Biomedical subjects

S T Crooke

Publications and source records attributed to S T Crooke.

At least 217 records · Page 12Linked to original sources

PubMed1985

Application of a tissue culture microtiter test for the detection of cytotoxic agents from natural products.

A method is described by which the growth inhibitory effects of cytotoxic compounds and fermentation broth cultures on adherent tumor cell lines can be quantitated. Cells are seeded into 96-well microtiter plates and 16 hours later the test compounds or broths are added to the wells. Cell growth is measured after three days (B16 mouse melanoma cells) or six days (HT-29, human colon carcinoma cells) by first fixing adherent cells, staining with Giemsa stain, washing away excess stain, then solubilizing stained cells with HCl. Absorbance is determined using a microELISA spectrophotometer and the data are transferred to and analyzed by a computer. The assay is rapid and reproducible and can be used to identify fermentation broths with cytotoxic components. Addition of DNA into the assay mixture (cells plus compound) inhibits the cytotoxic activities of certain DNA-reactive agents. The results of this study demonstrate the application of this assay system for primary and secondary evaluation of fermentation broths for in vitro antitumor activity.

Animals↗

PubMed1985

Characterization of guinea pig myocardial leukotriene C4 binding sites. Regulation by cations and sulfhydryl-directed reagents.

Using [3H]leukotriene C4 (LTC4) and radioligand-binding techniques, specific leukotriene C4 binding sites have been identified in membranes derived from guinea pig ventricular myocardium. High performance liquid chromatography analyses indicated that, in the presence of the gamma-glutamyl transpeptidase inhibitor L-serine-borate (80 mM), less than 2% of membrane-bound [3H]LTC4 was converted at 20 degrees to [3H]leukotriene D4 or [3H]leukotriene E4. The specific binding of 4 nM [3H]LTC4, in the presence of 80 mM L-serine-borate, reached a stable steady state within 15 min at 20 degrees (pH 7.5). A monophasic Scatchard plot of saturation binding data yielded a dissociation constant (Kd) of 27.5 +/- 6.0 nM and a maximum number of binding sites (Bmax) of 19.9 +/- 5.2 pmol/mg of membrane protein. Competition binding studies of [3H]LTC4 with synthetic leukotriene C4, leukotriene D4, and leukotriene E4 and the putative peptidoleukotriene antagonists FPL 55712, SKF 88046, and 4R-hydroxy-5S-1-cysteinylglycine-6Z-nonadecanoic acid revealed an order of potency of leukotriene C4 much greater than 4R-hydroxy-5S-1-cysteinylglycine-6Z-nonadecanoic acid greater than SKF 88046 greater than LTE4 greater than LTD4 greater than FPL 55712. The specific [3H]LTC4 binding was stimulated by the divalent cations Ca2+, Mg2+, and Mn2+ and to a lesser degree by the monovalent cations Na+, K+, Li+, and NH4+. CaCl2 (3 mM) and NaCl (150 mM) stimulated the LTC4 binding by increasing the Bmax to 42.6 +/- 5.9 and 35.0 +/- 2.0 pmol/mg, respectively, but had minimal effects on Kd. Pretreatment of the heart membranes with the sulfhydryl reagent N-ethylmaleimide decreased the specific [3H]LTC4 binding in a concentration-dependent manner. The N-ethylmaleimide-induced inactivation of [3H]LTC4 binding sites was protected by occupation of the binding site with the agonist leukotriene C4, but no protection was observed with the antagonist SKF 88046. Scatchard analyses of saturation isotherms indicated that 30 microM N-ethylmaleimide pretreatment reduced the Bmax of the [3H]LTC4 binding to 8.2 +/- 3.1 pmol/mg with minimal effects on Kd. The data provide direct biochemical evidence for specific [3H]LTC4 binding sites in the guinea pig heart membranes. The [3H]LTC4 binding sites appear to be modulated by divalent and monovalent cations and free sulfhydryl group(s) may be associated with the agonist-binding site. The results suggest that the physiological effects of the leukotrienes on the guinea pig heart may be mediated through membrane-bound receptors.

Animals↗

PubMed1985

Characterization of [3H]leukotriene D4 binding sites in guinea-pig ventricular myocardium.

[3H]Leukotriene (LT) D4 was used to identify specific LTD4 binding sites in guinea-pig ventricular myocardial membranes. High-performance liquid chromatography analyses indicated that, in the presence of the gamma-glutamyl transpeptidase inhibitor L-serine-borate (80 mM), less than 3% of membrane-bound [3H]LTD4 was converted to [3H]LTC4 or [3H]LTE4 at 30 degrees C. The specific [3H] LTD4 binding, assayed in the presence of 80 mM L-serine-borate, reached a stable steady state within 45 min at 30 degrees C (pH 7.5). A monophasic Scatchard plot of saturation binding data yielded an apparent dissociation constant (Kd) of 3.4 +/- 2.1 nM and a maximum number of binding sites of 850 +/- 91 fmol/mg of protein. Competition binding studies with [3H]LTD4, synthetic 5S, 6R-LTD4 (LTD4) and its diastereoisomer 5R,6S-LTD4, LTE4, LTC4 and the putative LT antagonists FPL 55712, 4R-hydroxy-5S-1-cysteinylglycine-6Z-nonadecanoic acid (2-nor-LTD1) and SKF 88046 demonstrated a potency order of LTD4 greater than LTE4 greater than LTC4 greater than 5R,6S-LTD4 much greater than 2-nor-LTD1. FPL 55712 and SKF 88046 were ineffective in displacing the specific [3H]LTD4 binding. Pretreatment of the heart membranes with the sulfhydryl reducing reagent dithiothreitol decreased the specific [3H]LTD4 binding in a concentration-dependent manner. Scatchard analyses of saturation isotherms indicated that 0.3 mM dithiothreitol pretreatment of heart membranes decreased the maximum number of binding sites of the [3H]LTD4 binding to 368 +/- 61 fmol/mg of protein with minimal effects on the apparent Kd.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1985

Molecular heterogeneity of leukotriene receptors: correlation of smooth muscle contraction and radioligand binding in guinea-pig lung.

The [3H]leukotriene C4 ([3H]LTC4) and [3H]leukotriene D4 ([3H] LTD4) specific binding sites in guinea-pig lung membranes were characterized and correlated with smooth muscle contractile activities of a series of LTC-, D- and E-type analogs. [3H]LTC4 bound to the specific sites with high affinity (dissociation constant Kd = 15 +/- 5 nM), saturable capacity (maximum binding = 68 +/- 15 pmol/mg of membrane protein), stereoselectivity and specificity. The [3H]LTC4 specific binding sites were detected in the membranes isolated from leukotriene sensitive (e.g., lung and heart) or insensitive (e.g., brain and red blood cells) tissues. [3H] LTD4 also bound to specific sites with high affinity (Kd = 0.20 +/- 0.05 nM), low capacity (maximum binding = 1.1 +/- 0.2 pmol/mg of membrane protein) stereoselectivity and specificity. The [3H] LTD4 specific binding sites were detected in the membranes isolated from lung and trachea. [3H]LTC4 specific binding was inhibited by treatment of the membranes with the sulfhydryl alkylating agent N-ethylmaleimide. [3H]LTD4 specific binding was more sensitive to heat treatment and p-hydroxymercuribenzoate than the [3H]LTC4 specific binding. Radioligand competition activities of the LTD- and LTE-type analogs correlated well with the agonist and antagonist smooth muscle contractile activities. In contrast, the radioligand competition activity of the LTC-type analogs did not correlate with smooth muscle contractile activities. These results indicate that the [3H]LTC4 and [3H]LTD4 specific binding sites in guinea-pig lung membranes are chemically and physically distinct. The [3H]LTD4 specific binding sites represent physiologically and pharmacologically important receptors, and the smooth muscle contraction induced by LTD-, and possible LTE-, type analogs are mediated through the LTD4 receptors.

Animals↗

PubMed1985

Evaluation of the in vivo antitumor activity and in vitro cytotoxic properties of auranofin, a coordinated gold compound, in murine tumor models.

The coordinated gold compound, 2,3,4,6-tetra-O-acetyl-1-thio-beta-D-glucopyranosato-S-triethyl phosphine gold (auranofin; Ridaura), was evaluated for antitumor activity in a variety of mouse tumor models. Of the 15 tumor models evaluated, auranofin was found to be active only against i.p. P388 leukemia. A number of dose schedules was used to measure activity against P388 with optimal activity observed at 12 mg/kg given daily, i.p., on Days 1 to 5. Auranofin was active against i.p. P388 leukemia only when administered i.p.; the drug was completely inactive when administered i.v., s.c., or p.o. on Days 1 to 5. Evaluation of the effects of auranofin in vitro demonstrated that survival curves for B16 melanoma cells as measured by the clongenic and dye exclusion assays were exponential and monophasic; cell cycle distribution was not altered, and auranofin displayed no preferential cytotoxicity to logarithmic or plateau growth phase cell populations; auranofin inhibited DNA, RNA, and protein synthesis at cytotoxic concentrations but showed no selective effect; the cytotoxic activity and cellular association of gold from auranofin were dose, time, and temperature dependent; and binding of auranofin gold to serum proteins markedly decreased cellular uptake of gold and cytotoxicity of auranofin in vitro.

Animals↗

PubMed1985

Effects of structural modifications of antitumor antibiotics (luzopeptins) on the interactions with deoxyribonucleic acid.

Luzopeptins consist of two identical, substituted quinolines linked to a cyclic decadepsipeptide, with a 2-fold symmetry. Luzopeptin A, with two acetylated sites in its peptide ring, is active against several experimental animal tumor systems. Luzopeptin B (one acetylated site) is less active, and luzopeptin C (no acetylation) is inactive. Our studies showed that all three luzopeptins and a half-molecule of luzopeptin C exhibited similar fluorescence (400 to 700 nm, with a peak at 490 to 496 nm) with an excitation spectrum in the 200 to 450 nm range (with a peak at 250 to 252, 337, and 385 nm). The half-molecule had the strongest fluorescence, followed in order by luzopeptins A, B, and C. DNA binding quenched both fluorescence and absorption of luzopeptins. Studies of the DNA-induced fluorescence and absorption quenching and the drug-induced viscosity and gel electrophoretic mobility changes of DNA suggested that luzopeptin C was slightly more effective than luzopeptins B and A in both the bifunctional DNA intercalation and the drug-induced DNA-DNA intermolecular cross-linking. Thus, the lack of antitumor activity of luzopeptin C is not the result of the lack of interactions with DNA. The half-molecule of luzopeptin C (quinoline with a pentapeptide) and smaller fragments (quinoline alone or with one to four peptide residues) did not react with DNA. Thus, the planar quinoline chromophore alone is unable to intercalate with DNA. The peptidic cyclic structure of luzopeptins is essential for the bifunctional intercalation of the twin chromophores, probably by providing proper conformational orientations of the chromophores.

Antibiotics, Antineoplastic↗

PubMed1984

Regulation of ligand binding to leukotriene D4 receptors: effects of cations and guanine nucleotides.

High affinity, stereoselective specific binding sites for [3H]leukotriene D4 [( 3H]LTD4) have been demonstrated in guinea pig lung membranes. Purine nucleotides quantitatively reduced [3H]LTD4 specific binding with a rank order potency of guanosine-5'-O-3-thiotriphosphate (GTP gamma S) = guanyl-5'-yl-imido-diphosphate [Gpp(NH)p] greater than GTP greater than ATP greater than GDP. In the presence of 1 microM Gpp(NH)p, the maximum number (Bmax) of [3H]LTD4 specific binding sites was reduced to 41 +/- 10 percent of the control level (950 +/- 150 fmol/mg membrane protein). In the presence of 3 microM Gpp(NH)p, the rate of association of [3H]LTD4 to the specific sites was estimated to have increased 2.5-fold. The rate of dissociation of [3H]LTD4 from the specific sites was also increased significantly in the presence of 50 microM Gpp(NH)p. The divalent cations, Ca2+ and Mg2+ (10 mM), increased the Bmax 2-fold and had minimal effects on the dissociation constant (Kd) of [3H]LTD4 specific binding. Sodium ions, at a concentration of 50 mM, reduced the Bmax, and had minimal effects on the Kd of [3H]LTD4 specific binding. These data indicate that guanine nucleotides, Na+, Mg2+ and Ca2+ regulate [3H]LTD4 binding to its receptors in guinea pig lung.

Animals↗

PubMed1984

Leukotriene C4 ([3H]-LTC4) binding to membranes isolated from a hamster smooth muscle cell line (DDT1MF2).

Leukotriene C4 (LTC4) has been demonstrated to induce contraction of the smooth muscle cell line DDT1MF2. A partially purified membrane fraction obtained from these cells exhibited a high affinity binding site for LTC4. Binding of [3H]-LTC4 was saturable, specific and reversible with a dissociation constant (Kd) of 21 +/- 4 nM. The maximum number of binding sites (Bmax) was 55 +/- 5 pmol/mg of protein. Specificity was demonstrated in competition studies in which the Ki of LTC4 against specifically bound [3H]-LTC4 was 12 nM whereas Leukotriene D4 (LTD4) and Leukotriene E4 (LTE4) had a Ki of 38 +/- 4 and 4.7 +/- 0.5 nM respectively. A previously described antagonist of leukotriene-induced smooth muscle contraction PFL 55712 had a Ki of 23 +/- 2 nM as determined by competition binding experiments.

Animals↗

PubMed1984

Deoxyribonucleic acid breaks produced by 4'-(9-acridinylamino)methanesulfon-m-anisidide and copper.

We have demonstrated that 4'-(9-acridinyl-amino)methanesulfon-m-anisidide (mAMSA), in the presence of Cu(II) ion, causes the breakage of plasmid pDPT275 and pBR322 superhelical form I DNA. In neutral pH, the degradative product was nicked, relaxed form II DNA, resulting from single-stranded DNA breakage. The extent of DNA breakage was both mAMSA concentration and Cu(II) concentration dependent. DNA breakage increased with increasing time of drug treatment. The mAMSA-Cu(II)-induced DNA breakage varied with pH values and also with the nature of the buffer systems. In both Tris-HCl and borate buffers the extent of DNA breakage increased with increasing pH. In Tris-HCl buffer (pH 7-9), only single-strand breaks were obtained, whereas in borate buffer (pH 9-10.5), linear form III DNA was obtained. At equivalent pH, the optimum buffer was borate. No breakage was observed at pH values below 6. The interaction of Cu(II) with mAMSA was examined by using absorption and fluorescence spectroscopies. Interaction of Cu(II) with mAMSA was characterized by a decrease in the absorption at 435 and 420 nm with a simultaneous increase at 330 nm. A highly fluorescent product was obtained upon reacting mAMSA with Cu(II), with an emission spectrum (excitation at 400 nm) showing a doublet at 430 and 450 nm and a shoulder around 480 nm. The spectral changes are also dependent similarly on the pH and the nature of buffer. Other divalent metal ions such as Co(II), Cd(II), Ni(II), and Zn(II) do not induce DNA breakage or spectral changes. The oAMSA isomer, which has no antitumor activity, is less effective in inducing DNA breakage than the mAMSA.

Aminoacridines↗

PubMed1984

Mechanism of deoxyribonucleic acid breakage induced by 4'-(9-acridinylamino)methanesulfon-m-anisidide and copper: role for cuprous ion and oxygen free radicals.

4'-(9-Acridinylamino)methanesulfon-m-anisidide (mAMSA) interacts with Cu(II) ion, as indicated by changes in the mAMSA absorption spectrum induced by Cu(II). The spectral changes are due to the oxidation of mAMSA by Cu(II), resulting in an oxidized mAMSA product and Cu(I). Two lines of evidence for the oxidation of mAMSA are as follows: (1) The spectral changes induced by manganese oxide, an oxidizing agent, were similar to those induced by Cu(II), and (2) the Cu(II)-induced spectral changes were reversed by a reducing agent, NADPH. Thin-layer chromatographic studies showed the oxidized mAMSA product to be N1-methylsulfonyl-N4-(9-acridinyl)-3-methoxy-2,5-cyclohexadiene-1, 4-diimine (mAQDI). The involvement of Cu(I) in the reaction was demonstrated by the use of two Cu(I)-specific chelating agents, neocuproine and bathocuproine. Neocuproine or bathocuproine chelated the Cu(I) ions in the mixture, producing Cu(neocuproine)2+ complex or Cu(bathocuproine)2+ complex. The stoichiometry of mAMSA-Cu(II) interactions was determined by titrating the mAMSA-Cu(II) mixtures with bathocuproine. Job plots of the absorbance at 480 nm showed a clear end point at a Cu(II)/mAMSA ratio of 1.5/1, indicating that 1.5 equiv of Cu(II) reacts with 1 equiv of mAMSA to produce 1.5 equiv of Cu(I). Cu(I) plays an important role in the mAMSA-Cu(II)-induced DNA breakage, since in the presence of neocuproine the DNA breakage is inhibited. Up to 200 microM, Cu(I) by itself is virtually ineffective, in contrast to the mixture of mAMSA and Cu(II).(ABSTRACT TRUNCATED AT 250 WORDS)

Aminoacridines↗

PubMed1984

Characterization of the leukotriene D4 receptor in guinea-pig lung.

The effects of monovalent, divalent cations, buffere species and pH dependency on [3H]leukotriene D4 binding to the receptor have been characterized in vitro by using a radioligand binding assay. It was found that Ca2+, Mg2+, Co2+ and Mn2+ enhanced the specific binding. High concentrations of NaCl (150-300 mM) inhibited the specific binding to the receptor. The specific binding was also found to be higher in Pipes buffer (pH 6.5) than in Tris, Hepes and phosphate buffer at pH 7.0-8.0. Conversion of [3H]leukotriene D4 was minimized by inclusion of 1 mM cysteine, glycine in the incubation buffer and maintaining the temperature at 22 degrees C. Under the conditions employed, the dissociation constant (KD) and the receptor density (Bmax) were calculated to be 1.8 +/- 0.9 nM and 2100 +/- 375 fmol/mg protein respectively. The leukotriene antagonist FPL 55712, agonist 5R, 6S-LTD4 and LTE4 competed with the [3H]LTD4 binding to the receptor. Prostaglandins, alpha-, beta-adrenergic and dopaminergic receptor agonists and antagonists did not compete significantly.

Animals↗

PubMed1984

Identification of leukotriene D4 specific binding sites in the membrane preparation isolated from guinea pig lung.

A radioligand binding assay has been established to study leukotriene specific binding sites in the guinea pig and rabbit tissues. Using high specific activity [3H]-leukotriene D4 [( 3H]-LTD4), in the presence or absence of unlabeled LTD4, the diastereoisomer of LTD4 (5R,6S-LTD4), leukotriene E4 (LTE4) and the end-organ antagonist, FPL 55712, we have identified specific binding sites for [3H]-LTD4 in the crude membrane fraction isolated from guinea pig lung. The time required for [3H]-LTD4 binding to reach equilibrium was approximately 20 to 25 min at 37 degrees C in the presence of 10 mM Tris-HCl buffer (pH 7.5) containing 150 mM NaCl. The binding of [3H]-LTD4 to the specific sites was saturable, reversible and stereospecific. The maximal number of binding sites (Bmax), derived from Scatchard analysis, was approximately 320 +/- 200 fmol per mg of crude membrane protein. The dissociation constants, derived from kinetic and saturation analyses, were 9.7 nM and 5 +/- 4 nM, respectively. The specific binding sites could not be detected in the crude membrane fraction prepared from guinea pig ileum, brain and liver, or rabbit lung, trachea, ileum and uterus. In radioligand competition experiments, LTD4, FPL 55712 and 5R,6S-LTD4 competed with [3H]-LTD4. The metabolic inhibitors of arachidonic acid and SKF 88046, an antagonist of the indirectly-mediated actions of LTD4, did not significantly compete with [3H]-LTD4 at the specific binding sites. These correlations indicated that these specific binding sites may be the putative leukotriene receptors in the guinea-pig lung.

Animals↗

PubMed1984

Cloning and expression of a yeast copper metallothionein gene.

The induction of a copper-binding metallothionein (Cu-MT) was studied in yeast, Saccharomyces cerevisiae, and a relationship between copper resistance and intracellular levels of Cu-MT in these eukaryotes was established. Poly(A)-containing RNA from a copper-resistant (Cur) yeast strain, which synthesized abundant quantities of Cu-MT and in which Cu-MT gene transcription was enhanced 50-fold upon exposure to CuSO4, was used to screen yeast genomic DNA clones. Restriction analysis revealed common XbaI and KpnI sites in five genomic clones isolated. The transcription of these clones was regulated by copper. Transformation of a copper-sensitive (Cus) yeast strain by one of these clones confers copper resistance in yeast. The results suggest that the expression of the Cu-MT gene is, in part, responsible for mediating copper resistance in yeast.

Cloning, Molecular↗

PubMed1984

Identification of specific binding sites for leukotriene C4 in human fetal lung.

Specific leukotriene C4 (LTC4)1 binding sites were identified in membrane preparations from human fetal lung. Specific binding of [3H]-LTC4 represented 95 percent of total binding, reached steady-state within 10 minutes and was rapidly reversible upon addition of excess unlabeled LTC4. Binding assays were performed at 4 degrees C under conditions which prevented metabolism of [3H]-LTC4 (80 mM serine-borate, 10 mM cysteine, 10 mM glycine). Under these conditions, greater than 95 percent of the membrane bound radioactivity, as analyzed by high performance liquid chromatography, co-eluted with the LTC4 standard. Computer-assisted analyses of saturation binding data showed a single class of binding sites with a dissociation constant (Kd) of 26 + 6 nM and a density (Bmax) of 84 + 18 pmol/mg protein. Pharmacological specificity was demonstrated by competition studies in which specific binding of [3H]-LTC4 was displaced by LTC4 and its structural analogs with inhibition constants (Ki) of 10 to 30 nM, whereas LTD4, diastereoisomers of LTD1, LTE4 and the end organ antagonist FPL 55712 were 150 to 700 fold less potent competitors than LTC4. These results provide evidence for specific, reversible, saturable, high affinity binding sites for [3H]-LTC4 in human fetal lung membranes.

Binding Sites↗

PubMed1984

Copper metallothionein of yeast, structure of the gene, and regulation of expression.

Addition of copper to yeast cells leads to the induction of a low molecular weight, cysteine-rich protein that binds copper. This protein, termed copper chelatin or thionein, is related to the metallothionein family of proteins that are induced in response to cadmium and zinc in vertebrate cells. We have determined the structure of the yeast copper-binding protein by DNA sequence analysis of the gene. Although the 6573-dalton yeast protein is substantially divergent from vertebrate metallothioneins, the arrangement of 12 cysteine residues, which is a hallmark of metal-binding proteins, is partially conserved. We analyzed the regulatory DNA sequence of the gene by fusing it with the Escherichia coli galactokinase gene and assaying the levels of enzyme activity in yeast in response to copper. The transcriptional activation has a specific requirement for copper. Zinc, cadmium, and gold were unable to regulate the galactokinase activity. The yeast copper metallothionein regulatory sequences represent a previously unreported class of yeast promoter that is regulated by copper.

Amino Acid Sequence↗

PubMed1984

Improved genetically modified Escherichia coli strain for prescreening antineoplastic agents.

Clinical experience suggests that drugs that interact with and damage DNA are useful in cancer chemotherapy (H. Umezawa , p. 43-72, in V. T. DeVita , Jr., and H. Busch [ed.], Methods in Cancer Research; Cancer Drug Development, vol. XVI, 1979). Prescreening systems for antitumor agents in natural products require assays that are exquisitely sensitive, since the active components are often produced in quantities of micrograms per milliliter or less. One assay used to identify agents that interact with DNA is the biochemical induction assay, utilizing Escherichia coli BR 513 (R. K. Elespuru and R. J. White, Cancer Res. 43:2819-2830, 1983). In this paper we describe a genetic modification of strain BR 513 that displays an expanded spectrum of activity. This strain may provide an improved prescreen for detecting natural products that interact with DNA.

Ampicillin↗

PubMed1983

Molecular mechanisms of action of auranofin and other gold complexes as related to their biologic activities.

The molecular mechanisms of action of gold complexes in the treatment of rheumatoid arthritis are partially known, as are the mechanisms of action and potential utility of gold complexes in the treatment of neoplastic disease. In this paper, data relative to the mechanism of cytotoxicity and structure activity relationships are presented. Concepts of future research are also discussed.

Anti-Inflammatory Agents↗

PubMed1983

Peptidoleukotrienes: distinct receptors for leukotriene C4 and D4 in the guinea-pig lung.

Using [3H]-leukotriene C4 ([3H]-LTC4) and [3H]-leukotriene D4 ([3H]-LTD4), specific peptidoleukotriene receptors have been identified in membranes derived from guinea-pig lung. In the presence of 0.1 mM guanyl-5'-yl-imidodiphosphate, which completely inhibits [3H]-LTD4 binding, [3H]-LTC4 binding was protein- and temperature-dependent, reached equilibrium within 15 minutes at 20 degrees C and was reversible. Guanine nucleotides had no effect on the [3H]-LTC4 binding. Competition studies with [3H]-LTC4, peptidoleukotrienes C4, D4, E4 and the peptidoleukotriene antagonist FPL 55712 revealed an order of potency of leukotriene C4 much greater than E4 greater than D4 greater than FPL 55712. [3H]-LTD4 competition studies indicated an order of potency of LTD4 greater than LTE4 greater than LTC4 much greater than FPL 55712. Bioconversion of [3H]-LTC4, as determined by high performance liquid chromatography, was less than 3 percent. The data suggest the guinea-pig lung may contain biochemically distinct receptors for LTC4 and LTD4.

Animals↗