Prognosis of autoimmune hemolytic anemia in critically ill patients.

Patients with autoimmune hemolytic anemia (AIHA) may require intensive care unit (ICU) admission. In order to describe the characteristics of AIHA patients in ICU and identify prognosis factors, clinical and biological data from 44 patients admitted in one ICU between 2002 and 2015 were retrospectively analyzed. The main reasons for ICU admission were profound anemia without any organ failure in 19 patients (either for safer transfusion or continuous monitoring only). Twenty-five (57%) patients had a past history of hemopathy. Twenty patients presented with a direct anti-globulin test (DAT) positive for immunoglobulin G (DAT-IgG) only (46%), 8 with a DAT positive for both IgG and complement (DAT-IgG+C) (36%), and 16 with a DAT positive for complement only (DAT-IgG+C) (18%). Corticosteroids and rituximab were administered to respectively 44 (100%) and 12 (25%) patients. Red blood cell transfusion was required in 28 (64%) patients. Ten (23%) patients received vasopressors. Renal replacement therapy was necessary in 14 (31.8%) patients. Thirteen (30%) patients died in the ICU. There was no difference between survivors and non-survivors regarding associated comorbidities like hemopathy (18/31 [58%] vs. 7/13 [54%], p = 0.80). In decedents, age was higher (72 years [57.8-76.3] vs. 50 years [34.3-64], p < 0.01) and organ dysfunctions were more severe at day 1 (SOFA 8 [7-11] vs. 5.5 [3-7], p < 0.01). Patients with a DAT-IgG displayed poorer outcome in comparison with patients with DAT-IgG+C/C (hospital mortality 69% vs. 36%, p = 0.04). Mortality rate of AIHA patients requiring ICU admission is consequential and appears to be impacted by age, organ failures, and DAT-IgG.

Severe infections requiring intensive care unit admission in kidney transplant recipients: impact on graft outcome.

BACKGROUND: Kidney transplant recipients are at risk for life-threatening infections, which may affect the long-term prognosis. METHODS: We retrospectively included all kidney transplant recipients admitted for sepsis, severe sepsis, or septic shock to the medical intensive care unit (ICU) of the Saint-Louis Hospital, Paris, France, between 2000 and 2010. The main objective was to identify factors associated with survival without graft impairment 90 days after ICU discharge. RESULTS: Data were available for 83 of 100 eligible patients. The main sites of infection were the lungs (54%), urinary tract (24%), and bloodstream (22%). Among documented infections (55/83), 80% were bacterial. Fungal infections were more common among patients transplanted after 2005 (5% vs. 23%, P = 0.02). Mechanical ventilation was used in 46 (56%) patients, vasopressors in 39 (47%), and renal replacement therapy (RRT) in 34 (41%). In-hospital and day-90 mortality rates were 20% and 22%, respectively. On day 90, among the 65 survivors, 39 (47%) had recovered their previous graft function and 26 (31%) had impaired graft function, including 16 (19%) who were dependent on RRT. Factors independently associated with day-90 survival and graft function recovery were baseline serum creatinine (odds ratio [OR] for a 10 µmol/L increase 0.94, 95% confidence interval [CI] 0.88-1.00) and cyclosporine therapy (OR 0.30, 95% CI 0.11-0.79). CONCLUSION: Sepsis was chiefly related to bacterial pneumonia or urinary tract infection. Pneumocystis jirovecii was the leading opportunistic agent, with a trend toward an increase over time. Infections often induced severe graft function impairment. Baseline creatinine and cyclosporine therapy independently predicted the outcome.

Psychological impact of medical evacuation for ICU saturation in Covid-19-related ARDS patients.

PURPOSE: Psychological impact of Medical Evacuation (MEDEVAC) in Covid-19 patients is undetermined. The objectives were to evaluate: Post-traumatic Stress Disorder (PTSD) in MEDEVAC patients hospitalized in ICU for Covid-19-related acute respiratory distress syndrome (ARDS) compared to control group; anxiety, depression rates and outcomes in patients and PTSD in relatives. MATERIAL AND METHODS: This is a retrospective multicentric 1/1 paired cohort performed in 10 ICUs in the West of France. Evaluation was performed 18 months after discharge. Patients and closest relatives performed IES-R (Impact and Event Scale-Revised) and/or HADS (Hospital Anxiety and Depression Scale) scales. RESULTS: Twenty-six patients were included in each group. Patients were 64 ± 11 years old, with 83% male. We report 12 vs 20% of PTSD in control vs MEDEVAC groups (p = 0.7). Anxiety disorder affected 43.5 vs 28.0% (p = 0.26) and depression 12.5 vs 14.3% (p > 0.99) in control vs MEDEVAC groups. PTSD affects 33.3 vs 42.1% of closest relatives (p = 0.55). Ways of communication were adapted: video calls were more frequent in MEDEVAC patients (8.7 vs 60.9%, p < 0.01) whereas physical visits concerned more control group (45.8 vs 13.0%, p = 0.01). CONCLUSIONS: PTSD rate were similar between groups. Adaptive ways of communication, restricted visits and global uncertainties could explain the absence of differences.

Dexamethasone in patients with acute lung injury from acute monocytic leukaemia.

The use of steroids is not required in myeloid malignancies and remains controversial in patients with acute lung injury (ALI) or acute respiratory distress syndrome (ARDS). We sought to evaluate dexamethasone in patients with ALI/ARDS caused by acute monocytic leukaemia (AML FAB-M5) via either leukostasis or leukaemic infiltration. Dexamethasone (10 mg every 6 h until neutropenia) was added to chemotherapy and intensive care unit (ICU) management in 20 consecutive patients between 2005 and 2008, whose data were compared with those from 20 historical controls (1994-2002). ICU mortality was the primary criterion. We also compared respiratory deterioration rates, need for ventilation and nosocomial infections. 17 (85%) patients had hyperleukocytosis, 19 (95%) had leukaemic masses, and all 20 had severe pancytopenia. All patients presented with respiratory symptoms and pulmonary infiltrates prior to AML FAB-M5 diagnosis. Compared with historical controls, dexamethasone-treated patients had a significantly lower ICU mortality rate (20% versus 50%; p = 0.04) and a trend for less respiratory deterioration (50% versus 80%; p = 0.07). There were no significant increases in the rates of infections with dexamethasone. In conclusion, in patients with ALI/ARDS related to AML FAB-M5, adding dexamethasone to conventional chemotherapy seemed effective and safe. These results warrant a controlled trial of dexamethasone versus placebo in AML FAB-M5 patients with noninfectious pulmonary infiltrates.

Nationwide survey on training and device utilization during tracheal intubation in French intensive care units.

BACKGROUND: Intubation is a lifesaving procedure that is often performed in intensive care unit (ICU) patients, but leads to serious adverse events in 20-40% of cases. Recent trials aimed to provide guidance about which medications, devices, and modalities maximize patient safety. Videolaryngoscopes are being offered in an increasing range of options and used in broadening indications (from difficult to unremarkable intubation). The objective of this study was to describe intubation practices and device availability in French ICUs. MATERIALS AND METHODS: We conducted an online nationwide survey by emailing an anonymous 26-item questionnaire to physicians in French ICUs. A single questionnaire was sent to either the head or the intubation expert at each ICU. RESULTS: Of 257 ICUs, 180 (70%) returned the completed questionnaire. The results showed that 43% of intubators were not fully proficient in intubation; among them, 18.8% had no intubation training or had received only basic training (lectures and observation at the bedside). Among the participating ICUs, 94.4% had a difficult intubation trolley, 74.5% an intubation protocol, 92.2% a capnography device (used routinely to check tube position in 69.3% of ICUs having the device), 91.6% a laryngeal mask, 97.2% front-of-neck access capabilities, and 76.6% a videolaryngoscope. In case of difficult intubation, 85.6% of ICUs used a bougie (154/180) and 7.8% switched to a videolaryngoscope (14/180). Use of a videolaryngoscope was reserved for difficult intubation in 84% of ICUs (154/180). Having a videolaryngoscope was significantly associated with having an intubation protocol (P = 0.043) and using capnography (P = 0.02). Airtraq® was the most often used videolaryngoscope (39.3%), followed by McGrath®Mac (36.9%) then by Glidescope® (14.5%). CONCLUSION: Nearly half the intubators in French ICUs are not fully proficient with OTI. Access to modern training methods such as simulation is inadequate. Most ICUs own a videolaryngoscope, but reserve it for difficult intubations.

Influence of severity of myocardial injury on distribution of macromolecules: extravascular versus intravascular gadolinium-based magnetic resonance contrast agents.

OBJECTIVES: This study sought to 1) compare the distribution of extravascular (573 Da) and intravascular (92 kDa) magnetic resonance (MR) contrast agents in reperfused infarcted myocardium, and 2) investigate the effect of injury severity on these distribution patterns. BACKGROUND: Myocardial distribution of low and high molecular weight contrast agents depends on vascular permeability, diffusive/convective transport within the interstitium and accessibility of the intracellular compartment (cellular integrity). METHODS: To vary the severity of myocardial injury, 72 rats were subjected to 20, 30, 45 or 75 min (n = 18, respectively) of coronary artery occlusion. After 2 h of reflow, the animals received either 0.05 mmol/kg of gadolinium-diethylenetriaminepentaacetic acid-bismethylamide (Gd-DTPA-BMA) (n = 24), (Gd-DTPA)30-albumin (n = 24) or saline (control group, n = 24). Three minutes after injection, the hearts were excised and imaged (spin-echo imaging parameters: repetition time 300 ms, echo time 8 ms, 2-tesla system), followed by triphenyltetrazolium chloride staining for infarct detection and sizing. RESULTS: Histomorphometric and MR infarct size (expressed as percent of slice surface) correlated well: r = 0.96 for Gd-DTPA-BMA; r = 0.95 for (Gd-DTPA)30-albumin. On Gd-DTPA-BMA-enhanced images, reperfused myocardial infarctions were homogeneously enhanced. The ratio of signal intensity of infarcted/ normal myocardium increased with increasing duration of ischemia (overall p < 0.0001, analysis of variance [ANOVA]), indicating an increase in the distribution volume of Gd-DTPA-BMA in postischemic myocardium. On (Gd-DTPA)30-albumin-enhanced images, reperfused infarctions consisted of a bright border zone and a less enhanced central core. The extent of the core increased with increasing duration of ischemia (overall p value < 0.0001, ANOVA). CONCLUSIONS: At 2 h of reperfusion, the distribution of MR contrast agents in postischemic myocardium is 1) specific for extravascular and intravascular agents, and 2) modulated by the duration of ischemia.

Effects of dietary supplementation with alpha-tocopherol on myocardial infarct size and ventricular arrhythmias in a dog model of ischemia-reperfusion.

OBJECTIVES: We investigated whether dietary supplementation with the antioxidant vitamin alpha-tocopherol (500 mg daily) might reduce lethal ventricular arrhythmias and infarct size. BACKGROUND: Previous studies suggested that dietary supplementation with alpha-tocopherol may be associated with a reduced risk of ischemic heart disease. However, the mechanism of this protection remains unknown. METHODS: Beagle dogs were randomized to either a supplemented or a control group. Because of the low mortality rate in the supplemented group, five dogs were added to the control group. After 2 months, dogs were anesthetized and underwent a 2-h coronary artery occlusion and 6-h reperfusion. Plasma vitamin E, retinol and malondialdehyde concentrations were assessed in all dogs. RESULTS: Fourteen dogs (11 of 25 control vs. 3 of 19 supplemented dogs, p < 0.05) developed ventricular fibrillation during either ischemia or reperfusion. Malondialdehyde concentrations were higher in dogs that subsequently developed arrhythmias (2.7 +/- 0.2 mumol/liter, mean +/- SEM) compared with dogs that did not (2.1 +/- 0.2 mumol/liter, p = 0.03). Among survivors with significant ischemia, infarct size was larger in supplemented (n = 12, 58.5 +/- 3.3% of area at risk) than in control (n = 11, 41.9 +/- 6.5%, p < 0.04) dogs. In addition, for a given collateral flow, supplemented dogs (n = 16) developed larger infarct size than control dogs (n = 15, p < 0.001, analysis of covariance). CONCLUSIONS: The data suggest that dietary alpha-tocopherol supplementation prevented lethal ventricular arrhythmias associated with ischemia and reperfusion. However, its influence on infarct size and long-term prognosis warrants further investigation.

Investigation of S-farnesyl transferase substrate specificity with combinatorial tetrapeptide libraries.

Using biased tetrapeptide libraries made up of proteinogenic amino acids of the general formula Cys-O2-X3-X4, we searched for new substrates of partly purified rat brain S-farnesyl transferase (FTase). To achieve this task, an assay was developed in which the consumption of the co-substrate (farnesyl pyrophosphate) was measured. After three steps of deconvolution including each synthesis and enzymatic assay, the most efficient substrates found under these particular conditions were Cys-Lys-Gln-Gln (peptide I) and Cys-Lys-Gln-Met (peptide II). As a control, we used another tetrapeptide library (Cys-Val-O3-X4) in which the valine position was arbitrarily fixed, corresponding to Cys-Val-Ile-Met in the CAAX box of K-RasB, although this sublibrary was only marginally active compared with Cys-Lys-X3-X4 in the first round of deconvolution. The best substrate sublibrary was Cys-Val-Thr-X4, threonine being more favourable than the aliphatic amino acids (Val, Ile, Leu, Ala) in this position. Deconvolution finally led to Cys-Val-Thr-Gln, -Met, -Thr and -Ser as the most efficient substrates of FTase. Those tetrapeptides were not substrates of a partly purified geranylgeranyl transferase 1 (GGTase1). We also investigated the influence of the -1 position (at the N-terminus of cysteine) on the specificity of the enzyme, by using a series of pentapeptides constructed on the basis of the best tetrapeptide core (peptide 1). Among this family of analogues, only His-Cys-Lys-Gln-Gln did not behave as a substrate, whereas all the other pentapeptides were measurable substrates, with Gly-, Asn- and Thr-Cys-Lys-Gln-Gln displaying kinetic constants similar to that of Cys-Lys-Gln-Gln. The present work provides strong evidence that the best tetrapeptide substrates of FTase do not necessarily belong to the classical CAAX box, in which A's are lipophilic residues, but rather contain hydrophilic amino acids in the middle of their sequences. Among them, peptides I and II are potent FTase in vitro substrates that are not recognised by GGTase1 and might be new starting points for the design of FTase inhibitors.

[Hyperviscosity syndrome]. / Syndrome d'hyperviscosité plasmatique.

Hyperviscosity syndrome is a life-threatening complication. Clinical manifestations include neurological impairment, visual disturbance and bleeding. Measurement of plasma or serum viscosity by a viscometer assesses the diagnosis. Funduscopic examination is a key exam because abnormalities are well-correlated with abnormal plasma viscosity. Etiologies are various but symptomatic hyperviscosity is more common in Waldenström's macroglobulinemia and multiple myeloma. Prompt treatment is needed: treatment of the underlying disease should be considered, but generally not sufficient. Symptomatic measures aim to not exacerbate blood viscosity while urgent plasmapheresis effectively reduces the paraprotein concentration and relieves symptoms.

Acute renal failure is prevalent in patients with thrombotic thrombocytopenic purpura associated with low plasma ADAMTS13 activity.

BACKGROUND: Among patients with thrombotic microangiopathies, acute kidney injury (AKI) is the hallmark of hemolytic uremic syndrome (HUS) and is largely underestimated in patients with thrombotic thrombocytopenic purpura (TTP). OBJECTIVE: We sought to report AKI features and outcomes in patients with TTP. METHODS: We conducted a retrospective study of 92 patients with TTP assessed by low ADAMTS13 activity (< 10%) between 2001 and 2013. A logistic regression identified variables independently associated with AKI. RESULTS: Among the 92 patients, 54 (58.7%) presented with AKI, including 25 (46.3%) with stage 3 AKI. Fourteen (27.4%) patients had a nephrotic-range proteinuria and 21 (45.6%) had hemoglobinuria. Hematuria and leucocyturia were detected in 19 (41.3%) and 16 patients (36.4%), respectively. Renal replacement therapy (RRT) was required in 14 patients (25.9%). Six months after TTP remission, RRT-free patients had median (IQR) MDRD (Modification of Diet in Renal Disease formula estimating the glomerular filtration rate) of 93 mL min(-1) per 1.73 m(2) (68.8-110) and three patients required long-term dialysis. Mild or moderate chronic renal disease occurred in 23/54 (42.6%) AKI patients. By multivariate analysis, serum level of complement component 3 at admission was the only factor independently associated with AKI (OR per 0.25 unit decrease of C3, 0.85; CI, 1.82-8.33; P = 0.001). CONCLUSIONS: In patients with TTP, AKI is present in more than half the patients, and half of those will have lasting renal effects. Further studies to better understand the pathophysiology of renal involvement in patients with TTP and to identify a subset of patients with TTP syndrome overlapping HUS are warranted.

Life-threatening complications and outcomes in patients with malignancies and severe pulmonary embolism.

BACKGROUND: Data are scarce about ICU patients with malignancy and severe pulmonary embolism. Here, our main objective was to identify risk factors for life-threatening complications, organ failures, and death in ICU patients with severe pulmonary embolism, with special attention to the impact of malignancy. We also described the clinical features of PE in patients with and without malignancies. METHODS: Data from consecutive adults admitted to our ICU in 2002-2011 with severe pulmonary embolism were collected retrospectively. Multivariate analysis was performed to look for factors associated with death, organ failures, or life-threatening complications (major bleeding, recurrent PE, and cardiac arrest). RESULTS: Of 119 included patients (42 [35%] with bilateral pulmonary embolism), 41 had solid malignancies, 27 hematological malignancies, and 51 no malignancies. The most common symptoms were syncope (40%) and hemoptysis (18%) in patients with solid and hematological malignancies, respectively. Life-threatening complications occurred in 23 (19%) patients; risk factors were obesity (OR, 13.22; 1.93-90.70), disseminated intravascular coagulation/ischemic hepatitis (OR, 27.06; 5.14-142.46), fluid load ≥1000 mL/24 h (OR, 6.42; 1.60-25.76), and solid malignancy (OR, 5.45; 1.15-25.89). Inhospital mortality was 27/119 (23%) and respiratory or circulatory failure developed in 36 (30%) patients. Risk factors for these adverse outcomes were older age (OR, 1.04/year; 1.01-1.07), higher oxygen flow rate (OR, 1.28/L; 1.13-1.45); and renal failure (OR, 8.08; 2.50-26.11); whereas chest pain was protective (OR, 0.13; 0.04-0.48). CONCLUSION: In this study, solid malignancy was a risk factor for life-threatening complications but not for death.

Activation of MMP-9 by neutrophil elastase in an in vivo model of acute lung injury.

The effect of neutrophil elastase on the functional status of gelatinases was studied in an hamster model developed by intratracheal administration of lipopolysaccharide followed by in situ cell activation with phorbol myristate acetate. This resulted in the production in bronchoalveolar lavage fluids, in addition to the matrix metalloproteinase MMP-9, of a 75 kDa gelatinase associated with collagenolytic activity. Treatment in vivo with an elastase inhibitor abolished the latter activity. Since, in addition, elastase activates in vitro purified MMP-9 gelatinase into a similar 75 kDa entity, these data suggest that elastase may be a physiological activator of MMP-9 in vivo.

Dual inhibition of human leukocyte elastase and lipid peroxidation: in vitro and in vivo activities of azabicyclo[2.2.2]octane and perhydroindole derivatives.

A series of potent and selective human leukocyte elastase (HLE) inhibitors of the Val-Pro-Val type has been developed. Initially, the central proline residue was replaced by nonnatural amino acids Phi ((2S,3aS,7aS)-perhydroindole-2-carboxylic acid) and Abo ((3S)-2-azabicyclo-[2.2.2]octane-3-carboxylic acid), and secondly several groups able to confer antioxidant properties to the molecule were introduced at the lipophilic N-terminal side chain. When compared to reference inhibitors, in vitro HLE inhibitory potency was maintained (10-100 nM) both with compounds containing the antioxidant moiety at the end of the N-terminal side chain and with compounds in which the N-terminal valine of the tripeptidic sequence had been replaced by a epsilon-substituted lysine. The lipidic peroxidation inhibitory potency of this series of inhibitors was found to be similar to that of the reference antioxidant compounds (around 1 microM). Moreover, HLE-induced hemorrhage in the hamster lung was effectively prevented (40-60% at 15 micrograms/kg) by most of the inhibitors tested when administered intratracheally 3 h before instillation of elastase. Among the most active analogs, compounds 11a,c,g were still active when administered 18 h before elastase. Interestingly, compound 14a was able to prevent HLE-mediated lung damage when administered 72 h prior to enzymatic challenge, indicating exceptional stability and retention in the lung. Finally, in a 14-day chronic model of emphysema in the hamster, 14a significantly conserved alveolar spaces, a marker of lung tissue destruction, and was more potent than reference inhibitor ICI 200 880. This indicates that addition of peroxidation inhibitory properties to an HLE inhibitor can provide a powerful in vivo inhibitor of pulmonary tissue destruction.

Design and synthesis of new linear and cyclic bradykinin antagonists.

We report here on the synthesis and pharmacological properties of a new series of small linear and cyclic peptides derived from the five C-terminal amino acid residues of second-generation bradykinin receptor antagonists. Variations of the two first residues of the pentapeptide (Thi-Ser-D-Tic-Oic-Arg) were shown to modulate the biological activities of the analogs on bradykinin-induced smooth muscle contractions in rabbit jugular vein (RJV), a tissue preparation specific of the B2 bradykinin receptor. Several analogs showed pA2 values around 7 on this tissue preparation, and one cyclic compound, c[-Gly-Thi-D-Tic-Oic-Arg-], 24, in which Thi-Ser was replaced by Gly-Thi, displayed a pA2 of 7.4 on RJV. On the basis of these results, three cyclic molecules and their linear counterparts (compounds 22-24 and 4-6, respectively) were tested on human umbilical vein, a tissue specific of the human B2 receptor. The pKB values obtained for these compounds on these tissue preparations were equivalent to those obtained for the decapeptide NPC 567 (4.8 < pA2 < 5.1). NMR and molecular modeling studies performed on compound 24 clearly demonstrated a type II' beta-turn structure. This analog may serve as a new lead for the design of nonpeptide ligands of the bradykinin B2 receptor subtype.

Tetrapeptide tachykinin antagonists: synthesis and modulation of the physicochemical and pharmacological properties of a new series of partially cyclic analogs.

We report on the synthesis and the pharmacological properties of a new series of tachykinin antagonists based on the pseudopeptide pharmacophore cyclo[-Abo-Asp(D-Trp-Phe-N(Me)Bzl)-] which contains the 2-azabicyclo[2.2.2]octane-3(S)-carboxylic acid (Abo) residue. Variation of the substituents on the tryptophan indole nitrogen was shown to modulate water solubility and transport properties of the analogs as well as potency in classical in vitro response and binding assays. One water-soluble compound, 16, in which the substituent was 3-carbonylpropionate, strongly prolonged the reaction time in the mouse hot-plate test both after iv or oral administration and was devoid of degranulating activity in rat peritoneal mast cells.

Bradykinin and changes in microvascular permeability in the hamster cheek pouch: role of nitric oxide.

1. The objective of this study in the hamster cheek pouch was to investigate the role of nitric oxide in bradykinin-induced microvascular leakage. The cheek pouch microcirculatory bed of the anaesthetized hamster was directly observed under microscope and vascular leakage was evidenced by dextranfluorescein isothiocyanate (FITC-dextran) extravasation. 2. Bradykinin superfusion (but not [des-Arg9]-bradykinin up to 3 x 10(-6) M) induced an increase in microvascular permeability (log EC50: -6.5 +/- 0.4) which was exclusively located on the post-capillary venule. Plasma extravasation was blocked by intravenous pretreatment with Hoe 140, a bradykinin B2 receptor antagonist (estimated log ID50: -9.5 +/- 0.2). 3. The effects of bradykinin (3 x 10(-7) M) superfusion were partially but significantly inhibited by indomethacin (10(-5) M, P < 0.05) and abolished by pretreatment with L-nitro-arginine (L-NOARG; 10(-5) M). 4. Acetylcholine (10(-6) M, which releases endothelial nitric oxide (NO), and sodium nitroprusside (10(-6) M, a nitrovasodilator) superfusion did not induce any changes in permeability, per se. Cromakalim (10(-5) M, a potassium channel opener) superfusion induced a moderate but significant plasma extravasation. 5. The effects of bradykinin, blocked by L-NOARG pretreatment, were restored by the co-perfusion of either sodium nitroprusside or cromakalim. Conversely vasoconstriction, produced by a stable analogue of thromboxane A2 (U46619, 3 x 10(-7) M), inhibited the increase in permeability produced by bradykinin. 6. The measurement of arteriolar diameter showed that bradykinin induced a vasodilatation which was blocked by L-NOARG. L-NOARG in itself was a powerful vasoconstrictor. Sodium nitroprusside and cromakalim, in the presence of L-NOARG, were able to restore the inhibited vasodilator response to bradykinin. 7. These results suggest: (1) bradykinin-induced microvascular leakage is mediated by bradykinin B2 receptor activation; (2) the increase in permeability is due to two different independent phenomena, i.e. post-capillary venular endothelial gap formation and arteriolar vasodilatation which increases the post-capillary venular transmural pressure: (3) NO is only involved in the arteriolar dilatation component of the bradykinin-induced increase in microvascular permeability.

Effects of tachykinins and capsaicin on the mechanical and electrical activity of the guinea-pig isolated trachea.

1. The effects of tachykinins and capsaicin were studied by means of intracellular membrane potential and isometric tension recordings in the isolated trachea of the guinea-pig. 2. The basal membrane potential averaged -51 mV, and most preparations demonstrated spontaneous slow waves. Tetraethylammonium (TEA), a potassium channel blocker (8 x 10(-3) M), depolarized the membrane potential to -44 mV and induced a rhythmic activity. 3. In control solution, substance P (10(-8)-10(-6) M), [Nle10]-neurokinin A(4-10) (10(-8)-10(-6) M) and capsaicin (10(-7)-10(-6) M) induced concentration-dependent depolarizations which were statistically significant at the highest concentration tested (depolarization by 10(-6) M: 8, 11 and 16 mV for the NK1 agonist, the NK2 agonist and capsaicin, respectively). 4. In the presence of TEA (8 x 10(-3) M), the three substances induced depolarizations which were statistically significant at the highest concentration tested for substance P (10(-6) M) and at 10(-7) and 10(-6) M for both [Nle10]-neurokinin A(4-10) and capsaicin (depolarization by 10(-6) M: 11, 17 and 10 mV for substance P, [Nle10]neurokinin A(4-10) and capsaicin, respectively). 5. In the presence or absence of tetraethylammonium, [MePhe7]-neurokinin B (10(-8)-10(-6) M) did not induce any significant changes in membrane potential. 6. The depolarizing effects of substance P (10(-6) M) and [Nle10]-neurokinin A(4-10) (10(-6) M) were blocked only by the specific antagonists for NK1 and NK2 receptors, SR 140333 (10(-7) M) and SR 48968 (10(-7) M), respectively. The effects of capsaicin (10(-6) M) were partially inhibited by each antagonist and fully blocked by their combination. 7. Substance P (10(-9) to 10(-4) M), [Nle10]-neurokinin A(4-10) (10(-10) to 10(-5) M), [MePhe7]-neurokinin B and capsaicin (10(-7) to 10(-5) M) evoked concentration-dependent contractions. 8. The contractions to substance P were significantly inhibited by SR 140333 (10(-8) to 10(-6) M) but unaffected by SR 48968 (10(-8) to 10(-6) M). Furthermore, the response to [Nle10]-neurokinin A(4-10) was significantly inhibited by SR 48968 and unaffected by SR 140333 at the same concentrations. Although SR 48968 (10(-7) M) alone did not influence the effects of substance P, it potentiated the inhibitory effect of SR 140333 (10(-7) M). A similar synergetic effect of these two compounds was observed in the inhibition of the contractile response to [Nle10]-neurokinin A(4-10). 9. Neither SR 140333 (10(-7) M) nor SR 48968 (10(-7) M) alone influenced the contractions to [MePhe7]-neurokinin B and capsaicin. However, the combination of the two antagonists abolished the contractions to either peptide. 10. These results demonstrate that the stimulation of both NK1 and NK2 tachykinin-receptors induced contraction and depolarization of the guinea-pig tracheal smooth muscle and that both receptors were stimulated during the endogenous release of tachykinins by capsaicin. There was no evidence for a major role of NK3 receptors in the contractile and electrical activity of the guinea-pig isolated trachea.

Agonistic and antagonistic properties of the bradykinin B2 receptor antagonist, Hoe 140, in isolated blood vessels from different species.

1. Hoe 140, a recently described bradykinin B2 antagonist, and NPC 567 from an earlier generation of bradykinin B2 antagonists, were tested in rabbit and sheep isolated blood vessels. 2. In rabbit jugular vein, a bradykinin B2 preparation, NPC 567 was an antagonist (apparent pA2: 8.67 +/- 0.16) with marked residual agonistic activity (log[EC50]: -7.29 +/- 0.13). Hoe 140 was a potent non-competitive antagonist devoid of agonistic properties (slope of the Schild plot: 2.02; estimated pA2: 9.04). 3. In rabbit aorta, a bradykinin B1 preparation, NPC 567 was a competitive antagonist (pA2: 6.32 +/- 0.13) but Hoe 140 was ineffective. The two antagonists did not show any agonistic properties in this tissue. 4. In sheep femoral artery without endothelium, bradykinin and Hoe 140 induced contractions with identical efficacy and similar potency (log[EC50]: -8.05 +/- 0.12, -7.73 +/- 0.10; maximal contraction in % of KCl [60 mM]: 59.5 +/- 15.1, 62.0 +/- 13.1; for bradykinin and Hoe 140, respectively). In contrast NPC 567 was an extremely weak agonist. The contractile responses to bradykinin and Hoe 140 were inhibited by NPC 567 (apparent pKB: 6.89 +/- 0.22 and 6.58 +/- 0.08 versus bradykinin and Hoe 140, respectively) but not by a B1 bradykinin antagonist, suggesting that the receptor involved was a bradykinin B2 receptor. 5. In sheep femoral artery with endothelium, bradykinin induced a biphasic response: an endothelium-dependent relaxation and a contraction which were both inhibited by NPC 567 (apparent pKB: 7.10 +/- 0.15) and Hoe 140 (pA2: 8.38 +/- 0.12). As bradykinin B2 receptor antagonists, Hoe 140 and NPC 567 were less potent in the sheep femoral artery than in the rabbit jugular vein. Neither Hoe 140 nor NPC 567 were agonists for the endothelial receptor.6. This study demonstrates that Hoe 140, a new bradykinin B2 receptor antagonist, is more selective and more potent than NPC 567; however, it may possess, depending on the tissue studied, marked residual agonistic properties. Furthermore, bradykinin B2 receptors are subject to important species specificity. Finally, two different bradykinin B2 receptor subtypes may coexist in the sheep femoral artery with endothelium.

Endothelium-derived factors and hyperpolarization of the carotid artery of the guinea-pig.

1. Transmembrane potentials were recorded from isolated carotid arteries of the guinea-pig superfused with modified Krebs-Ringer bicarbonate solution. Smooth muscle cells were impaled from the adventitial side with intracellular glass microelectrodes filled with KCl (30-80 M omega). 2. Acetylcholine (1 microM) in the presence of inhibitors of nitric oxide synthase, (N omega-nitro-L-arginine (L-NOARG) 100 microM) and cyclo-oxygenase, (indomethacin 5 microM) induced an endothelium-dependent hyperpolarization (-18.9 +/- 1.6 mV, n = 15). 3. In the presence of these two inhibitors, S-nitroso-L-glutathione (10 microM), sodium nitroprusside (10 microM), 3-morpholinosydnonimine (SIN-1, 10 microM) and iloprost (0.1 microM) induced endothelium-independent hyperpolarizations of the smooth muscle cells (respectively: -16.0 +/- 2.3, -16.3 +/- 3.4, -12.8 +/- 2.0 and -14.5 +/- 1.5 mV, n = 4-6). 4. The addition of glibenclamide (1 microM) did not influence the acetylcholine-induced L-NOARG/ indomethacin-resistant hyperpolarization (-18.0 +/- 1.8 mV, n = 10). In contrast, the responses induced by S-nitroso-L-glutathione, sodium nitroprusside, SIN-1 and iloprost were abolished (changes in membrane potential: -0.8 +/- 1.1, 1.3 +/- 3.9, 4.5 +/- 4.6 and 0.3 +/- 0.8 mV respectively, n = 4-5). 5. In the presence of NO synthase and cyclo-oxygenase inhibitors, charybdotoxin (0.1 microM) or apamin (0.5 microM) did not influence the hyperpolarization produced by acetylcholine. However, in the presence of the combination of charybdotoxin and apamin, the acetylcholine-induced L-NOARG/indomethacin-resistant hyperpolarization was converted to a depolarization (4.4 +/- 1.2 mV, n = 20) while the endothelium-independent hyperpolarizations induced by S-nitroso-L-glutathione, sodium nitroprusside, SIN-1 and iloprost were not affected significantly (respectively: -20.4 +/- 3.4, -22.5 +/- 4.9, -14.5 +/- 4.7 and -14.5 +/- 0.5 mV, n = 4-5). 6. In the presence of the combination of charybdotoxin and apamin and in the absence of L-NOARG and indomethacin, acetylcholine induced a hyperpolarization (-19.5 +/- 3.7 mV, n = 4). This hyperpolarization induced by acetylcholine was not affected by the addition of indomethacin (-18.3 +/- 4.6 mV, n = 3). In the presence of the combination of charybdotoxin, apamin and L-NOARG (in the absence of indomethacin), acetylcholine, in 5 out of 7 vessels, still produced hyperpolarization which was not significantly smaller (-9.1 +/- 5.6 mV, n = 7) than the one observed in the absence of L-NOARG. 7. These findings suggest that, in the guinea-pig isolated carotid artery, the endothelium-independent hyperpolarizations induced by NO donors and iloprost involve the opening of KATP channels while the acetylcholine-induced endothelium-dependent hyperpolarization (resistant to the inhibition of NO-synthase and cyclo-oxygenase) involves the opening of Ca(2+)-activated potassium channel(s). Furthermore, in this tissue, acetylcholine induces the simultaneous release of various factors from endothelial origin: hyperpolarizing factors (NO, endothelium derived hyperpolarizing factor (EDHF) and prostaglandins) and possibly a depolarizing factor.

Inhibitors of the cytochrome P450-mono-oxygenase and endothelium-dependent hyperpolarizations in the guinea-pig isolated carotid artery.

1. Transmembrane potentials were recorded from isolated carotid arteries of the guinea-pig superfused with modified Krebs-Ringer bicarbonate solution. Smooth muscle cells were impaled with sharp intracellular microelectrodes. 2. Acetylcholine (1 microM) induced an endothelium-dependent hyperpolarization (14.3 +/- 2.8 mV, n = 6) which was not affected (15.1 +/- 1.1 mV, n = 35) by inhibitors of cyclo-oxygenase (indomethacin, 5 microM) and nitric oxide synthase (N omega nitro-L-arginine: L-NOARG, 100 microM). 3. The hyperpolarization produced by acetylcholine was abolished in the presence of elevated potassium (35 mM) in the superfusing physiological saline solution. 4. The acetylcholine-induced hyperpolarization was not affected by the inhibitors of cytochrome P450 mono-oxygenases, SKF525a (10 and 100 microM, 13.9 +/ 2.2 and 15.3 +/- 4.6 mV), metyrapone (100 microM, 13.1 +/- 1.9 mV), clotrimazole (100 microM, 13.5 +/- 2.7 mV), 17-octadecynoic acid (5 microM, 16.5 +/- 1.9 mV), methoxsalen (10 microM, 15.3 +/- 1.6 mV), the inhibitor of phospholipase A2 quinacrine (10 microM 12.8 +/- 2.5 mV) and the non specific lipoxygenases/cyclo-oxygenases/cytochrome P450 inhibitor, eicosatetraynoic acid (50 microM, 15.0 +/- 2.2 mV). However, the muscarinic antagonist, atropine (100 nM), abolished the hyperpolarization. 5. These results suggest that in guinea-pig carotid artery, the metabolism of arachidonic acid, either through cyclo-oxygenase, lipoxygenase or cytochrome p450 mono-oxygenase, is not involved in acetylcholine-induced endothelium-dependent hyperpolarizations.