重组人白细胞介素-11对肺腺癌A549 细胞增殖、 迁移和侵袭的影响

摘要:目的 观察重组人白细胞介素-11 （rhIL-11） 对肺腺癌 A549 细胞增殖、 迁移和侵袭的影响, 并初步探讨其影响机制。方法 分别以 0、 10、 20、 50、 100 μg/L 终浓度的 rhIL-11 作用于 A549 细胞, MTT 法测定 A549 细胞增殖程度; 用 0、 10、 40、 80 μg/L 终浓度的 rhIL-11 作用于 A549 细胞, 划痕实验和 Transwell 侵袭实验测定 A549 细胞的迁移和侵袭能力; Western blot 检测基质金属蛋白酶 （MMP） -2 和 MMP-9 蛋白水平的表达。结果 rhIL-11 对 A549 细胞增殖能力无影响; rhIL-11 作用于 A549 细胞后, 细胞的迁移和侵袭能力显著增强; rhIL-11 能显著上调 MMP-2 和 MMP-9 的表达, 且表达随 rhIL-11 浓度的增高而增高 （P < 0.05）。结论 rhIL-11 能够促进 A549 细胞的迁移和侵袭, 促进 MMP-2、 MMP-9 上调为其可能的机制之一。     

Are Changes in Breathing Pattern on Exposure to Ozone Related to Changes in Pulmonary Surfactant?

Abstract

Rats respond to the inhalation of ozone with changes in breathing pattern during the exposure and the development of a pulmonary inflammatory response 24–48 h postexposure. We report experiments designed to investigate the relationships between changes in breathing pattern and the composition and surface tension-reducing properties of pulmonary surfactant immediately after the exposure. A total of 64 male Fischer 344 rats were exposed to 0.8 ppm O₃ for 4 h in 4 replicate exposures with matched purified air control exposures and 8 rats per exposure group. Those exposed to O₃ developed the rapid-shallow breathing pattern characteristic of oxidant pulmonary irritation during exposure. The rats were sacrificed immediately following the exposure, and pulmonary surfactant was isolated from samples of bronchoalveolar lavage fluid pooled from groups of eight rats. After esterification, the fatty acid methyl ester composition was measured using GC-MS. in the ozone-exposed animals, the decreases in unsaturated species (linoleic, oleic, palmitoleic, and an unidentified fatty acid) relative to the major saturated component, palmitic acid, were highly statistically significant, while stearic acid showed no significant change. Total protein in the lavage fluids of the exposed animals was not elevated, indicating that sacrifice and analysis were performed early in the O₃ injury-inflammation response sequence, and suggesting that the fatty acid changes in the pulmonary surfactant may be due in part to a direct reaction with inhaled O₃. The group mean change in breath frequency (as percent of matched purified air control values) was significantly correlated with the percent change in lnoleic acid fraction among replicate exposures, suggestive of a possible relationship between ozone-induced changes in pulmonary surfactant and changes in breathing patterns. There was no significant change in the surface pressure-area isotherms of monolayers of pulmonary surfactant upon ozone exposure. However, comparison to isotherms from an in vitro exposure of a synthetic mixture of saturated and unsaturated phospholipids suggests that changes due to the observed change in the fatty acid composition in the in vivo experiments may be too small to be observed. Furthermore, the pulmonary surfactant isolation procedure was specifically designed to recover the undamaged surfactant and may discriminate against products of reaction with ozone; hence the isotherms may not necessarily reflect the actual changes during the exposure. Further experiments to elucidate the interaction of inhaled O₃ with pulmonary surfactant and its relationship to changes in breathing pattern are discussed.     

P2-purinoceptors regulate surfactant secretion from rat isolated alveolar type II cells.

Rat isolated alveolar Type II cells were utilized to examine the effect of purine and pyrimidine analogues on secretion of pulmonary surfactant. ATP potently stimulated [3H]-phosphatidylcholine ([3H]-PC) secretion in a time- and dose-dependent manner. The effect of ATP was noted by one hour of exposure and persisted for three hours. The EC50 (concentration producing 1/2 the maximal response) for ATP-induced [3H]-PC secretion was 100 nM. ADP was also a potent secretagogue for surfactant secretion, but AMP and adenosine had no significant effect on surfactant secretion at concentrations less than or equal to 250 microM. The EC50 for ADP-induced [3H]-PC secretion was 250 nM. Other purine and pyrimidine nucleotides (ITP, GTP, CTP, TTP) were examined for their effect on [3H]-PC secretion. All purine and pyrimidine triphosphates examined significantly augmented [3H]-PC secretion, but were much less potent than ATP. The EC50s were ITP = 10 microM; GTP = 100 microM; CTP = 250 microM; TTP = 100 microM. Neither 8-phenyltheophylline (10 microM, a P1-purinoceptor antagonist), propranolol (100 microM, a beta-adrenoceptor antagonist), nor indomethacin (10 microM, a prostaglandin synthetase inhibitor) inhibited ATP-induced [3H]-PC secretion from isolated Type II cells. These data provide evidence for regulation of surfactant secretion from alveolar Type II cells by a P2-purinoceptor.     

肺表面活性物质预防早产儿呼吸窘迫综合征的疗效观察

目的:探讨早产儿生后6 h内使用牛肺表面活性剂(PS)预防新生儿呼吸窘迫综合征(NRDS)的疗效。方法:我院出生的76例胎龄28～32周的早产儿随机分为两组,预防组38例给予气管内注入40～100 mg/kg牛肺表面活性剂与未用PS的对照组38例进行前瞻性临床研究,比较两组NRDS的发生情况。结果:预防组发生NRDS 5例,对照组发生NRDS 17例,两组比较差异有统计学意义(P<0.05)。预防组吸氧天数(9.2±6.1)d、呼吸机使用天数(6.5±4.9)d及住院天数(29.3±7.8)d,较对照组(16.7±6.4)d、(9.8±7.0)d、(37.4±7.2)d明显缩短,差异有统计学意义(P<0.05)。预防组肺部并发症如肺部感染及支气管肺发育不良较对照组明显减少,两组比较差异有统计学意义(P<0.05)。结论:早产儿预防性使用牛肺表面活性剂能减少NRDS的发生率,减少呼吸机的使用,减少肺部并发症发生,改善早产儿预后。     

Possible involvement of long chain fatty acids in the spores of Ganoderma lucidum (Reishi Houshi) to its anti-tumor activity

During our isolation of biologically active substances from the spores of Ganoderma lucidum (Reishi Houshi, G. lucidum) guided by the inhibitory activity on HL-60 cell proliferation, NMR spectroscopic and mass spectrometric data indicate the substance contains a mixture of several long chain fatty acids. Hence, in this study, we have examined the inhibitory effects of an ethanolic extract of the spores of G. lucidum as the spore extract, on the proliferation of various human cancer cell lines by comparison with several authentic long chain fatty acids. Of the fatty acids we examined nonadecanoic acid (C19:0) showed the highest inhibitory activity for HL-60 cell proliferation with IC(50) values of 68+/-7 microM followed by heptadecanoic acid (C17:0, 120+/-23 microM), octa- (C18:0, 127+/-4 microM) and hexadecanoic acids (C16:0, 132+/-25 microM), respectively. The corresponding unsaturated fatty acids containing one double bond such as cis-10-nonadecenoic acid (C19:1), cis-9-octadecenoic acid (C18:1), cis-10-heptadecenoic acid (C17:1) and cis-9-hexadecenoic acid (C16:1) were less effective. The ethanolic extract of spores of G. lucidum were shown by annexin-V FITC/PI double staining to induce apoptosis of HL-60 cells in a similar way to cis-10-nonadecenoic acid (C19:1). These unsaturated fatty acids probably inhibit tumor necrosis factor production induced by lipopolysaccharide in a mouse macrophage preparation. Our results suggest the spores of G. lucidum contain 19-carbon fatty acids as one of the components for characteristics of its physiological effects.     

Phosphatidylcholine synthesis in isolated type II pneumocytes from ozone-exposed rats

Phosphatidylcholine (PC) synthesis by alveolar type II cells, as an indicator for the production of pulmonary surfactant, was studied after a 4-h exposure of rats to 4 mg ozone/m³ (2 ppm). Lung ravage fluid analysis after exposure revealed significant increases in proteins, which is indicative for pulmonary injury. When type II cells were isolated immediately and thereafter cultured for 20 h, the rate of PC synthesis in cells derived from ozone-exposed rats was not significantly different from that in cells from unexposed controls. Yet, a decreased rate of PC synthesis was observed when these cells were subsequently exposed to ozone in vitro. The activity of the enzyme glycerolphosphate acyltransferase (GPAT) was slightly enhanced in cultured type II cells isolated from ozone-exposed rats, while the lysophosphatidylcholine acyltransferase (LPCAT) activity was unchanged. However, ozone exposure of rats did result in a significant decrease of PC synthesis when measured in freshly prepared type II cell suspensions, although both GPAT and LPCAT activities were not affected. It is concluded that a decrease in pulmonary surfactant related PC synthesis after ozone exposure of rats can be demonstrated in freshly isolated type II pneumocytes. Cultured type II cells from exposed rats lack this effect and are therefore less useful to study changes in phospholipid biosynthesis after in vivo ozone exposure. The data on in vitro ozone exposure of cultured type II cells, however, support the view that ozone may impair pulmonary surfactant production.Abbreviations PC phosphatidylcholines - GPAT glycerolphosphate acyltransferase - LPCAT lysophosphatidylcholine acyltransferase     

THERMAL DECOMPOSITION OF PHOSPHOLIPID SECONDARY OZONIDES: IMPLICATIONS FOR THE TOXICITY OF INHALED OZONE

While inhalation of ozone is known to cause a variety of health effects, the reactions at a molecular level that lead to these effects are not well understood. One potential path is the reaction of ozone with the unsaturated fatty acid components of pulmonary surfactant at the air-water interface in the lung to form secondary ozonides. These have been proposed to decompose to free radicals, which can then initiate the well-known inflammatory response. We report here the first kinetic studies of the thermal decomposition of the cis and trans secondary ozonides of 1-palmitoyl-2-oleoyl- sn -glycero-3-phosphocholine (POPC), a phospholipid found in significant quantities in lung surfactant. The ozonides were synthesized by reaction of O3 with POPC adsorbed on a glass surface, and their thermal decomposition kinetics were followed using high-performance liquid chromatography (HPLC) over the tem perature range from 50 C to 106 C in either methanol or 1,1,1,2-tetrachloroethane. The Arrhenius parameters for the thermal decomposition in methanol are A = 10 8.7+/-0.3 s-1 and E = 19.6 +/- 0.6 kcal mol-1 for the a cis ozonide, and A = 10 8.7+/-0.6 s-1 and Ea = 19.8 +/- 1.0 kcal mol-1 for the trans ozonide. In 1,1,1,2-tetrachloroethane, the parameters are A = 10 8.3+/-2.1 s-1 and Ea = 18.4 +/- 3.4 kcal mol-1 for the cis ozonide, and A = 10 9.3+/-3.2 s-1 and Ea = 20.2 +/- 5.2 kcal mol-1 for the trans ozonide (all errors cited are +/-2). Within experimental error, there is no difference in the kinetics of decomposition in the two solvents. However, both the activation energy and the preexponential factor for the decomposition of the phospholipid ozonides are significantly lower than those for decomposition of the long-chain alkene ozonide 1-octene ozonide, determined to be Ea = 26.7 +/- 3.2 kcal mol-1 and A = 10 12.7+/-1.9 s-1. The latter reaction has been proposed to be initiated by scission of the O-O bond, followed by decomposition of the peroxy biradical to generate free radicals. The kinetics for the decomposition of the POPC ozonides in solution are similar to those of simple alkene ozonides in the gas phase, where a concerted mechanism involving simultaneous intramolecular hydrogen transfer and O-O bond cleavage has been proposed. The only high-molecular-weight major product of the POPC ozonide decomposition identified using liquid secondary ion mass spectrometry (LSIMS) was the lipid acid 1-palmitoyl-2-azelaoyl- sn -glycero-3-phosphocholine, which was observed as a product in both solvents. The mechanism and implications for the toxicology of inhaled ozone are discussed.     

Baicalin is anti-inflammatory in cigarette smoke-induced inflammatory models in vivo and in vitro: A possible role for HDAC2 activity

Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory disease characterized by airway obstruction and progressive lung inflammation, which is insensitive to corticosteroids therapies. In this study, we investigated the mechanism underlying the attenuation of cigarette smoke (CS)-induced respiratory inflammation by baicalin, a flavonoid compound isolated from the root of Scutellaria baicalensis Georgi, in vivo and in vitro. In vivo, mice were exposed to smoke of 15 cigarettes for 1 h/day, 6 days/week for 3 months and dosed with baicalin (25, 50 and 100mg/kg) or dexamethasone (1mg/kg). In vitro, A549 cells were incubated with baicalin (10, 50 and 100 μM) or dexamethasone (10(-12), 10(-10), 10(-8) and 10(-6)M) followed by treatments with cigarette smoke extract (CSE, 2.5 and 5%), or TNF-α (10 ng/ml), or trichostatin A (TSA, 100 ng/ml). We found that baicalin significantly protected pulmonary function and attenuated CS-induced inflammatory response by decreasing inflammatory cells and production of TNF-α, IL-8 and MMP-9. This result was not found in the group treated with dexamethasone. Baicalin also showed efficacy in enhancing histone deacetylase (HDAC)2 activity and protein expression, however, it did not affect HDAC2 mRNA. Further studies revealed that baicalin inhibited HDAC2 phosphorylation, suggesting that it may directly affect the protein structure and effect by modification at post-translational level. Together these results suggest that baicalin has anti-inflammatory effects in cigarette smoke induced inflammatory models in mice and A549 cells, possibly achieved by modulating HDAC2.     

Similar binding of 3H-ADTN and 3H-apomorphine to calf brain dopamine receptors.

The binding of 3H(+/-)-ADTN (of high specific activity; 7.6 Ci/mmole) to homogenates of calf striatum was investigated. The dissociation constant (KD) for the specific, saturable binding of 3H-(+/-)-ADTN was 1 nM and the density of specific sites was 100 fmoles/mg protein. The IC50 values (nM concentrations inhibiting specific binding by 50%) were 0.9 for (+/-)-N-propyl-norapomorphine, 3.0 for dopamine, 7 for (--)-adrenaline, 60 for (--)-noradrenaline and 4000 for isoproterenol, a series of potencies compatible with properties for a dopaminergic site. The (+)-enantiomer of ADTN was 10 times more potent than (--)-ADTN in competing for 3H-(+/-)-ADTN, while the (--)-enantiomer of 5-OH-dipropyl-ATN was 40 times more potent than the (+)-isomer. The IC50 values for various agonists against 3H-(+/-)-ADTN were similar to those against 3H-apomorphine or 3H-dopamine in the calf striatum. A comparison of these 3H-(+/-)-ADTN data to those for 3H-spiperone suggests that the two 3H-ligands label different receptor sites.     

Formation of secondary ozonides from the reaction of an unsaturated phosphatidylcholine with ozone

Phosphatidylcholines are significant components of pulmonary surfactant in the alveolar region of the lung, where they play a major role in lung function due to their surface tension reducing properties. However, separation and the direct identification of many of the primary products of reaction of phosphatidylcholines with inhaled pollutant gases has not been possible until recently due to the lack of suitable analytical techniques, so that compounds such as fatty acid methyl esters generally have been used as analogues for the phospholipids. We report here the first isolation and identification of the products of reaction of ozone with one of the unsaturated components of lung surfactant, beta-oleoyl-gamma-palmitoyl-L-alpha-phosphatidylcholine (OPPC), using a combination of high-performance liquid chromatography, fast atom bombardment mass spectrometry, and Fourier transform infrared, ultraviolet absorption, and nuclear magnetic resonance spectrometry as well as gas chromatography. The products are shown to be the cis and trans secondary ozonides of the parent phosphatidylcholine, analogous to those previously observed by other researchers in the reactions of the simple fatty acid methyl esters with ozone. This also appears to be the first report of fast atom bombardment mass spectra of these phospholipid secondary ozonides. The implications of this work for the inhalation of ozone, formed in photochemical smog, are discussed.     

Synthesis, pharmacological characterization, and molecular modeling of heterobicyclic amino acids related to (+)-2-aminobicyclo[3.1.0] hexane-2,6-dicarboxylic acid (LY354740): identification of two new potent, selective, and systemically active agonists for group II metabotropic glutamate receptors

As part of our ongoing research program aimed at the identification of highly potent, selective, and systemically active agonists for group II metabotropic glutamate (mGlu) receptors, we have prepared novel heterobicyclic amino acids (-)-2-oxa-4-aminobicyclo[3.1. 0]hexane-4,6-dicarboxylate (LY379268, (-)-9) and (-)-2-thia-4-aminobicyclo[3.1.0]hexane-4,6-dicarboxylate (LY389795, (-)-10). Compounds (-)-9 and (-)-10 are structurally related to our previously described nanomolar potency group II mGlu receptor agonist, (+)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylate monohydrate (LY354740 monohydrate, 5), with the C4-methylene unit of 5 being replaced with either an oxygen atom (as in (-)-9) or a sulfur atom (as in (-)-10). Compounds (-)-9 and (-)-10 potently and stereospecifically displaced specific binding of the mGlu2/3 receptor antagonist ([3H]LY341495) in rat cerebral cortical homogenates, displaying IC50 values of 15 +/- 4 and 8.4 +/- 0.8 nM, respectively, while having no effect up to 100 000 nM on radioligand binding to the glutamate recognition site on NMDA, AMPA, or kainate receptors. Compounds (-)-9 and (-)-10 also potently displaced [3H]LY341495 binding from membranes expressing recombinant human group II mGlu receptor subtypes: (-)-9, Ki = 14.1 +/- 1.4 nM at mGlu2 and 5.8 +/- 0.64 nM at mGlu3; (-)-10, Ki = 40.6 +/- 3.7 nM at mGlu2 and 4.7 +/- 1.2 nM at mGlu3. Evaluation of the functional effects of (-)-9 and (-)-10 on second-messenger responses in nonneuronal cells expressing human mGlu receptor subtypes demonstrated each to be a highly potent agonist for group II mGlu receptors: (-)-9, EC50 = 2.69 +/- 0.26 nM at mGlu2 and 4.58 +/- 0.04 nM at mGlu3; (-)-10, EC50 = 3.91 +/- 0.81 nM at mGlu2 and 7.63 +/- 2. 08 nM at mGlu3. In contrast, neither compound (up to 10 000 nM) displayed either agonist or antagonist activity in cells expressing recombinant human mGlu1a, mGlu5a, mGlu4a, or mGlu7a receptors. The agonist effects of (-)-9 and (-)-10 at group II mGlu receptors were not totally specific, however, as mGlu6 agonist activity was observed at high nanomolar concentrations for (-)-9 (EC50 = 401 +/- 46 nM) and at micromolar concentrations (EC50 = 2 430 +/- 600 nM) for (-)-10; furthermore, each activated mGlu8 receptors at micromolar concentrations (EC50 = 1 690 +/- 130 and 7 340 +/- 2 720 nM, respectively). Intraperitoneal administration of either (-)-9 or (-)-10 in the mouse resulted in a dose-related blockade of limbic seizure activity produced by the nonselective group I/group II mGluR agonist (1S,3R)-ACPD ((-)-9 ED50 = 19 mg/kg, (-)-10 ED50 = 14 mg/kg), indicating that these molecules effectively cross the blood-brain barrier following systemic administration and suppress group I mGluR-mediated limbic excitation. Thus, heterobicyclic amino acids (-)-9 and (-)-10 are novel pharmacological tools useful for exploring the functions of mGlu receptors in vitro and in vivo.     

New selective and potent 5-HT(1B/1D) antagonists: chemistry and pharmacological evaluation of N-piperazinylphenyl biphenylcarboxamides and biphenylsulfonamides

A series of new analogues of N-[4-methoxy-3-(4-methylpiperazin-1-yl)phenyl] 2'-methyl-4'-(5-methyl-1,2,4-oxadiazol-3-yl)biphenyl-4-carboxamide (1; GR127935) as potent and selective 5-HT(1B/1D) antagonists were synthesized and evaluated pharmacologically. Their receptor binding profiles were comparable to that of 1. The 1,3,4-oxadiazole isomer 2 and the 4'-aminocarbonyl and 4'-amidinyl analogues (9 and 10) of 1 had higher affinities at the rat 5-HT(1B) receptor (IC(50) = 0.93, 1. 3, and 0.5 nM, respectively) and calf 5-HT(1D) receptor (IC(50) = 37, 10, and 3 nM, respectively) than did 1 (1.6 and 52 nM for rat 5-HT(1B) and calf 5-HT(1D) receptors, respectively). In the functional in vitro testing of 5-HT(1B/1D) antagonistic properties, 2, 9, 10, 11b (O-demethylated derivative of 2), 13a (O-methylsulfonyl analogue of 2), and 16 (which differs from 2 with a sulfonamide linker) showed more pronounced effects in the K(+)-induced 5-HT release in the cortex of guinea pig than did 1 and 3 (SB224289). Compounds 2, 9, and 10 were equally potent as 1 in rabbit saphenous vein model (pA(2) > 9). A biochemical study of 2 with in vivo microdialysis in the rat brain showed that it is capable of augmenting citalopram (a selective serotonin reuptake inhibitor, SSRI) induced 5-HT release in rat ventral hippocampus, while preventing the decrease in acetylcholine release elicited by citalopram administration. The molecular structure of 2 was determined by single-crystal X-ray analysis. The log P and log D values of these compounds were calculated. This study contributes to the SAR study of N-piperazinylphenyl biphenylcarboxamides as selective and potent 5-HT(1B/1D) antagonists.     

Effects of ambroxol on pulmonary surfactant--analysis of the fatty acid composition of phosphatidylcholine in the sputum and normal respiratory tract fluid in rabbits

The mechanism of action of secretagogic expectorants (ex., ambroxol) has not been clarified. Recently, attention has been directed to the relationship of their action to pulmonary surfactants. In the present study, the fatty acid (FA) composition of phosphatidylcholine (PC), which is the main physiological surfactant, was investigated using sputum, respiratory tract fluid, mucin-like substance, lung washings and lung tissue of rabbits. Effects of ambroxol (20 mg/kg, i.s.) on several parameters such as output volume, FA contents of PC, protein content and viscosity of respiratory tract fluid of rabbits were also investigated. In respiratory tract fluid, lung washings and mucin-like substance of rabbits, saturated FA, especially C16:0, were predominant components of PC; while in sputum and lung tissue of rabbits and respiratory tract fluid of hens, unsaturated FA, especially C18:1 and C18:2, were more predominant components in comparison with those in the above specimens. Ambroxol significantly increased the contents of C16:0, saturated FA and total FA of PC, and it also increased protein content with an increase in the viscosity of respiratory tract fluid. These results suggest that in the respiratory tract fluid of rabbits, PCs are pulmonary surfactants, and the increasing secretion of pulmonary surfactant is likely to be involved in the expectorant action of ambroxol.     

Enzyme-linked immunosorbent assays for kinins using high-affinity monoclonal kinin antibodies

Splenocytes from mice immunized either with bradykinin conjugated with carbodiimide to keyhole limpet hemocyanin or ovalbumin were fused using polyethylene glycol with the mouse myeloma cell line SP2/o. Nine monoclonal antibodies reactive with kinins were obtained from two fusions. All of the antibodies were of the IgG1k isotype, except for one, which was an IgG2ak. Based on their reactivities with biologically active kinins and biologically inactive degradation products, the antibodies were separated into three groups. The first group, which had the highest affinities for bradykinin, displayed about equal reactivities for bradykinin and des-Arg9-bradykinin, but little reactivities for the kinin fragments, des-Arg1-bradykinin and des-Phe8-Arg9-bradykinin, or for lysyl-bradykinin and methionyl-lysyl-bradykinin. The second group was similar to the first except that it showed about a 2.5- to 3.5-fold greater reactivity for des-Arg9-bradykinin than for bradykinin. The third group, which had the lowest affinities for bradykinin [50% inhibition of antibody binding to an enzyme-linked immunosorbent assay (ELISA) plate occurring with bradykinin concentrations ranging from about 8 to 39 nM], showed little reactivities with des-Arg1-bradykinin, des-Arg9-bradykinin and des-Phe8-Arg9-bradykinin, but 50-100% cross-reactivities with lysyl-bradykinin and methionyl-lysyl-bradykinin. The useful ranges for bradykinin detection (ng/well, 50 microL assay volume) using the highest affinity antibody in each group in ELISAs were: 0.01 to 0.5, 0.03 to 3, and 0.1 to 3 for groups 1, 2, and 3, respectively.     

Relaxation to bradykinin in bovine pulmonary supernumerary arteries can be mediated by both a nitric oxide-dependent and -independent mechanism

1. The aim of the present study was to determine the relative contribution of prostanoids, nitric oxide and K(+) channels in the bradykinin-induced relaxation of bovine pulmonary supernumerary arteries. 2. In endothelium-intact, but not denuded rings, bradykinin produced a concentration-dependent relaxation (pEC(50), 9.6+/-0.1), which was unaffected by the cyclo-oxygenase inhibitor indomethacin. The nitric oxide scavenger hydroxocobalamin (200 micro M, pEC(50), 8.5+/-0.2) and the nitric oxide synthase inhibitor L-NAME (100 micro M, pEC(50), 8.9+/-0.1) and the combination of L-NAME and hydroxocobalamin (pEC(50), 8.1+/-0.2) produced rightward shifts in the bradykinin concentration response curve. 3. The guanylyl cyclase inhibitor ODQ (10 micro M, pEC(50), 9.6+/-0.4) did not affect the response to bradykinin. 4. Elevating the extracellular [K(+)] to 30 mM did not affect the response to bradykinin but abolished the response when ODQ or L-NAME was present. 5. The K(+) channel blocker apamin (100 nM), combined with charybdotoxin (100 nM), produced a small reduction in the maximum response to bradykinin but they abolished the response to bradykinin when ODQ, L-NAME or hydroxocobalamin were present. Apamin (100 nM) combined with iberiotoxin (100 nM) also reduced the response to bradykinin in the presence of hydroxocobalamin or L-NAME. 6. The concentration response curve for sodium nitroprusside-induced relaxation was abolished by ODQ (10 micro M) and shifted to the right by apamin and charybdotoxin. 7. These studies suggest that in bovine pulmonary supernumerary arteries bradykinin can stimulate the formation of nitric oxide and activate an EDHF-like mechanism and that either of these pathways alone can mediate the bradykinin-induced relaxation. In addition nitric oxide, acting through guanylyl cyclase, can activate an apamin/charbydotoxin-sensitive K(+) channel in this tissue.     

N-hydroxy-3-phenyl-2-propenamides as novel inhibitors of human histone deacetylase with in vivo antitumor activity: discovery of (2E)-N-hydroxy-3-[4-[[(2-hydroxyethyl)[2-(1H-indol-3-yl)ethyl]amino]methyl]phenyl]-2-propenamide (NVP-LAQ824)

A series of N-hydroxy-3-phenyl-2-propenamides were prepared as novel inhibitors of human histone deacetylase (HDAC). These compounds were potent enzyme inhibitors, having IC(50)s < 400 nM in a partially purified enzyme assay. However, potency in cell growth inhibition assays ranged over 2 orders of magnitude in two human carcinoma cell lines. Selected compounds having cellular IC(50) < 750 nM were tested for maximum tolerated dose (MTD) and for efficacy in the HCT116 human colon tumor xenograft assay. Four compounds having an MTD > or = 100 mg/kg were selected for dose-response studies in the HCT116 xenograft model. One compound, 9 (NVP-LAQ824), had significant dose-related activity in the HCT116 colon and A549 lung tumor models, high MTD, and low gross toxicity. On the basis, in part, of these properties, 9 has entered human clinical trials in 2002.     

Synthesis of a 6-methyl-7-deaza analogue of adenosine that potently inhibits replication of polio and dengue viruses

Bioisosteric deaza analogues of 6-methyl-9-β-D-ribofuranosylpurine, a hydrophobic analogue of adenosine, were synthesized and evaluated for antiviral activity. Whereas the 1-deaza and 3-deaza analogues were essentially inactive in plaque assays of infectivity, a novel 7-deaza-6-methyl-9-β-D-ribofuranosylpurine analogue, structurally related to the natural product tubercidin, potently inhibited replication of poliovirus (PV) in HeLa cells (IC(50) = 11 nM) and dengue virus (DENV) in Vero cells (IC(50) = 62 nM). Selectivity against PV over cytotoxic effects to HeLa cells was >100-fold after incubation for 7 h. Mechanistic studies of the 5'-triphosphate of 7-deaza-6-methyl-9-β-D-ribofuranosylpurine revealed that this compound is an efficient substrate of PV RNA-dependent RNA polymerase (RdRP) and is incorporated into RNA mimicking both ATP and GTP.     

Acute ozone-induced lung injury in rats: structural-functional relationships of developing alveolar edema.

As part of a study on the effects of acute ozone stress on the lung surfactant system, we correlated morphometric, biochemical, and functional indices of lung injury using male rats exposed to 3 ppm ozone for 1, 2, 4, and 8 hr. Evaluation of lung mechanics, using the Pulmonary Evaluation and Diagnostic Laboratory System, revealed a significant decrease in dynamic lung compliance (ml/cmH2O/kg) from a control value of 0.84 +/- 0.02 (SEM) to 0.72 +/- 0.04 and 0.57 +/- 0.06 at 4 and 8 hr, respectively. At 2 hr there was a transient increase in PaO2 to 116 torr (control = 92 torr) followed by a decrease at 4 hr (65 torr) and 8 hr (55 torr). Morphometry of lung tissue, fixed by perfusion of fixative via the pulmonary artery at 12 cm H2O airway distending pressure, demonstrated an increase in the area of the intravascular compartment at 8 hr, in association with a 65 and 39% replacement of the alveolar area by fluid in ventral and dorsal lung regions, respectively. There was a positive correlation (r = 0.966) between alveolar edema and transudated proteins in lavage fluid. A stepwise multiple regression model, with edema as the dependent variable, suggested that pulmonary vasodilatation, hypoxemia, and depletion of surfactant tubular myelin in lavage fluid were indices for predicting alveolar edema. In a second model, with lavage protein concentration as the dependent variable, decreasing dynamic compliance and hypoxemia were predictors of progressive, intraalveolar transudation of plasma proteins. The above structural-functional relationships support the concept that ozone-induced high-protein alveolar edema is pathogenetically linked to pulmonary hyperemia, deficiency of surfactant tubular myelin, and associated lung dysfunctions.     

非离子表面活性剂逆转肿瘤细胞多药耐药性的机制

目的探讨非离子表面活性剂聚氧乙烯醚﹑聚乙二醇300﹑曲拉通X100对肺癌耐药细胞A549/DDP(耐顺铂)多药耐药性的逆转及其机制。方法用四氮唑盐(MTT)法进行体外耐药逆转实验,Westernblot检测P-gp(P-糖蛋白)蛋白的表达。结果①合用非离子型表活剂前后,A549/DDP对化疗药物阿霉素、顺铂、丝裂霉素、5-氟尿嘧啶、依托泊甙和长春新碱的敏感性显著增加(各组都为P<0.01);②三种非离子型表面活性剂能抑制P-gp蛋白的表达。结论三种非离子表面活性剂具有逆转A549/DDP耐药性的作用,逆转机制与其抑制P-gp蛋白的表达有关。