The characteristics of calcium antagonism and vascular effect of 2-methoxyethyl (E)-3-phenyl-2-propen-1-yl (±)-1, 4-dihydro-2, 6-dimethyl-4-(3-nitrophenyl) pyridine-3, 5-dicarboxylate (FRC-8653) were investigated. FRC-8653 inhibited an increase in intracellular free calcium concentration during membrane depolarization in PC12 cells. FRC-8653 also inhibited the specific binding of ³H-nitrendipine to cardiac membranes, in a similar manner to nifedipine and nicardipine. FRC-8653 inhibited KCl- and CaCl₂-induced contractions in isolated rabbit aorta, but failed to affect norepinephrine-induced contraction. The vasorelaxing effect of FRC-8653 in rabbit aorta developed more slowly than those of nifedipine and nicardipine. In ex vivo experiment, the inhibitory effect of orally administered FRC-8653 against KCl-contraction in rat aorta lasted longer than that of nifedipine. These findings suggest that FRC-8653 dilates blood vessels by blocking calcium influx via dihydropyridine-sensitive, voltage-dependent calcium channels and that the vascular effects are slow in development and long in duration.

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The influence of para substituent of phenol on the property of phospholipid dependence of uridine diphosphate-glucuronyltransferase (UDPGT) was studied using hepatic microsomes of the rats. para Substituents used were p-ethyl, p-tert-butyl and p-phenyl. Glucuronidations of phenol and its derivatives were inhibited with each other. When phospholipids of the microsomes were removed with phospholipase A₂, V_max and K_m decreased in the order phenol, p-ethylphenol, p-tert-butylphenol, p-phenylphenol, indicating that this change by delipidation increased with van der Waals volume (V_w) of para substituent. Arrhenius plots of glucuronidation for phenol and p-ethylphenol exhibited a change in slope, responding to thermotropic property of the membrane labelled with 1, 6-diphenyl-1, 3, 5-hexatriene (DPH). In contrast, p-tert-butyl and p-phenyl substituents showed linear Arrhenius plots. The activation energy (E_a) for glucuronidation of p-phenylphenol was increased by delipidation, whereas E_a of other substrates did not show any remarkable change.

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The H₂-receptor antagonists, cimetidine, ranitidine, famotidine and nizatidine, were tested for their effect on the isolated smooth muscle strips from the rabbit stomach fundus and sigmoid colon. These H₂-receptor antagonists were found to possess a concentration-dependent contractile effect on the above smooth muscle preparations and the order of potency was : ranitidine = nizatidine > famotidine > cimetidine. In addition, the smooth muscle preparations from the sigmoid colon were significantly more sensitive to the above compounds than the smooth muscle preparations from the stomach fundus.

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An enzyme immunoassay (EIA) for assessment of cell activation and proliferation was developed by the use of monoclonal antibodies (MoAb) directed against a ubiquitious growth-associated antigen, gp125. Test cells distributed in a microtest plate were labeled with anti-rat gp125 MoAb, B3, for rat cells or anti-human gp125 MoAb, HBJ127, for human cells and subsequently with rabbit anti-mouse immunoglobulins. The cell-bound antibody was assessed by a colorimetric enzyme assay using horseradish peroxidase-modified protein A and 2, 2'-azinobis (3-ethylbenzthiazoline sulfonic acid). To terminate the reaction and to make the reaction mixture transparent, sodium dodecyl sulfate (SDS) was added to the mixture. The intensity of the developed color was measured by use of a multiwell scanning spectrometer. The titration curves obtained by the EIA for cells were practically similar to those obtained by the conventional ³H-thymidine (Tdr) uptake method, and the appropriate cell number to be used for the assay was indicated to be 3 to 50×10³ cells per well. This method was applicable not only for quantitation of cell number of growing cells but also for measuring mitogenic responses of lymphocytes as revealed by the data obtained from Con A stimulation of lymphocytes. These results indicate that this new EIA method using anti-gp125 antibodies is useful for quantitative assays of cell growth and cell activation in the research fields of oncology and immunology.

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We studied the effect of feeding ursodeoxycholylcysteic acid, the cysteic acid conjugated analog of ursodeoxycholyltaurine, on serum and liver cholesterol levels and on intestinal absorption of cholesterol and bile salts in hamsters. Addition of ursodeoxycholylcysteic acid to the cholesterolenriched diet reduced the elevation of serum and liver cholesterol levels caused by feeding cholesterol. However, supplementation with ursodeoxycholylcysteic acid to the standard diet did not show any significant change in serum and liver cholesterol levels. Administration of ursodeoxycholylcysteic acid caused a decrease in dietary cholesterol absorption but did not interfere with the ileal transport of endogenous bile salts. Hence the hypocholesterolemic activity of dietary ursodeoxycholylcysteic acid is thought to be the effect on intestinal absorption of cholesterol but not to be the interruption of the enterohepatic circulation of bile salts.

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Glucocorticoids inhibit IgE antibody-mediated passive cutaneous anaphylaxis (PCA) and chemical mediator-induced cutaneous reactions elicited in the mouse ear. In the present study, we investigated the effect of actinomycin D, a protein synthesis inhibitor, on dexamethasone-caused inhibition of PCA and histamine-induced cutaneous reaction in the mouse ear. Tyrosine aminotransferase (TAT) activity in the liver, which was estimated as an index for protein synthesis, significantly increased by the administration of hydrocortisone, prednisolone and dexamethasone. Significant increase in TAT activity was observed from 2h after glucocorticoid administration and peaked at 4h, and declined gradually thereafter. Cycloheximide even at high doses of 100 and 300 mg/kg failed to affect the increase in TAT activity by dexamethasone. On the contrary, actinomycin D at doses of 1 and 10 mg/kg abrogated the TAT activity increase by dexamethasone almost completely. Treatment with 1 mg/kg of actinomycin D, however, failed to affect the inhibition of PCA and histamine-induced cutaneous reaction by dexamethasone. These results suggest that glucocorticoids exhibit their inhibitory action of PCA and chemical mediator-induced cutaneous reactions in mice through a mechanism resistant to actinomycin D treatment.

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The interaction of pranoprofen, pranoprofen glucuronide and pranoprofen methylester with human serum albumin (HSA), was investigated by equilibrium dialysis and spectroscopic techniques. The binding affinities of pranoprofen glucuronide and pranoprofen methylester to HSA were found to be almost the same, although they were remarkably small as compared to that of the parent compound, pranoprofen. Pranoprofen and pranoprofen methylester showed steroselective affinities to HSA. It was found from the competitive displacement experiments using the fluorescent probes that the specific binding site for pranoprofen was site II, the diazepam site, and that the binding sites of pranoprofen glucuronide and pranoprofen methylester were site I, the warfarin site. In addition, from the binding data with modified HSA, it seemed that tyrosine-411 was specifically involved in the pranoprofen binding. The absorption spectral changes which accompanied the binding of pranoprofen and pranoprofen methylester to HSA or detergents implied that the HSA binding site of pranoprofen consisted of a cationic site on the surface of the albumin molecule with a hydrophobic region to accommodate the aromatic ring and that the binding site for pranoprofen methylester seemed to occupy a wide hydrophobic area. These limited data indicated differences in the location and microenvironments of binding sites for pranoprofen and its glucuronide on the HSA molecule.

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Constant exposure of mastocytoma P-815 cells to adenosine 3', 5'-cyclic decylphosphoramidate (1), which is permeable to the cell membrane and resistant to the action of phosphodiesterase, caused a dose-dependent (1 to 50μM) inhibition in the synthesis of DNA and cell proliferation. Pretreating the cells with compound 1 (20μM, 4h) caused considerable inhibition of the incorporation of [³H] thymidine ([³H] TdR) into [³H] deoxythymidine 5'-triphosphate ([³H] dTTP) and that of [¹⁴C] hypoxanthine into nucleic acid, but not the synthesis of [¹⁴C] dTTP from [U-¹⁴C] aspartate. These results indicate that compound 1 preferentially inhibits the salvage synthesis of intracellular nucleotides and nucleic acids. Thymidine kinase, a key enzyme in salvage synthesis of nucleotides, was almost undetectable in cells pretreated with compound 1 at 20 μM for 4 h or at 5μM for 15h. On the other hand, compound 1 activated partially purified cAMP-dependent protein kinase A from bovine heart. Judging from these observations, it is likely that compound 1 readily permeates the cell membrane, activates cAMP-dependent protein kinase, then inhibits the salvage synthesis of nucleotides and nucleic acids by inhibiting thymidine kinase, which results in the inhibition of cell growth.

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