Role of Na(+)-K(+)-ATPase in sodium nitroprusside-induced relaxation of pulmonary artery under hypoxia.

BACKGROUND: The sodium pump (Na(+)-K(+)-ATPase) plays a part in the regulation of smooth muscle contractility, and alterations of enzyme activity by hypoxia could contribute to the mechanism of hypoxic pulmonary vasoconstriction. OBJECTIVE: To determine the role of Na(+)-K(+)-ATPase in the sodium nitroprusside (SNP)-induced relaxation of pulmonary artery in hypoxia. METHODS: Using isolated canine pulmonary arterial rings, we measured the relaxant responses of KCI-contracted tissues to SNP under hyperoxic (95% O2, 5% O2) and hypoxic conditions (5% O2, 5% CO2, 90% N2 in vitro. Na(+)-K(+)-ATPase activity was assessed by measuring ouabain-sensitive (86)Rb uptake. RESULTS: The SNP-induced relaxation was reduced under hypoxia, so that the maximal relaxation decreased from 80.1 +/- 8.6 to 57.8 +/- 6.8% (p < 0.01) and the concentration of SNP required to produce 50% relaxation increased from 1.9 +/- 0.4 x 10(-6) to 2.6 +/- 0.6 x 10(-5) M (p < 0.01). Addition of ouabain, an Na(+)-K(+)-ATPase inhibitor, attenuated the relaxant response to SNP and this inhibition was still observed under hypoxia. Incubation of endothelium-denuded rings with SNP caused dose-dependent increases in intracellular cGMP levels and ouabain-sensitive (86)Rb uptake, and these effects were not significantly altered by hypoxia. CONCLUSION: These results suggest that sarcolemmal Na(+)-K(+)-ATPase activity may be implicated in the mechanism of nitrovasodilator-induced vasodilation of pulmonary artery and may still be functioning under hypoxia.

Ginsenoside-induced relaxation of human bronchial smooth muscle via release of nitric oxide.

Ginsenoside, an extract of Panax ginseng, is an essential constituent of anti-asthmatic Chinese herbal medicine. To elucidate whether ginsenoside affects airway smooth muscle tone and, if so, what the mechanism of action is, we studied relaxant responses of human bronchial strips under isometric condition in vitro, and directly measured the release of nitric oxide (NO) by an amperometric sensor for this molecule. Addition of ginsenoside relaxed the tissues precontracted with acetylcholine in a dose-dependent manner, the maximal relaxation and the ginsenoside concentration required to produce 50% relaxation being 67+/-8% and 210+/-29 microg ml(-1), respectively. The relaxant responses to ginsenoside were inhibited by N(G)-nitro-L-arginine methylester (L-NAME) and removal of the epithelium, but not by N(G)-nitro-D-arginine methylester (D-NAME) or tetrodotoxin. This inhibitory effect of L-NAME was reversed by L-arginine but not by D-arginine. Addition of ginsenoside to the medium containing bronchial tissues dose-dependently increased NO-selective electrical current, and this effect was greatly attenuated by the epithelial removal or Ca(2+)-free medium. Ginsenoside also increased tissue cyclic GMP contents, an effect that was abolished in the presence of L-NAME. It is concluded that ginsenoside induces relaxation of human bronchial smooth muscle via stimulation of NO generation predominantly from airway epithelium and cyclic GMP synthesis. This action might account for the anti-asthmatic effect of Panax ginseng.

Effect of inhaled indomethacin in asthmatic patients taking high doses of inhaled corticosteroids.

BACKGROUND: Cyclooxygenase products of arachidonic acid may play a part in bronchoconstriction and airway inflammation in asthma. OBJECTIVE: We sought to determine the effect of inhaled indomethacin on asthma control and asthma exacerbations during reduction of inhaled corticosteroids in patients with moderate-to-severe steroid-dependent asthma. METHODS: We conducted a double-blind, randomized, parallel-group, multicenter study in 38 patients with asthma taking high doses (> or =1500 microg/d) of beclomethasone dipropionate (BDP). After a run-in period, patients were assigned inhaled indomethacin (50 mg/d) or placebo for 6 weeks, during which the daily doses of BDP were reduced to half at week 2 and then to one third of the baseline dose at week 4. RESULTS: Data were available from 34 patients. After the reduction of BDP doses, FEV(1), peak expiratory flow, asthma symptoms, and exhaled nitric oxide concentrations deteriorated in both treatment groups, but these effects were less pronounced in the indomethacin group compared with the placebo group. During the 6-week treatment period, 89% of the patients receiving placebo had relapse of asthma, whereas only 38% of those receiving inhaled indomethacin did so (P =.003). CONCLUSION: Inhalation of indomethacin can reduce asthma exacerbations induced by reduction of high-dose inhaled corticosteroid in steroid-dependent asthma.

Inhibition of airway smooth muscle tone by Chinese herbal medicines.

The aim of the present study was to elucidate whether Chinese traditional herbal drugs, Gorei-San (TJ-17) and Toki-Shakuyaku-San (TJ-23), affect airway smooth muscle tone and, if so, to determine what the mechanism of action is. Rabbit tracheal segments were isolated and the contractile responses to electrical field stimulation and acetylcholine were measured before and after the application of TJ-17 or TJ-23 under isometric conditions in vitro. Ouabain-sensitive rubidium-86 (86Rb) uptake by tissues in response to each drug was also measured. Each herbal medicine attenuated the contractile responses to electrical field stimulation and acetylcholine in a concentration-dependent manner, the maximal inhibition of acetylcholine-induced contraction being 37.5+/-4.9% for TJ-17 and 42.4+/-5.3% for TJ-23 (p<0.05 for each). These effects were not altered by mechanical removal of the epithelium, indomethacin, the nitric oxide synthase inhibitor NG -nitro-L-arginine methyl ester, the cyclic adenosine monophosphate (cAMP)-dependent protein kinase inhibitor adenosine 3'5'-cyclic monophosphorothioate (Rp-cAMPS), the cyclic guanosine monophosphate (cGMP)-dependent protein kinase inhibitor KT5823, or the calcium (Ca2+)-activated potassium (K+) channel inhibitor charybdotoxin, but were greatly inhibited in the presence of the sodium (Na+)-K+ adenosine triphosphatase (ATPase) inhibitor ouabain. Incubation of tissues with TJ-17 and TJ-23 dose dependently increased ouabain-sensitive 86Rb uptake. The results of the study suggest that both Gorei-San and Toki-Shakuyaku-San reduce airway smooth muscle tone via a postjunctional mechanism probably through stimulation of the sodium pump and the subsequent hyperpolarization/repolarization of the cell membrane. These effects may contribute to the antiasthmatic properties of these herbal medicines.

Repetitive use of a phosphate-binding module in DNA polymerase beta, Oct-1 POU domain and phage repressors.

Motifs for sequence specific-protein-DNA interactions, such as helix-turn-helix, zinc finger and leucine zipper, are now better understood as a result of extensive studies of three-dimensional (3D) structures of transcription factors. On the other hand, little attention has been paid to motifs for sequence nonspecific binding, namely DNA-phosphate binding. To address the question whether different transcription factors and DNA manipulation enzymes, that is enzymes that work on DNA, share a similar mode of phosphate binding, we surveyed interactions between DNA and protein module, a structural unit of a globular protein. We analyzed the modular organization of DNA polymerase beta and found that residues making contact with DNA phosphates were localized to five modules. Structural comparison of these phosphate-binding modules against others in transcription factors and DNA manipulation enzymes revealed that DNA polymerase beta, the Oct-1 POU domain, 434 Cro and the Arc repressor have a phosphate-binding module with 3D structures similar to one another. This newly detected module, the phosphate-binding helix-turn-helix (pbHTH) module, named for its function and 3D structure, interacts with DNA by (i) making hydrogen bonds between a DNA phosphodiester oxygen and an amino hydrogen of the main chain located at the N-terminus of a C-terminal alpha-helix, and (ii) making electrostatic interactions between DNA phosphates and side chains of lysine or arginine. Finding structurally and functionally similar phosphate-binding units in different transcription factors and DNA manipulation enzymes suggests that shuffling of modules is not limited to the DNA base-recognition motif. Phosphate-binding modules are apparently also shuffled in DNA-binding proteins.

[A case of pulmonary eosinophilic granuloma undergoing spontaneous remission].

A case of pulmonary eosinophilic granuloma that underwent spontaneous remission is presented. A 23-year old man presented with dry cough and fever. Chest X-ray film revealed diffuse reticulo-nodular infiltrates in the middle and upper lung fields. Chest CT and HRCT showed multiple cystic lesions with thick walls and small nodules predominantly in the inner zone. Based on radiographic findings, pulmonary eosinophilic granuloma was suspected. Bronchoalveolar lavage cell data showed lymphocyte and eosinophil alveolitis with no increase of CD 1 lymphocytes. The symptoms and radiographic findings improved markedly within 4 months after the onset of symptoms without treatment and upon cessation of smoking. Chest CT and HRCT showed that the cystic walls were thinner and that the small nodules had decreased. Thoracoscopic lung biopsy revealed granulomatous lesions consisting of CD 1 and S-100 protein positive histiocytes with infiltration of eosinophils and fibrous lesions. Pulmonary eosinophilic granuloma was diagnosed. There has been no recurrence for 1 year.

Purification and properties of a proline iminopeptidase from apricot seeds.

A proline iminopeptidase was purified about 18,000-fold from apricot seeds (Prunus armeniaca LINN.) by a five-step procedure comprised of extraction from seeds, ammonium sulfate fractionation, DEAE-cellulose chromatography, CM-Sepharose chromatography, and rechromatography on CM-Sepharose. The purified enzyme had a molecular weight of 220,000 by gel filtration and 55,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This indicates that the native enzyme may be composed of four identical subunits. The isoelectric point was 6.2 as determined by gel electrofocusing. The pH optimum for L-proline beta-naphthylamide was between pH 7.5 and 8.0, and the enzyme was stable in the pH 6.5-to-8.0 region and up to 40 degrees C. The enzyme was specific for L-proline beta-naphthylamide among various amino acid beta-naphthylamides, and it also hydrolzyed L-prolylglycine and L-prolylglycylglycine. The enzyme was strongly inhibited by p-chloromercuribenzoate, 5,5'-dithiobis(2-nitrobenzoic acid), N-ethylmaleimide, and heavy metal ions, but was not activated significantly by thiol compounds. Moreover, the enzyme was inactivated by diethyl pyrocarbonate, p-bromophenacyl bromide, and photooxidation, but was not affected by diisopropyl fluorophosphate, phenylmethanesulfonyl fluoride, bestatin, puromycin, or metal chelating agents. No activation of the enzyme was observed on addition of metal ions. These results suggest that the enzyme is not classifiable as a metalloenzyme, and that cysteine and histidine residues may participate in the enzyme activity.