentiation of thrombus formation was observed when Feiba was added to melagatran, compared with controls. NovoSeven at high doses (2-10 mg/kg) produced a nonsignificant trend in reduction of blood loss and with the highest dose (10 mg/kg) producing only a mild nonsignificant reduction in bleeding time. The prolonged prothrombin time (PT) and the ecarin clotting time (ECT) were more effectively shortened by Feiba than by NovoSeven. In contrast, whole blood clotting time (WBCT) was more effectively shortened by NovoSeven than by Feiba. Activated partial thromboplastin time (APTT) was shortened by NovoSeven but was prolonged by Feiba. Thrombin-antithrombin (TAT) complex formation was increased in a dose-dependent fashion more effectively by Feiba than by NovoSeven. CONCLUSION: Feiba (APCC) reversed prolonged bleeding time and blood loss in rats treated with high doses of melagatran and compared with the control group thrombus formation was not potentiated. NovoSeven (r-F VIIa) at high doses had less pronounced effects on blood loss and bleeding times compared with Feiba.

Effects of agents, used to treat bleeding disorders, on bleeding time prolonged by a very high dose of a direct thrombin inhibitor in anesthesized rats and rabbits.

Melagatran is the active form of the oral, direct thrombin inhibitor, H 376/95, that is under evaluation in clinical trials for the prevention and treatment of thromboembolism. In this study, a single dose, calculated on body weight basis, of antifibrinolytic treatment, factor VIIa, factor VIII with and without von Willebrand factor (vWF), factor IX, activated (APCC) or nonactivated (PCC) prothrombin complex concentrates was given intravenously to rats and rabbits, in an attempt to reverse the prolonged bleeding time during intensive anticoagulation with melagatran (2 micromol/kg/h). The doses used were at or above human therapeutic doses. The cutaneous tail bleeding time in the rat, as well as the ear incision bleeding time and cuticle bleeding time, and the blood loss in the rabbit were used for evaluation of the hemostatic effects of these agents. In vivo Feiba (APCC) and Prothromplex-T (PCC) shortened the prolonged cutaneous bleeding times in rats (P<.05); Feiba and Autoplex (APCC) shortened the cutaneous bleeding times in rabbits (P<.05). In contrast, Prothromplex-T prolonged bleeding times and blood loss in the rabbits (P<.05). Ex vivo Feiba, Autoplex and NovoSeven (rF VIIa) significantly (P<.05) shortened the prolonged whole blood clotting time (WBCT). Prothromplex-T significantly prolonged WBCT, activated clotting time (ACT) and activated partial thromboplastin time (APTT). Feiba, Autoplex, and Prothromplex-T increased thrombin generation measured as increased thrombin-antithrombin complex (TAT) formation. In conclusion, APCCs were found to be the most effective agents for reversing bleeding time induced by a very high plasma concentration of melagatran. APCC and recombinant activated factor FVII (rF VIIa) effectively shortened the prolonged WBCT. Thus, stimulating thrombin generation with the use of APCC may counteract the anticoagulant effect observed with a very high dose of a thrombin inhibitor.

The effects of oral and intravenous direct thrombin inhibitors on the size of photochemically induced cortical infarction in rats.

Oral thrombin inhibitors are under development as potential drugs for prophylaxis and treatment of thrombotic events. The effect of pretreatment with two direct thrombin inhibitors, melagatran and inogatran, was evaluated in a rat model of cerebral infarction. Ischaemic stroke was induced by photochemical reaction after an injection of Rose Bengal and focused posterior and to the right of the intersection of the coronal and sagittal sutures on the intact calvarium. A single oral dose of melagatran (30 micromol/kg) significantly reduced the volume of the cortical infarct by 53% (P<.05) compared with control. In addition, following intravenous inogatran (6 micromol/kg) or oral inogatran (100 micromol/kg), the volume of the cortical infarct decreased by 83% and 19%, respectively, compared with control. This study showed that experimental focal ischaemic infarction, elicited by photochemically induced endothelial cell damage, can be significantly reduced with melagatran and inogatran, direct thrombin inhibitors.

In vitro effects of inogatran, a selective low molecular weight thrombin inhibitor.

The thrombin inhibitor inogatran is a synthetic peptidomimetic with a molecular weight of 439 dalton. In vitro studies have shown that inogatran is a classical competitive inhibitor of the active site of thrombin with a Ki of 15 x 10(-9) mol/l. Inogatran doubles the thrombin clotting time in human plasma at 20 x 10(-9) mol/l, APTT at 1.2 x 10(-6) mol/l, and prothrombin time at 4 x 10(-6) mol/l. The effects on rat and dog plasma are similar although slightly weaker. IC50 for inhibition of thrombin-induced aggregation of human platelets is 17 x 10(-9) mol/l. Inogatran has no effect on platelet aggregation induced by ADP or collagen. Up to a concentration of 10 x 10(-6) mol/l inogatran does not inhibit t-PA-induced fibrinolysis as seen in an ECLT system. Inogatran has good selectivity for thrombin as compared to several other serine proteases occurring in the blood. It is concluded that the properties of inogatran in vitro make the compound suitable for further studies in animals and man.

The direct thrombin inhibitor melagatran and its oral prodrug H 376/95: intestinal absorption properties, biochemical and pharmacodynamic effects.

UNLABELLED: Suboptimal gastrointestinal absorption is a problem for many direct thrombin inhibitors. The studies presented herein describe the new oral direct thrombin inhibitor H 376/95, a prodrug with two protecting residues added to the direct thrombin inhibitor melagatran. Absorption properties in vitro: H 376/95 is uncharged at intestinal pH while melagatran is charged. H 376/95 is 170 times more lipophilic (octanol water partition coefficient) than melagatran. As a result, the permeability coefficient across cultured epithelial Caco-2 cells is 80 times higher for H 376/95 than for melagtran. Pharmacokinetic studies in healthy volunteers: H 376/95 is converted to melagatran in man. Oral bioavailability, measured as melagatran in plasma, is about 20% after oral administration of H 376/95, which is 2.7-5.5 times higher than after oral administration of melagatran. The variability in the area under the drug plasma concentration vs. time curve (AUC) is much smaller with oral H 376/95 (coefficient of variation 20%) than with oral melagatran (coefficient of variation 38%). Pharmacodynamic properties: H 376/95 is inactive towards human alpha-thrombin compared with melagatran [inhibition constant (K(i)) ratio, 185 times], a potential advantage for patients with silent gastrointestinal bleeding. In an experimental thrombosis model in the rat, oral H 376/95 was more effective than the subcutaneous low molecular weight heparin dalteparin in preventing thrombosis. CONCLUSION: By the use of the prodrug principle, H 376/95 endows the direct thrombin inhibitor melagatran with pharmacokinetic properties required for oral administration without compromising the promising pharmacodynamic properties of melagatran.