Increased lung neutrophil apoptosis and inflammation resolution in nonresponding pneumonia.

Neutrophil activation state and its relationship with an inflammatory environment in community-acquired pneumonia (CAP) remain insufficiently elucidated. We aimed to evaluate the neutrophil apoptosis and cytokine pattern in CAP patients after 72 h of treatment, and their impact on infection resolution. Apoptosis of blood and bronchoalveolar lavage (BAL) neutrophils was measured in nonresponding CAP (NCAP), in responding CAP (blood only) and in patients without infection (control). Pro-inflammatory (interleukin (IL)-6, IL-8) and anti-inflammatory (IL-10) cytokines were measured. Main outcomes were clinical stability and days of hospitalisation. Basal neutrophil apoptosis was higher in the BAL and blood of NCAP, whereas spontaneous apoptosis (after 24 h culture) was lower. Cytokines in NCAP were higher than in responding CAP and control: IL-6 was increased in BAL and blood, IL-8 in BAL and IL-10 in blood. An increased basal apoptosis (≥20%) in BAL of NCAP was associated with lower systemic IL-10 (p<0.01), earlier clinical stability (p=0.05) and shorter hospital stay (p=0.02). A significant correlation was found for systemic IL-6 and IL-10 with days to reach stability and length of stay. After 72 h of treatment, an increased basal alveolar neutrophil apoptosis might contribute to downregulation of inflammation and to faster clinical stability.

Bioconversion of polyphenols and organic acids by gut microbiota of predigested Hibiscus sabdariffa L. calyces and Agave (A. tequilana Weber) fructans assessed in a dynamic in vitro model (TIM-2) of the human colon.

The present work aimed at understanding gut microbiota bioconversion of phenolic compounds (PC) and organic acids in predigested Hibiscus sabdariffa (Hb) calyces and the mixture of Hb and Agave (Agave tequilana Weber) fructans (AF). With this purpose, dried Hb and Hb/AF were predigested with enzymatic treatment, and then fermented in a dynamic in vitro model of the human colon (TIM-2). After HPLC-ESI-QToF-MS analysis of samples taken at 0, 24, 48 and 72 h of fermentation, it was observed that hydroxycinnamic acids, flavanols, flavonols, and anthocyanins were mainly transformed into derivatives of hydroxyphenylpropionic, hydroxyphenylacetic and hydroxybenzoic acids. Moreover, organic acids, such as hydroxycitric and hibiscus acids, were formed along with unidentified lactones and reduced compounds. Interestingly, no differences were observed between microbial-derived metabolites formed after the fermentation of Hb and Hb/AF. In conclusion, colonic fermentation of polyphenol-rich Hb yields a wide range of microbial phenolic metabolites with potential effects on health.

Experimental confounding factors affecting stability, transport and metabolism of flavanols and hydroxycinnamic acids in Caco-2 cells.

Studying bioavailability of polyphenols is essential to understand the health effects of these compounds. Human epithelial cells are commonly used in intestinal absorption and transport experiments but the changes polyphenols undergo during incubation, due to their chemical instability under the cell culture conditions, are scarcely known and might lead to inaccurate conclusions. Based on abundance of flavanols and hydroxycinnamic acids in the diet, epicatechin, epicatechin-3-gallate and procyanidin B2 as flavanols along with 5-caffeoylquinic and 3,5-dicaffeoylquinic acids as hydroxycinnamic acids were selected to comparatively evaluate their absorption and metabolism using an in vitro Caco-2 cell model. Special emphasis was paid to the structure-stability relationship of these phenolic compounds in Dulbecco's Modified Eagle's Medium (DMEM) under the cell culture conditions. The tested compounds were scarcely absorbed and minimally metabolized by the intestinal epithelium cells. The cell transport study showed prevalent efflux for flavanols opposite to absorption for hydroxycinnamates. Intestinal metabolism revealed that hydroxycinnamates were preferentially hydrolyzed and subsequently methylated, whereas hydrolysis of flavanols could not be confirmed, being mostly conjugated to sulfate, methyl- and methyl-sulfate derivatives. It is noteworthy that methyl derivatives of procyanidin-B2 were detected inside Caco-2 cells, confirming its absorption. In addition, culture medium influenced phenol isomerization to a higher extent than cells. In conclusion, hydroxycinnamates were better absorbed than flavanols although their bioavailability was limited in this intestinal cell model.

Iron status biomarkers in iron deficient women consuming oily fish versus red meat diet.

Specific recommendations for anemic individuals consist in increasing red meat intake, but the population at large is advised to reduce consumption of red meat and increase that of fish, in order to prevent the risk of developing cardiovascular disease. This study aimed to determine the effects of consuming an oily fish compared to a red meat diet on iron status in women with low iron stores. The study was designed attending the Consolidated Standards of Reporting Trials (CONSORT) statement guidelines. It was a randomised crossover dietary intervention study of two 8-week periods. Twenty-five young women with low iron stores completed the study. Two diets containing a total of 8 portions of fish, meat and poultry per week were designed differing only in their oily fish or red meat content (5 portions per week). At the beginning and the end of each period blood samples were taken and hemoglobin, hematocrit, serum ferritin, serum iron, serum transferrin, serum transferrin receptor-2 and the Zn-protoporphyrin/free-protoporphyrin ratio were determined. Food intake and body weight were monitored. During the oily fish diet, PUFA intake was significantly higher (p=0.010) and iron intake lower (mean+/-SD, 11.5+/-3.4 mg/day vs. 13.9+/-0.1 mg/day, p=0.008), both diets providing lower mean daily iron intake than recommended for menstruating women. Although there were no significant differences after 16 weeks, serum ferritin moderately decreased and soluble transferrin receptor increased with the oily fish, while changes with the red meat diet were the opposite. In conclusion, an oily fish diet compared to a red meat diet does not decrease iron status after 8 weeks in iron deficient women.

A comparative study of iron bioavailability from cocoa supplemented with ferric pyrophosphate or ferrous fumarate in rats.

BACKGROUND: Food iron fortification can be a good strategy to prevent iron deficiency. Iron bioavailability from cocoa powder enriched with ferric pyrophosphate encapsulated in liposomes or ferrous fumarate was assessed in rats. METHODS: Three groups of rats consumed during 28 days either a control diet or two diets prepared with ferric pyrophosphate- or ferrous fumarate-enriched cocoa powder as the unique source of iron. Body weight and food intake were monitored and last-week feces were collected. On day 28, animals were sacrificed and livers and spleens were removed. Hemoglobin and total iron binding capacity (TIBC) were determined. RESULTS: There were no significant differences in body weight and food intake. Apparent iron absorption and % absorption/intake were significantly lower in rats consuming enriched cocoa compared to the control group, without significant differences due to the iron form. Enriched cocoa groups showed significantly lower spleen iron content and concentration than the control. Liver iron was lower in the ferric pyrophosphate group compared to the other two groups. Hemoglobin and TIBC values showed a deficient iron status in ferric pyrophosphate rats. CONCLUSION: Cocoa powder is a good vehicle for iron fortification when enriched with ferrous fumarate compared to ferric pyrophosphate encapsulated in liposomes.

Does bicarbonated mineral water rich in sodium change insulin sensitivity of postmenopausal women?

AIM: To study the effects of drinking 0.5 L of two sodium-rich bicarbonated mineral waters (BMW-1 and 2), with a standard meal, on postprandial insulin and glucose changes. And to determine, if the effects vary depending on insulin resistance, measured by homeostasis model assessment (HOMA). METHODS: In a 3-way randomized crossover study, 18 healthy postmenopausal women consumed two sodium-rich BMWs and a low-mineral water (LMW) with a standard fat-rich meal. Fasting and postprandial blood samples were taken at 30, 60 and 120 min. Serum glucose, insulin, cholesterol and triacylglycerols were determined. Insulin resistance was estimated by HOMA and insulin sensitivity was calculated by quantitative insulin sensitivity check index (QUICKY). RESULTS: Glucose levels did not change. HOMA and QUICKY values were highly inversely correlated (r = -1,000; p < 0.0001). Insulin concentrations showed a significant time effect (p < 0.0001) and a significant water x time interaction (p < 0.021). At 120 min insulin levels with BMW-1 were significantly lower than with LMW (p = 0.022). Postprandial insulin concentrations showed significantly different patterns of mineral water intake depending on HOMA n-tiles (p = 0.016). CONCLUSION: Results suggests an increase in insulin sensitivity after BMWs consumption. This effect is more marked in the women, who have higher HOMA values. These waters should be considered part of a healthy diet in order to prevent insulin resistance and cardiovascular disease.

Antiproliferative and cytotoxic effects of green coffee and yerba mate extracts, their main hydroxycinnamic acids, methylxanthine and metabolites in different human cell lines.

This work aimed at studying the effects of green coffee bean (GCBE) and yerba mate (YME) extracts, their main phenolic components (5-caffeoylquinic acid, 5-CQA; 3,5-dicaffeoylquinic acid, 3,5-DCQA) and metabolites (ferulic acid, FA; caffeic acid, CA; dihydrocaffeic acid, DHCA; and dihydroferulic acid, DHFA) along with caffeine (CAF) on the viability and proliferation of different human cell lines. Extracts (10-1000 µg/mL) and standards (10-1000 µM) were assayed in colon (Caco-2), lung (A549), oesophageal (OE-33), urinary bladder (T24) human carcinoma cells, and a non-cancer cell line (CCD-18Co). YME significantly reduced viability of cancer cells at all assayed concentrations, the higher doses also reducing cell proliferation. GCBE effects on cell viability were more effective at 100 and 1000 µg/mL, showing modest effects on cell proliferation. The highest doses of 5-CQA and 3,5-DCQA reduced cell viability and proliferation in all cell lines, whereas FA, DHCA and DHFA had lower and variable effects. Caffeine had no effect. Dietary-attainable concentrations (0.1, 1 and 10 µg/mL) of YME were tested for cytotoxicity and reactive oxygen species generation, showing no cytotoxic effect. Low concentrations of all tested compounds were non-cytotoxic to CCD-18Co cells. CONCLUSION: YME and to a lower degree GCBE, their phenolic components and metabolites may decrease cancer cell viability and proliferation.

A phenolic extract from grape by-products and its main hydroxybenzoic acids protect Caco-2 cells against pro-oxidant induced toxicity.

Grape/wine industry produces large amounts of by-products, however knowledge on their health-promoting qualities is limited. This study investigated the effects of a grape phenolic extract (GPE) and its phenolic compounds, gallic acid (GA) and syringic acid (SA) on human intestinal Caco-2 cells, directly or after cytotoxicity induced by tert-butylhydroperoxide (t-BOOH). Direct treatment with 0.1-10 µg/mL GPE, or 0.1-10 µM GA and SA produced no major cytotoxic effect, either changes in antioxidant defences (glutathione content, glutathione peroxidase and reductase activities) or protein damage (carbonyl groups). However, 10 µg/mL GPE, 1 and 10 µM GA and 10 µM SA decreased reactive oxygen species (ROS) production. Pre-treatment with GPE, SA and GA at the same concentrations for 20 h showed that 10 µg/mL GPE and 10 µM GA or SA significantly counteracted ROS increase induced by t-BOOH. 10 µg/mL GPE and 1-10 µM GA or 10 µM of SA significantly reduced pro-oxidant-induced cytotoxicity. 1-10 µg/mL GPE, 1-10 µM GA and 10 µM SA significantly recovered both depleted glutathione and enhanced glutathione reductase and peroxidase activities, and reduced protein oxidative damage. Therefore, treatment with realistic concentrations of GPE and its main hydroxybenzoic acids protected Caco-2 cells against induced oxidative stress.

Estimation of iron absorption in humans using compartmental modelling.

OBJECTIVE: To evaluate whether a compartmental model could estimate iron absorption as accurately as the well-validated technique of plasma area under the curve using labelled test meals. DESIGN: The study is a randomised cross-sectional intervention. SETTING: The study was carried out at the Human Nutrition Unit at the Institute of Food Research, Norwich, UK. SUBJECTS: A total of nine female volunteers, aged 33+/-8 y. INTERVENTIONS: Volunteers were given an oral dose (approximately 5 mg) of Fe-57 as iron sulphate in an orange juice test drink and simultaneously infused Fe-58 (approximately 200 microg) as iron citrate over 90 min. Multiple blood samples were taken for the following 6 h. The samples were analysed by mass spectrometry and iron absorption was estimated using a mathematical model based on the appearance of Fe isotopes in plasma and the area under the curve technique. RESULTS: The geometric mean (-1 s.d., +1 s.d.) absorption of the model estimate is 16% (9, 31) and the area under curve estimate is 18% (8, 29). CONCLUSIONS: Results indicate that a compartmental model can be used to estimate labelled iron absorption although it is unlikely that this new method will be used in favour of an existing one. Further studies are now needed with unlabelled iron to assess whether the technique could have application in the assessment of total (haem+nonhaem) iron absorption from food.

Antagonism of calcium by zinc in guinea-pig isolated taenia caeci and trachealis muscle.

1 In guinea-pig isolated taenia caeci and trachealis bathed in a K+-rich, Ca2+-free medium, CaCl2 (0.01-10 mM) produced a concentration-dependent contraction. Zn2+ (0.01-1 mM), Cd2+ (0.01-1 mM), verapamil (0.01-100 microM) and trifluoperazine (1-100 microM) were effective antagonists of CaCl2-induced responses. 2 Zn2+ and Cd2+ in concentrations from 0.01 to 1 mM were without effect on the tone of taenia and trachea in normal Tris solution. Conversely, Zn2+ and Cd2+, in concentrations of 1 mM, caused contraction of these tissues in a K+-rich, Ca2+-free medium. Zn2+ (1 mM)-induced contractions of taenia and trachea were completely inhibited by verapamil (10 microM). 3 In taenia and trachea skinned of their plasma membranes, tension development induced by Ca2+ (10 microM or 1 microM, respectively) was unaffected by verapamil (100 microM), whereas trifluoperazine (100 microM) depressed the maximal tension produced by Ca2+. Segments of skinned preparations contracted in response to low concentrations of Zn2+ (10 microM) or Cd2+ (10 microM). 4 It is concluded that Zn2+ may suppress Ca2+-induced spasm by a direct action on the binding sites of the Ca2+ channel.

Contractile activity of three endothelins (ET-1, ET-2 and ET-3) on the human isolated bronchus.

1. The effects of three endothelins: (i) the classical or human/porcine endothelin (ET-1); (ii) [Trp6, Leu7] endothelin (ET-2) and (iii) [Thr2, Phe4, Thr5, Tyr6, Lys7, Tyr14] endothelin or rat endothelin (ET-3) were tested on the human isolated bronchus. 2. ET-1 produced a concentration-dependent contraction of the human isolated bronchus that proceeded in two different steps. The first step was observed at very low concentrations (pD2 = 11.01 +/- 0.17, n = 10) but corresponded to a low intrinsic activity (Emax = 15.6 +/- 1.8% of Emax = 26.1 +/- 2.9% of ACh 3 x 10(-3) M, n = 5, P less than 0.05), reduced by nicardipine 10(-6) M (Emax = 6.0 +/- 2.6% of ACh 3 x 10(-3) M, n = 5, P less than 0.05) and strongly inhibited in calcium-free medium. The second step of the action of ET-1 corresponded to a lesser potency (pD2 = 7.90 +/- 0.17, n = 9) but a higher intrinsic activity (Emax = 82.5 +/- 4.7% of ACh 3 x 10(-3) M). This effect was not significantly modified by nicardipine 10(-6) M or by Bay K 8644 10(-7) M. Neither of the two effects was modified by indomethacin 3 x 10(-6) M. 3. The effects of ET-2 and ET-3 were qualitatively similar to those of ET-1 but quantitatively different; for these two steps of contracting activity and for potency and efficacy the ranking was: ET-1 greater than ET-2 = ET-3. 4. Thus, ET-1 appears to be the most potent of these three substances in its effect on the human isolated bronchus. Its activity seems to involve the action of voltage-dependent calcium channels at low concentrations (10-12 to 10-9M), whereas other mechanisms are involved at higher concentrations (10-8 to 3 x 10-7M).

Functional, biochemical and morphological studies on human bronchi after cryopreservation.

1. Human isolated bronchi have been investigated as fresh tissue or after storage (7 and 30 days) at -196 degrees C in foetal calf serum containing 1.8 M dimethyl sulphoxide. 2. After cryopreservation, the maximal contractile response to acetylcholine (3 mM) was reduced (approximately 25%) but the difference did not reach significance statistically. Maximal responses to other spasmogens tested (histamine, [Nle10]NKA(4-10), bradykinin, leukotriene D4, U46619, and KCl) did not differ between unfrozen and frozen/thawed tissues. The sensitivity of cryopreserved tissues to the constrictor agents tested was similar to that of fresh tissues. 3. The accumulation of inositol phosphates produced by acetylcholine in human bronchus in vitro was similar in fresh and cryostored (30 days) tissues. 4. Relaxant responses of acetylcholine (0.3 microM)-precontracted preparations to theophylline, isoprenaline, rolipram and sodium nitroprusside were unchanged after storage with the exception of the sensitivity to rolipram which was diminished in the 30-days cryostorage group. 5. Light microscopic examination of sections taken from 30 days cryostored tissues indicates that the epithelium, submucosal tissue and smooth muscle were well preserved. 6. These experiments suggest that cryopreservation of human bronchi results in maintenance of several morphological, functional (contraction/relaxation) and biochemical properties.

Leukotriene receptors on human pulmonary vascular endothelium.

1. Cysteinyl-leukotrienes cause contractions and/or relaxations of human isolated pulmonary vascular preparations. Although, the localization and nature of the receptors through which these effects are mediated have not been fully characterized, some effects are indirect and not mediated via the well-described LT1 receptor. 2. In human pulmonary veins (HPV) with an intact endothelium, leukotriene D4 (LTD4) induced contraction above basal tone. This response was observed at lower concentrations of LTD4 in the presence of nitric oxide synthase inhibitor N omega-nitro-L-arginine (L-NOARG). Contractions (in the absence and presence of L-NOARG) were partially blocked by the LT1 antagonists (MK 571 and ICI 198615). 3. LTD4 relaxed HPV previously contracted with noradrenaline. This relaxation was potentiated by LT1 antagonists, but was abolished by removal of the endothelium. LTD4 also relaxed human pulmonary arteries (HPA) precontracted with noradrenaline but this effect was not modified by LT1 antagonists. 4. The results suggest that contraction of endothelium-intact HPV by LTD4 is partially mediated via LT1 receptors. Further, in endothelium-intact HPV, this contraction was opposed by a relaxation induced by LTD4, dependent on the release of nitric oxide, which was mediated, at least in part, via a non-LT1 receptor. In addition, LTD4 relaxation on contracted HPA was not mediated by LT1 receptors. 5. The mechanical effects of LTD4 on human pulmonary vasculature are complex and involve both direct and indirect mechanisms mediated via at least two types of cysteinyl-leukotriene receptors.

Effects of phorbol 12,13-diacetate on human isolated bronchus.

Protein kinase C appears to be involved in the regulation of airway contractility. Phorbol 12,13-diacetate (PDA; 0.01-10 microM), a protein kinase C activator, produced a transient relaxation followed by a sustained contraction of human isolated bronchus. Different protein kinase C inhibitors (calphostin C, staurosporine and 1-(5-isoquinolinesulfonyl)-2-methylpiperazine) (H-7), nifedipine (NIF; 1 microM) or incubation with Ca(2+)-free medium, inhibited the spasmogenic response to phorbol, while ouabain (10 microM) suppressed only the initial relaxation. These results indicate that the initial relaxation, in response to PDA, is related to the activation of Na(+)/K(+)-ATPase, while the ensuing contraction depends on extracellular Ca(2+) entry.Incubation with PDA (1-5 microM) depressed the maximal relaxation to theophylline and caffeine obtained at 37 degrees C but augmented the spasmogenic responses to methylxanthines (10 mM) obtained in cooled preparations. These effects do not result apparently from increased extracellular entry of Ca(2+), but instead, from facilitation of the release of Ca(2+) from intracellular stores.

Effect of epithelium removal and of enkephalin inhibition on the bronchoconstrictor response to three endothelins of the human isolated bronchus.

The three endothelins ET-1, ET-2 and ET-3 induced a potent contractile response in the human isolated bronchus with an intact epithelium, which proceeded on two different stages. The first stage was observed at low concentrations (high potency) (10(-12) to 10(-9) M) but corresponded to a low intrinsic activity (Emax maximal effect induced by ACh 10(-3) M), the second stage appeared at higher concentrations and corresponded to higher intrinsic activity. The rank order of potency for the two stages of contractile activity was ET-1 greater than ET-2 = ET-3. Removal of the epithelium significantly enhanced the two stages of the contractile responses to the three endothelins and abolished the differences in potency efficacy that were observed between ET-1, ET-2 and ET-3 when the epithelium was present. Phosphoramidon (10(-5) M), an enkephalinase inhibitor, was as potent as epithelium removal in enhancing the contractile responses to these agonists at low concentrations (first stage of contraction, 10(-16) to 10(-9) M). However, with high concentrations of endothelins (greater than 10(-9) M, second stage of contraction), phosphoramidon was less potent than epithelium removal in enhancing the contractile responses. In epithelium-denuded strips, preincubation with phosphoramidon did not further increase the maximal contractions induced by/or the potencies of ET-1, ET-2 or ET-3. After epithelium removal, responses to low doses of endothelins were attenuated by nicardipine (10(-6) M) whereas responses to high doses of the endothelins were not affected, as was also observed when the epithelium was present. (ABSTRACT TRUNCATED AT 250 WORDS)

Halothane inhibits endothelium-dependent relaxation elicited by acetylcholine in human isolated pulmonary arteries.

This study examined whether a clinically relevant concentration of the volatile anaesthetic halothane modifies the endothelium-dependent relaxation produced by acetylcholine (3 nM-10 microM), histamine (1 pM-0.1 microM) and anti-human immunoglobulin E (1:1000) in human isolated pulmonary arteries submaximally precontracted with noradrenaline. An inhibitor of nitric oxide formation, N(G)-nitro-L-arginine (100 microM), attenuated acetylcholine-induced relaxation but failed to inhibit histamine- and anti-human immunoglobulin E-induced relaxation. Indomethacin (2.8 microM, a cyclooxygenase inhibitor) preferentially reduced the relaxation to histamine and anti-human IgE. Halothane (2%) significantly attenuated the relaxation to acetylcholine but had no significant effect on the relaxation elicited by histamine and anti-human IgE. Halothane (2%) enhanced the basal release of prostaglandin I2 by human pulmonary arteries (control 0.31 +/- 0.04 ng mg(-1); treated tissues 0.50 +/- 0.06 ng mg(-1); n = 5; P < 0.05). Halothane (2%) did not alter the responsiveness and sensitivity of preparations to relaxants acting through activation of adenylyl cyclase (forskolin) or guanylyl cyclase (sodium nitroprusside) or by the opening of K(ATP) channels (cromakalim). In conclusion, halothane inhibits the endothelium-dependent relaxation of human pulmonary arteries to acetylcholine by interfering with the nitric oxide pathway at a site before activation of soluble guanylyl cyclase in vascular smooth muscle.

Calcium dependence of the contraction produced by endothelin (ET-1) in isolated guinea-pig trachea.

Endothelin (ET-1, 1 pM to 0.1 microM) produced a concentration-dependent contraction of isolated guinea-pig trachea. BAY K 8644 (1 microM) did not significantly alter the concentration-response curve for ET-1. Incubation with nicardipine (10 microM) partly inhibited responses to low concentrations (10 pM to 1 nM) of ET-1 while verapamil (10 microM) and diltiazem (10 microM) were ineffective. La3+ (10 microM) and Cd2+ (10 microM) preferentially depressed the responses evoked by high concentrations (30 nM-0.1 microM) of ET-1 without affecting the responses evoked by low concentrations of the peptide. Incubation in Ca2(+)-free (with EDTA, 1 mM) medium resulted in suppression of the responses elicited by low concentrations of ET-1 and partial inhibition of the responses elicited by high concentrations of the peptide. It is concluded that responses to ET-1 are dependent on extracellular Ca2+. The promotion of Ca2+ entry by ET-1 is not confined to a particular class of Ca2+ channel. An intracellular source of Ca2+ is also mobilized by high concentrations (greater than 10(-9) M) of ET-1.

Effect of dantrolene sodium in isolated guinea-pig trachea.

The effect of dantrolene sodium (3 microM-0.3 mM) on the spontaneous tone and responses to various contractile agonists was studied in isolated guinea-pig trachea. Dantrolene produced a concentration-related inhibition of the spontaneous tracheal tone, reaching a value of 94.8 +/- 4.8% of the relaxation induced by caffeine 10 mM. Removal of the epithelium did not affect the dantrolene-induced relaxation. Dantrolene did not alter the concentration-response curve for KCl and produced only small displacements of the concentration-response curves for CaCl2, acetylcholine and histamine, without affecting their maximal effects. Dantrolene dose relatedly inhibited the contraction induced by caffeine (1 mM) in Krebs solution containing indomethacin (2.8 microM) at 20 degrees C. The spasm induced by caffeine in Ca2(+)-free Krebs solution (20 degrees C, indomethacin 2.8 microM) was slightly depressed by dantrolene. Dantrolene did not depress the Ca2+ (1 microM)-induced contraction in skinned trachea. These results suggest that besides a possible intracellular site of action, the mechanism of the inhibitory effect of dantrolene in guinea-pig trachea may be related to interference with Ca2+ entry through pathways not susceptible to calcium channel blockers.

Modification by indomethacin of airway contractile responses in normal and sensitized guinea-pigs.

Active sensitization of guinea-pigs resulted in an increase in responsiveness and sensitivity of tracheal and lung parenchymal strips to CaCl2 (in K+-depolarised tissue), KCl, acetylcholine and histamine. Indomethacin (5 microM) preferentially enhanced the response of tracheal strips from normal animals to histamine and to a lesser extent acetylcholine but not to CaCl2 or KCl. A similar trend was observed in sensitized tissues. Indomethacin pretreatment did not cause changes in responsiveness or sensitivity of lung parenchymal strips from normal or sensitized guinea-pigs to the agonists tested. It is concluded that immunological sensitization produced a non-specific hyperresponsiveness in trachea and lung parenchymal strips. Conversely, cyclooxygenase inhibition by indomethacin elicited a selective increase in the responsiveness to certain agonists in central but not in the peripheral airways.

The relaxant effects of cromakalim (BRL 34915) on human isolated airway smooth muscle.

Cromakalim (BRL 34915) is a potassium channel opener with therapeutic potential as a bronchodilator in asthma. Cromakalim (0.1-30 mumol/l) inhibited the spontaneous tone of human isolated bronchi in a concentration-related manner being nearly as effective as isoprenaline or theophylline. The order of relaxant potencies (expressed as -log10 IC50 mol/l; mean +/- SEM) was isoprenaline (7.29 +/- 0.27; n = 8) > cromakalim (5.89 +/- 0.12; n = 7) > theophylline (4.07 +/- 0.13; n = 10). In human bronchi where tone had been raised by addition of histamine (0.1 mmol/l), acetylcholine (0.1 mmol/l) or leukotriene D4 (LTD4, 0.1 mumol/l), the relaxant effect of cromakalim was substantially reduced. Cromakalim suppressed the contraction produced by KCl (25 mmol/l) but not that produced by KCl (120 mmol/l). Tetraethylammonium (8 mmol/l) was without effect against the relaxant action of cromakalim but procaine (0.5-5 mmol/l) and glibenclamide (0.3 mumol/l) antagonised it. Cromakalim (10 mumol/l) produced an upward displacement of concentration-effect curves for KCl (1-100 mmol/l), acetylcholine (1 nmol/l-1 mmol/l) and histamine (1 nmol/l-1 mmol/l) but it did not alter the concentration-effect curve for LTD4 (0.1 nmol/l-0.1 mumol/l). When tissues were challenged in the presence of cromakalim (10 mumol/l) with KCl (100 mmol/l), acetylcholine (1 mmol/l) or histamine (1 mmol/l), an enhanced contraction was observed compared to control tissues. This enhancement by cromakalim was absent when tissues were challenged with acetylcholine or histamine in either a Ca(2+)-free medium (plus EGTA 0.1 mmol/l) or in the presence of verapamil (10 mumol/l).(ABSTRACT TRUNCATED AT 250 WORDS)