Epigallocatechin-3-gallate inhibits growth of activated hepatic stellate cells by enhancing the capacity of glutathione synthesis

Activation of hepatic stellate cells (HSC), the key effectors in hepatic fibrogenesis, is characterized by enhanced cell proliferation and overproduction of extracellular matrix. Oxidative stress promotes HSC activation. Glutathione (GSH) is the most important intracellular antioxidant, whose synthesis is mainly regulated by glutamate-cysteine ligase (GCL). We reported previously that (-)-epigallocatechin-3-gallate (EGCG), the major and most active component in green tea extracts, inhibited HSC activation. The aim of this study is to elucidate the underlying mechanisms. We hypothesize that this inhibitory effect of EGCG might mainly result from its antioxidant capability by increasing de novo synthesis of GSH. In this report, we observe that EGCG enhances the levels of cytoplasmic and mitochondrial GSH and increases GCL activity by inducing gene expression of the catalytic subunit GCLc, leading to de novo synthesis of GSH. Real-time polymerase chain reaction and Western blotting analyses show that de novo synthesis of GSH is required for EGCG to regulate the expression of genes relevant to apoptosis and to cell proliferation. Additional experiments demonstrate that exogenous transforming growth factor (TGF)-beta1 suppresses GCLc gene expression and reduces the level of GSH in cultured HSC. Transient transfection assays and Western blotting analyses further display that EGCG interrupts TGF-beta signaling by reducing gene expression of TGF-beta receptors and Smad4, leading to increased expression of GCLc. These results support our hypothesis and collectively demonstrate that EGCG increases the level of cellular GSH in HSC by stimulating gene expression of GCLc, leading to the inhibition of cell proliferation of activated HSC in vitro.     

HA autoreceptor assay with superfused slices of rat brain cortex and electrical stimulation

Slices of rat brain cortex previously loaded with [3H]histamine ([3H]HA) via de novo synthesis from [3H]histidine released tritiated histamine ([3H]HA) Ca2+ dependently in a superfused system. Both electrical field stimulation and high levels of K+ ions elicited this release. The extent of release depended on stimulation intensity. Rather strong stimuli, either by high frequency or longer stimulation, were required to elicit sufficient HA release for proper assessment of the concentration-dependence of release inhibition by drugs. The system showed marked depletion (less response per pulse) upon long-continued or successive stimulations. HA added to the superfusion medium inhibited the release evoked by stimulation at frequencies up to 10 Hz or with 30 mM K+ but not the release at higher frequencies or with 45 mM K+. The inhibition was mediated by H3 receptors, was concentration-dependent (pD2 = 7.4) and was complete at 10(-6) M. The H2 agonist impromidine antagonized the inhibition competitively (pA2 = 7.1). It is concluded that this assay in a superfusion system with electrical stimulation is suitable for the assessment of H3 receptor activity of drugs.     

Interleukin-1 alpha-stimulated fibroblast eicosanoid synthesis is not mediated by interleukin-6.

Adherent human dermal fibroblasts secreted interleukin-6, prostaglandin E2, prostaglandin I2 and 15-hydroxyeicosatetraenoic acid (assayed by radioimmunoassay) during a 3 h incubation period. Although human dermal fibroblasts did not secrete interleukin-1 alpha or interleukin-1 beta, human recombinant interleukin-1 alpha stimulated arachidonic acid metabolism and interleukin-6 synthesis. This effect was, at least partly, dependent on de novo protein synthesis. In contrast, human recombinant interleukin-6 had no effect on the synthesis and release of the eicosanoids measured. Human recombinant interleukin-1 alpha also stimulated the metabolism of [14C]arachidonic acid, but only if fibroblast were pre-incubated with the cytokine for three hours. Our data indicate that (a) fibroblasts secrete interleukin-6 but not interleukin-1, (b) interleukin-1 alpha, but not interleukin-6, stimulates fibroblast arachidonic acid metabolism and (c) the mechanisms involved in the metabolism of endogenous arachidonic acid are more sensitive to human recombinant interleukin-1 alpha than those involved in metabolism of the exogenous substrate.     

Effect of 1-methyl-2-nitrosoimidazole on intracellular thiols and calcium levels in Chinese hamster ovary cells

The cellular reduction of 2-nitroimidazoles under hypoxic conditions can lead to cell killing. One of the postulated toxic intermediates is the two-electron reduction product, the nitrosoimidazole. 1-Methyl-2-nitrosoimidazole (INO) was used as a model to study the reactivity of 2-nitrosoimidazoles with sulfhydryls. INO reacted within minutes with bovine serum albumin (BSA) in a stoichiometric fashion as measured by the loss of its characteristic absorption at 360 nm. It appeared to react specifically with the SH group of BSA as demonstrated by the loss of 5,5'-dithiobis-2- nitrobenzoic acid (DTNB) reactive groups and by the loss of INO reactivity if BSA was previously reacted with DTNB. INO also depleted glutathione (GSH) and protein sulfhydryls (Pr-SH) in Chinese hamster ovary (CHO) cells in a concentration-dependent fashion. INO at 25 microM, a non-toxic concentration in terms of cell colony-forming ability, depleted GSH to 10-20% of control levels within 5 min after treatment. Pr-SH were depleted more slowly to 60% of control levels. GSH recovered to near control levels over 3-4 hr but Pr-SH remained depressed. The recovery of GSH was blocked by buthionine sulfoximine (BSO), suggesting that the recovery was due to de novo synthesis of GSH. At a toxic concentration of INO (45 microM), GSH was again depleted to 10-20% and Pr-SH to 50% of control levels. No recovery of either was observed up to 4 hr. The effect of this extensive oxidative stress on intracellular calcium (Ca2+i) levels was monitored using 1-[2-amino-5-(6-carboxyindole-2-yl)-phenoxyl]-2- (2'-amino-5'-methylphenoxy)-ethane-N,N,N',N'-tetraacetic acid pentaacetoxy methylester (INDO-1 AM). At toxic concentrations of INO, Ca2+i increased in a sustained, non-physiological manner starting at approximately 60 min after the addition of INO. No increase in Ca2+i was observed when cells were treated with nontoxic concentrations of INO. INO toxicity may be modulated by an uncontrolled influx of Ca2+ which can trigger the activation of cellular enzymes and lead to cell death.     

Tissue-specific alterations of de novo fatty acid synthesis in 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-treated rats

De novo fatty acid synthesis was determined by the³H₂O method in numerous tissues and organs of TCDD-treated (125 g/kg), pair-fed and free-fed male Sprague-Dawley rats to investigate if this important pathway of intermediary metabolism is altered by TCDD. Of the 12 tissues and organs examined, liver showed an increased, and interscapular brown adipose tissue (IBAT) a decreased de novo fatty acid synthesis when comparing TCDD-treated to pair-fed or free-fed control rats. De novo fatty acid synthesis was unaffected in other organs and tissues examined, with the exception that the concentration of³H-fatty acids in plasma reflected the increased rate of synthesis seen in the liver of TCDD-treated animals. Increased de novo fatty acid synthesis in liver coincided with increased plasma triiodothyronine (T₃) concentrations, whereas decreased de novo fatty acid synthesis in IBAT parallelled decreased plasma thyroxine (T₄) levels. Thyroidectomy decreased de novo fatty acid synthesis, as expected, in both liver and IBAT. However, TCDD elicited no response in either of these organs in thyroidectomized rats. This finding suggests that changes observed in non-thyroidectomized rats are probably secondary effects. Indeed, known tissue-specific effects of T₃ on liver and T₄ on IBAT provide a likely explanation for the altered de novo fatty acid synthesis of these organs. It is suggested that increased de novo fatty acid synthesis in the liver of TCDD-treated rats might be responsible for the additional wasting away observable in these animals as compared to pair-fed controls.     

Effect of butylated hydroxytoluene on glutathione S-transferase and glutathione peroxidase activities in rat liver

When rats were fed a diet containing 0.4% (w/w) butylated hydroxytoluene (BHT), glutathione (GSH) S-transferase activity towards 1-chloro-2,4-dinitrobenzene (CDNB) increased approximately 3-fold in the liver. Immunotitration studies using the antibodies raised against rat liver GSH S-transferase B and GSH S-transferase A and C indicated that the increase in GSH S-transferase activity was probably due to de novo protein synthesis. Since some forms of rat liver GSH S-transferases express GSH peroxidase II activity, a concomitant increase in GSH peroxidase II was expected. However, GSH peroxidase II activity in the liver of BHT-treated rats remained unchanged. Gel filtration of supernatant fractions from livers of control and BHT-treated rats, followed by isoelectric focusing, indicated that BHT induced the activity of hepatic GSH S-transferases, without any apparent effect on GSH peroxidase II activity.     

Activation of plasma membrane reduced glutathione transport in death receptor apoptosis of HepG2 cells

Cells undergoing apoptosis release reduced glutathione (GSH) into the extracellular space; however, the physiological significance and the mechanism behind the GSH export remain unclear. The present study demonstrates that GSH is released by HepG2 cells undergoing Fas, tumor necrosis factor alpha (TNFalpha), or tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-stimulated cell death. GSH release was observed at times when extracellular lactate dehydrogenase (LDH) activity and propidium iodide (PI) incorporation were low, suggesting that the GSH release does not occur because of nonspecific cell damage, but is occurring through a specific transport system. Caspase 3-like proteases were activated before GSH was released, indicating that protease may be involved in signaling GSH release. To investigate the mechanism of GSH release, studies were performed in the presence of GSH transport inhibitors, as well as 25 mM GSH in the media. Two organic anion transporter inhibitors, probenecid and dibromosulfophthalein (DBSP), were effective in inhibiting Fas-stimulated GSH release. The addition of 25 mM GSH to the extracellular media also prevented the loss of intracellular GSH and delayed cell death. These findings suggest that an organic anion transporter is involved in GSH release during apoptosis, and that maintenance of intracellular GSH levels during apoptosis provides protection for the cell.     

Regulation of gamma-glutamate-cysteine ligase expression by oxidative stress in the mouse preimplantation embryo

The present study examined expression of gamma-glutamate-cysteine ligase (GLCL; also known as gamma-glutamylcysteine synthetase), the rate-limiting enzyme for de novo synthesis of glutathione, in the preimplantation mouse embryo. Previous studies indicated that the cleavage stage embryo is unable to synthesize glutathione de novo. It is hypothesized that GLCL mRNA and protein are not normally expressed in the cleavage stage embryo, but either glutathione depletion or oxidation may induce their expression. In untreated embryos, RT-PCR and Western blotting revealed GLCL heavy subunit (GLCL-H) mRNA and protein only at the blastocyst stage of development. Furthermore, while diethyl maleate (DEM) exposure to deplete cellular glutathione did not induce expression of GLCL-H, exposure to tertiary-butyl hydroperoxide (tBH), an oxidizing agent, resulted in significant upregulation of GLCL-H expression in two-cell embryos. Neither treatment affected expression in blastocysts. Finally, HPLC analysis confirmed that tBH-treated embryos experienced oxidative stress, as indicated by an increase in the ratio of oxidized to reduced glutathione. This oxidative stress induced de novo glutathione synthesis in the cleavage stage embryo, as demonstrated by the subsequent recovery of reduced glutathione levels following DEM-induced depletion. In the absence of tBH treatment, however, cleavage stage embryos could not recover GSH after DEM-mediated depletion. This study demonstrates that the preimplantation embryo has the capacity to upregulate glutathione synthesis in response to oxidative stress but not GSH depletion. These results suggest that, while the preimplantation embryo is well adapted to dealing with oxidative stress, it may be poorly protected from GSH-depleting toxicants.     

The effect of t-butylated hydroxytoluene on glutathione linked detoxification mechanisms in rat

When rats were fed a diet containing 0.4% (w/w) butylated hydroxytoluene (BHT), a three-fold increase in total glutathione (GSH) S-transferase activity towards 1-chloro-2,4-dinitrobenzene (CDNB) was observed in liver but not in lung or kidney. Hepatic GSH S-transferase activities towards styrene oxide (SO) and 1,2-epoxy-3-(p-nitrophenoxy)propane (EPNP) were also increased, but to a lesser extent. Isoelectric focusing studies indicated that the activities of most of the rat liver GSH S-transferase isoenzymes were induced. Immunoprecipitation studies of the native and induced enzymes suggested that de novo synthesis of these proteins caused the increase in GSH S-transferase activity in liver. A two-fold increase in glutathione reductase activity in liver upon dietary administration of BHT was observed. Kinetic and physical properties of the native and induced enzymes were similar which may indicate that the induction is due to the synthesis of this enzyme. A significant increase in reduced glutathione (GSH) content in liver and lung was also seen in rats treated with BHT.     

Development of a whole cell assay to measure methotrexate-induced inhibition of thymidylate synthase and de novo purine synthesis in leukaemia cells

The cellular pharmacology of methotrexate (MTX) is complex, involving the inhibition of both de novo thymidylate and purine biosynthesis. Measurement of MTX-induced inhibition of de novo thymidylate and purine biosynthesis may allow optimisation of MTX therapy, and the aim of this study was to develop an assay to measure the activity of both pathways in the same cell sample, and so determine the effects of MTX treatment. In situ thymidylate synthase (EC 2.1.1.45) activity was measured by the release of 3H2O from [5'-3H]deoxyuridine and de novo purine synthesis by the incorporation of [14C]formate into adenine and guanine. Incubation of human leukaemia CCRF-CEM cells for 22 hr with 50 nM MTX resulted in approximately 90% inhibition of in situ thymidylate synthase activity, relative to control untreated cells, and after exposure to 1000 nM MTX activity could not be detected. In contrast, de novo purine synthesis, measured in the same sample, was not inhibited by exposure to 50 nM MTX, although activity was again completely abolished by exposure to 1000 nM MTX. To demonstrate the utility of the assay, lymphoblasts isolated from a child with acute lymphoblastic leukaemia (ALL) were also incubated for 22 hr with 1000 nM MTX. Both in situ thymidylate synthase activity and de novo purine synthesis were significantly inhibited, by 70% and 60% respectively, relative to the activity in untreated cells.     

Inhibitions of acid secretion by E3810 and omeprazole, and their reversal by glutathione.

A substituted benzimidazole ([4-(3-methoxypropoxy)-3-methylpyridine-2-yl]methylsulfinyl)- 1H-benzimidazole sodium salt (E3810), is a gastric proton pump (H+, K(+)-ATPase) inhibitor. E3810 and omeprazole inhibited acid accumulation dose dependently as measured with aminopyrine uptake in isolated rabbit gastric glands, their IC50 values being 0.16 and 0.36 microM, respectively. The addition of exogenous reduced glutathione (GSH) to the gland suspension reactivated dose dependently the acid secretion which had been inhibited by 2 microM E3810 or omeprazole as a function of the incubation time. Furthermore, GSH at 1 and 3 mM reversed the antisecretory effect of E3810 more quickly than it did that of omeprazole. The antisecretory effect of E3810 was slightly greater than that of omeprazole in histamine-stimulated fistula dogs in vivo. The duration of the antisecretory activity of E3810 at concentrations of 2 and 4 mg/kg was shorter than that of omeprazole at the same concentrations in pentagastrin-stimulated fistula dogs. The reversal of the antisecretory activity of the inhibitors in dogs is suggested to be due to the action of endogenous extracellular GSH, in addition to de novo synthesis of the proton pump, because bullfrog gastric mucosae were found in the present study to secrete GSH into the mucosal solution at the rate of about 0.25 nmol/min/g tissue.     

Inhibition by lithium and rubidium of gentamicin-induced release of N-acetyl-β-D-glucosaminidase from perfused rat kidney

N-acetyl-β-D-glucosaminidase (NAG) is one of the sensitive hydrolytic lysosomal enzymes which is released after renal tubular damages. We studied gentamicin-induced nephrotoxicity by determining the NAG release in perfused rat kidney. 100 μg/ml of gentamicin caused a time-dependent increase in enzymuria, peaking at 90 min. At this time the released NAG is about sixfold more than the control. The effect of concurrent perfusion with 100 μg/ml gentamicin and with 0.5 mmol/l lithium chloride or 0.5 mmol/l rubidium chloride in the perfusion fluid was also studied by measuring NAG activity in the perfusate. Both cations decrease the gentamicin-induced NAG release. However, the inhibitory effect of lithium chloride may be due to interference of this ion with the polyphosphoinositide cycle in renal tubular lysosomal membranes. There is no obvious evidence for an inhibitory effect of rubidium chloride.     

Selective release of glutathione transferase subunits from primary cultured rat hepatocytes by carbon tetrachloride and deoxycholic acid

The change in the activity of glutathione (GSH) transferases by carbon tetrachloride or deoxycholic acid, which induced hepatotoxicity, was studied using primary cultured rat hepatocytes. The activity of GSH transferases in the hepatocytes was decreased after the treatment with carbon tetrachloride or deoxycholic acid in their concentration- and incubation time-dependent manners. On the other hand, these compounds elicited the release of the activity of GSH transferases into the medium. Glycyrrhizin, an antihepatotoxic agent, inhibited the release of both aspartate transaminase (AST) and GSH transferases induced by carbon tetrachloride or deoxycholic acid. All subunits comprised of GSH transferases could not be released by these compounds. The main subunits of GSH transferases released by hepatotoxicity were identified as 3 and 4. These results indicate that hepatotoxicity is accompanied by the selective release of GSH transferase isozymes (class mu) following the loss of the enzymes activity in the cells.     

Pro-oxidant shift in glutathione redox state during aging

The GSH:GSSG ratio, which is the primary determinant of the cellular redox state, becomes progressively more pro-oxidizing during the aging process due to an elevation in the GSSG content and a decline in the ability for de novo GSH biosynthesis. The K(m) of glutamate-cysteine ligase (GCL), the rate-limiting enzyme in de novo GSH biosynthesis, significantly increases during aging, which would adversely affect the ability for rapid GSH biosynthesis, especially under stressful conditions. Experimental studies suggest that age-related accumulation of homocysteine, an intermediate in the trans-sulfuration pathway, may be responsible for causing the loss of affinity between GCL and its substrates. Over-expression of GCL has been shown to prolong the life span of Drosophila by up to 50%, suggesting that perturbations in glutathione metabolism play a causal role in the aging process.     

Uracil nucleotide synthesis in a human breast cancer cell line (MCF-7) and in two drug-resistant sublines that contain increased levels of enzymes of the de novo pyrimidine pathway

Cultured wild-type MCF-7 human breast cancer cells and two MCF-7 sublines that overproduce enzymes of the de novo pyrimidine biosynthetic pathway were compared with regard to: rate of de novo biosynthesis of uracil nucleotides, sensitivity of the de novo and salvage pathways to the concentration of intracellular uracil nucleotides, and potential of exogenous uridine at concentrations equivalent to plasma levels to affect de novo pyrimidine biosynthesis. The PALAR MCF-7 subline, which is resistant to N-(phosphonacetyl)-L-aspartate and has 5.2 times the activity of the first de novo enzyme as the wild-type MCF-7 cells, synthesizes uracil nucleotides via the de novo pathway at a rate that is 5.8 times that of the wild type MCF-7 cells. The PYRR MCF-7 subline, which is resistant to pyrazofurin and has 15.1 times the activity of orotate phosphoribosyltransferase as the wild-type MCF-7 cells, synthesizes uracil nucleotides via the de novo pathway at a rate that is 1.4 times that of wild-type MCF-7 cells. These results are consistent with carbamyl phosphate synthetase being the rate-controlling step of de novo pyrimidine biosynthesis. In the presence of exogenous uridine at concentrations equivalent to that found in plasma (4.4-8.6 microM), the uracil nucleotide pool of wild-type MCF-7 cells was expanded by 20% and de novo synthesis was inhibited by 55%. Incubation of PALAR MCF-7 cells with uridine at concentrations between 7.3 and 16.8 microM caused a 40% increase in the uracil nucleotide pool and a 30% inhibition of de novo synthesis. De novo synthesis of uracil nucleotides in PYRR MCF-7 cells was not affected by a greater than 10-fold increase in the uracil nucleotide pool. Salvage of [14C] uridine was inhibited by an expanded uracil nucleotide pool in the wild-type and PYRR MCF-7 cells but was not inhibited in the PALAR MCF-7 cell line. These results demonstrate that, although the overproduced enzymes exhibit substrate affinities and specificities in cell-free preparations similar to those of the wild-type enzymes, in intact cells the resistant cell lines exhibit marked differences in the control of de novo and salvage pyrimidine biosynthetic pathways by intracellular uracil nucleotides.     

Dose-related effects of methotrexate on purine and pyrimidine nucleotides and on cell-kinetic parameters in MOLT-4 malignant human T-lymphoblasts

The effects of methotrexate (MTX) on cytotoxicity (trypan blue exclusion and soft agar clonal growth), cell cycle perturbation, and purine and pyrimidine ribonucleotide and deoxyribonucleotide pools have been studied in MOLT-4 malignant T-lymphoblasts. Two concentrations of MTX, 0.02 microM and 0.2 microM have been utilized, which can be maintained in vivo during many hours in the maintenance therapy of acute lymphoblastic leukemia (ALL). The results are correlated with the effects of MTX on the inhibition of purine de novo synthesis. Treatment with 0.02 microM MTX results in an accumulation of cells in early S phase after 20 hr, as measured by DNA flow cytometry and by a significant increase of dCTP levels, followed by a slow progression of a cohort of cells through the cell cycle. Cytotoxicity also becomes evident starting from this point of time. The effects on deoxyribonucleotide pools are discussed in correlation with the inhibition of DNA synthesis. The changes in ribonucleotide pools are associated with the partial inhibition of purine de novo synthesis at 20-28 hr and suggest an inhibition of RNA synthesis. After 48 hr a reutilization of nucleotide precursors due to nucleic acid breakdown and a recovery of purine de novo synthesis is shown, associated with a recovery of RNA synthesis, whereas cytotoxicity increases. Treatment of MOLT-4 cells with 0.2 microM MTX results in a rapid complete cessation of cell progression through all parts of the cell cycle after 8 hr, associated with a depletion of all deoxyribonucleotide pools, complete inhibition of purine de novo synthesis, inhibition of RNA synthesis and a marked cytotoxicity. Ribonucleotide pools demonstrate a reutilization of nucleotide precursors after 12 hr of incubation without a recovery of purine de novo synthesis and RNA synthesis. These data show a close dose- and time-dependent correlation of the effects of MTX on purine de novo synthesis, UMP levels and other (deoxy)ribonucleotide pools, and on RNA and DNA synthesis in MOLT-4 cells having an active purine de novo synthesis. This correlation is absent in normal bone marrow cells and peripheral blood lymphocytes. These data can be used in order to elucidate the synergistic effects of sequential administration of MTX and 6-mercaptopurine.     

The effect of age on the tyramine-sensitive intraneuronal pool of norepinephrine in rat heart

The effect of age on the amount of norepinephrine (NE) released by tyramine (TYR) was investigated in hearts from Fischer 344 rats, 6, 12, and 24 months of age. Isolated hearts were perfused through an aortic cannula with Krebs-Ringer solution containing 1.6 X 10(-4) M TYR. alpha-Methyl-p-tyrosine (7.5 X 10(-5) M) and fusaric acid (10(-5) M) were added to inhibit de novo synthesis of NE. The effluent from the heart was collected continuously throughout 4 h of perfusion with TYR. The content of NE in the perfusion effluent was measured by electrochemical (EC) detection methods after alumina extraction and high-performance liquid chromatography (HPLC) separation. In addition, the amount of NE remaining in the heart after TYR perfusion was measured with HPLC/EC methodology. The amount of NE released from hearts was not significantly different among the ages [6 months, 612.5 +/- 68.2 ng; 12 months, 640.2 +/- 53.0 ng; 24 months, 593.8 +/- 38.2 ng; p greater than 0.05, analysis of variance (ANOVA)]. These amounts represent more than 93% of total cardiac NE at all ages (6 months, 97.6 +/- 0.5%; 12 months, 94.8 +/- 1.3%; 24 months, 93.7 +/- 1.7%). The t1/2 for the decline in NE in hearts was also similar at the three ages (6 months, 45.0 +/- 4.9 min; 12 months, 53.6 +/- 5.2 min; 24 months, 60.4 +/- 7.8 min; p greater than 0.05, ANOVA). These results indicate that the amount of NE released by TYR does not change with increasing age.     

Modulation of the release of histamine and arachidonic acid metabolites from human basophils and mast cells by auranofin

In these experiments the effects of pharmacological concentrations of auranofin, a new absorbable gold compound, were assessed on the release of histamine and peptide leukotriene C4 (LTC4) from human basophils and lung mast cells. Auranofin, at pharmacological concentrations, inhibited in vitro histamine and LTC4 release from human basophils induced by anti-IgE. Inhibition began at about 3 X 10(-7) M and was maximum at 10(-5) M. We also evaluated the effect of auranofin on the release of histamine and LTC4 induced by anti-IgE from mast cells purified from human lung. Auranofin (3 X 10(-7) to 10(-5) M) dose-dependently inhibited the release of histamine and LTC4 from human lung mast cells. Thus pharmacological concentrations of auranofin cause dose-related inhibition of histamine release and de novo synthesis of LTC4 by human basophils and lung mast cells.