Biomedical subjects

M Mio

Publications and source records attributed to M Mio.

At least 37 records · Page 2Linked to original sources

PubMed1993

Ca(2+)-induced cortisol secretion from permeabilized bovine adrenocortical cells: the roles of calmodulin, protein kinase C and cyclic AMP.

In order to clarify the role of intracellular second messenger systems in the cortisol secretion from bovine adrenocortical (BAC) cells, the cells were permeabilized with beta-escin and stimulated intracellularly with various compounds. When the permeabilized BAC cells were exposed to submicromolar concentrations of Ca2+, a prompt cortisol secretion was elicited in a concentration-dependent manner. As the cells were stimulated with 12-O-tetradecanoyl-phorbol-13-acetate and 1-oleoyl-2-acetyl-glycerol, slow but persistent cortisol secretion was elicited, but in the case of 4 alpha-phorbol-12,13-didecanoate, no such effect was observed. The Ca(2+)-induced cortisol secretion was inhibited by simultaneous applications of calmodulin and protein kinase C (C kinase) inhibitors, but no significant inhibition was elicited by protein kinase A (A kinase) inhibitor. The results seem to indicate that in the Ca(2+)-induced cortisol secretion calmodulin may stimulate the initial stage, while C kinase may be involved mainly in the late phase of the secretion. In addition, cyclic AMP (cAMP) was also effective in activating cortisol secretion from permeabilized BAC cells. The cAMP-induced cortisol secretion was suppressed by an A kinase inhibitor but not affected by calmodulin or C kinase inhibitor. When Ca2+ and cAMP were added simultaneously at concentrations lower than those required to induce the cortisol secretion separately, a marked cortisol secretion was elicited, suggesting that a synergic action exists between Ca(2+)- and cAMP-activated systems. The Ca(2+)-induced cortisol secretion was suppressed by ruthenium red, an inhibitor of Ca2+ transport in the mitochondria. Although both NADP+ and NADPH elicited only a transient cortisol secretion, simultaneous addition of Ca2+ with NADP+ or NADPH caused a potent and sustained cortisol secretion. The augmentation due to Ca2+ on the NADP+ (or NADPH)-induced cortisol secretion was inhibited by the addition of a calmodulin inhibitor or a C kinase inhibitor, but not such effect was caused by A kinase inhibitor. From the present investigation, it was concluded that the Ca(2+)-dependent intracellular signal transduction may simulate the cortisol synthesis systems in the mitochondria of BAC cells.

Adrenal Cortex↗

PubMed1993

Inhibitory effects of emedastine difumarate on histamine release.

The inhibitory effects of emedastine difumarate on histamine release were studied in rat peritoneal mast cells. Emedastine significantly inhibited substance P (SP)-induced histamine release at concentrations above 10(-9) M in the presence of extracellular Ca2+ and at concentrations above 10(-11) M in its absence. At concentrations of 10(-8) M or higher, emedastine significantly inhibited SP-induced Ca2+ release from intracellular Ca stores and SP-induced 45Ca uptake into mast cells. Emedastine also inhibited passive peritoneal anaphylaxis in rats and guinea pigs. We conclude that the clinical antiallergic effects of emedastine involve the inhibition of histamine release and that this inhibition is mediated by the inhibition of Ca2+ release from intracellular Ca stores and the inhibition of Ca2+ influx into mast cells.

Anaphylaxis↗

PubMed1993

Antiallergic profile of the novel H1-antihistaminic compound levocabastine.

Levocabastine hydrochloride (R50 547, CAS79516-68-0) caused no inhibitory effect on the histamine release from rat peritoneal mast cells induced by compound 48/80, A23187 and concanavalin A. However, the drug inhibited histamine release from passively sensitized mast cells and passive peritoneal anaphylaxis in rats, though higher concentrations or doses were required. Moreover, levocabastine provided a relatively potent inhibitory effect on histamine release from lung pieces of actively sensitized guinea pigs exposed to antigen, and simultaneously the drug prevented a decrease in the cyclic AMP (cAMP) content. Levocabastine potently inhibited histamine-induced cutaneous reactions in rats and the drug also prevented histamine-induced contraction of isolated guinea pig ileum. Levocabastine did not induce any significant changes in platelet aggregation or in the contraction of guinea pig ileum induced by platelet activating factor (PAF). However, the drug inhibited eosinophil migration induced by PAF. The chemotaxis of neutrophils induced by N-formyl-methionyl-leucylphenylalanine (fMLP) was also inhibited by levocabastine in a dose-dependent fashion. Levocabastine has no influence on the order parameter tested with liposomes, suggesting that the drug provides no significant effect on the membrane fluidity of lipid bilayer. These results seem to indicate that the antiallergic effect of levocabastine is mainly dependent on its potent antihistaminic activity.

Anaphylaxis↗

PubMed1993

Histamine-induced production of interleukin-1 alpha from murine bone marrow stromal cells and its inhibition by H2 blockers.

In this study, the role of histamine in interleukin-1 (IL-1) formation in murine bone marrow stromal cells was investigated in vitro. It was found that histamine and 4-methylhistamine increased the number of granulocyte colony-forming units in murine bone marrow cells. A similar effect was elicited by dibutyryl-cAMP and theophylline. When histamine and H2 agonists, such as 4-methylhistamine and dimaprit, were added to the culture medium containing murine bone marrow stromal cells, thymocyte comitogenic activity detected in the medium increased significantly. However, no such effect was observed in the case of 2-methyl-histamine, an H1 agonist. Histamine-induced production of thymocyte comitogenic activity in bone marrow stromal cells was inhibited by some H2 antagonists, such as cimetidine, ranitidine, and famotidine, but not by the H1 antagonist pyrilamine. Histamine was also effective in inducing the colony-promoting activity in murine bone marrow stromal cells. This was also inhibited by H2 antagonists such as cimetidine, ranitidine, and famotidine. Histamine elicited an increase in cAMP content in bone marrow stromal cells. From gel filtration analysis, the molecular mass of the active substance produced by bone marrow stromal cells in response to histamine was in the range of 15 to 20 kDa. By means of Western blotting analysis, it was found that production of pro-IL-1 alpha in the bone marrow stromal cells was induced by histamine. The production of pro-IL-1 alpha in bone marrow stromal cells stimulated by histamine was inhibited not only by H2 antagonists, such as cimetidine, ranitidine, and famotidine, but also by the protein kinase A antagonist KT-5720. These results indicate that histamine stimulates the production of IL-1 alpha in bone marrow stromal cells and that this results in the proliferation and differentiation of neutrophil progenitor cells.

Animals↗

PubMed1992

de novo synthesis of calmodulin binding protein in substance P-induced steroidogenesis in bovine adrenocortical cells.

In order to clarify the mechanism of substance P (SP)-induced cortisol secretion from bovine adrenocortical (BAC) cells, protein synthesis at the early stage of SP-stimulation in BAC cells was investigated. Both SP and adrenocorticotropic hormone (ACTH) increased [3H]leucine uptake into BAC cells in a dose-dependent fashion. Although the SP-induced [3H]leucine uptake precedes the cortisol secretion, ACTH was slower in inducing [3H]leucine uptake and cortisol secretion. Protein synthesis inhibitors, actinomycin D and cycloheximide, were potent in inhibiting the SP-induced cortisol secretion. SDS-PAGE analysis, revealed that a 240 kDa protein is newly synthesized in BAC cells in response to SP but not ACTH. It was also indicated that the production of this 240 kDa protein was elicited about 30 min after stimulation by SP. Moreover, A23187 and 12-O-tetradecanoyl-phorbol-13-acetate (TPA) also caused a rapid [3H]leucine uptake and production of 240 kDa protein. In contrast, dibutyryl cAMP did not induce the synthesis of this 240 kDa protein. Calmidazolium, a calmodulin inhibitor, effectively inhibited not only [3H]leucine uptake but also 240 kDa protein production due to SP. On the other hand, KT-5720, an inhibitor of protein kinase A, had no effect on [3H]leucine uptake or 240 kDa production. Using the [125I]calmodulin-membrane overlay method, it was found that the 240 kDa protein was a newly synthesized calmodulin binding protein. From the present study, it was concluded that the de novo synthesis of this 240 kDa protein may be intimately related to the cortisol secretion in SP-stimulated BAC cells associated with an activation of the Ca-calmodulin pathway.

Adrenal Cortex↗

PubMed1992

Histamine-induced differentiation of HL-60 cells. The role of cAMP and protein kinase A.

When HL-60 cells were stimulated with histamine, a significant differentiation of the cells toward neutrophils was elicited. Histamine increased phagocytic activity, but it reduced myeloperoxidase activity of HL-60 cells. Histamine-induced differentiation in HL-60 cells was inhibited not only by H2 antagonists, such as cimetidine, ranitidine and famotidine, but also by an inhibitor of protein kinase A (A kinase), KT-5720. Histamine increased the cAMP level and A kinase activity in HL-60 cells; both increases preceded the cell differentiation. Histamine also enhanced phosphorylation of a 160 kD protein in HL-60 cells, while H2 antagonists and KT-5720 inhibited this phosphorylation. The results of the present study indicate that an activation of A kinase via H2 receptor stimulation may cause the phosphorylation of a 160 kD protein and that this phosphorylation is probably involved in the process leading to differentiation of HL-60 cells.

Carbazoles↗

PubMed1992

Cortisol secretion induced by substance P from bovine adrenocortical cells and its inhibition by calmodulin inhibitors.

When primary cultured bovine adrenocortical cells were treated with substance P (SP) at concentrations higher than 10 pM, cortisol output increased in a dose-dependent fashion. Although other neurokinins, such as neurokinin A (NKA) and neurokinin B (NKB), were also effective in secreting cortisol, SP was the most potent among the tested neurokinins, the potency order being SP greater than NKA much greater than NKB. This suggests that the NK-1 type receptor on adrenocortical cells may be the site of action of SP on cortisol secretion. The maximal response in SP-induced cortisol secretion was comparable to that elicited by adrenocorticotropic hormone (ACTH). SP-induced cortisol secretion was dependent upon extracellular Ca2+ concentrations, and 45Ca2+ uptake into adrenocortical cells treated with SP was long-lasting. While, in the case of ACTH, 45Ca2+ uptake proceeded transiently, the increase in intracellular cAMP content was much greater compared with that of SP. Although KT-5720, an inhibitor of protein kinase A, inhibited potently ACTH-induced cortisol secretion, SP-induced secretin was not affected by this inhibitor at all. On the other hand, calmodulin inhibitors, such as calmidazolium, trifluoperazine and N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide, were not more effective in inhibiting SP-induced cortisol secretion than secretion induced by ACTH. The present study indicates that SP may be one of the physiological stimulants of cortisol secretion and that an increase in intracellular Ca2+ concentration and the subsequent activation of calmodulin may precede SP-induced cortisol secretion.

Adrenal Cortex↗

PubMed1992

Antiallergic effect of epinastine (WAL 801 CL) on immediate hypersensitivity reactions: (I). Elucidation of the mechanism for histamine release inhibition.

Epinastine caused an inhibition of histamine release from rat peritoneal mast cells induced by both antigen-antibody reaction and compound 48/80. Epinastine was similarly effective in inhibiting compound 48/80-induced histamine release not only from isolated rat peritoneal mast cells but also from rat mesenterial pieces. Also, histamine release from lung pieces obtained from actively sensitized guinea pigs after exposure to antigen challenge was markedly inhibited by epinastine. The drug was effective in inhibiting not only Ca2+ uptake into lung mast cells in actively sensitized guinea pigs but also Ca2+ release from the intracellular Ca store of rat peritoneal mast cells exposed to both compound 48/80 and substance P. No significant changes were observed in phosphodiesterase activity in rat peritoneal mast cells treated with epinastine, while adenylate cyclase activity was augmented by epinastine. Epinastine has no inhibitory effect on histamine release induced by Ca2+ or IP3 from permeabilized mast cells. However, the drug significantly and dose-dependently suppressed calmodulin activity suggesting that histamine release inhibition due to epinastine may be partly attributable to Ca(2+)-calmodulin dependent process(es). The drug caused no visible changes in thermodynamic behavior of lipids, either in order parameter or in differential scanning calorimetry, indicating that the drug has no influence on membrane fluidity.

Adenylyl Cyclases↗

PubMed1992

Antiallergic effect of epinastine (WAL 801 CL) on immediate hypersensitivity reactions: (II). Antagonistic effect of epinastine on chemical mediators, mainly antihistaminic and anti-PAF effects.

Anti-histamine and anti-PAF effects of epinastine were tested in rats, guinea pigs and rabbits. Epinastine showed a potent histamine H1-blocking effect, but the potency was slightly less than that of ketotifen in histamine-induced contraction of guinea pig ileum and histamine-induced cutaneous reactions in rats. In histamine-induced dye leakage into the nasal cavity tested in rats, the drug was slightly more potent than ketotifen and azelastine. Epinastine as well as ketotifen suppressed rabbit platelet aggregation induced by PAF at higher concentrations compared with WEB 2086, a specific PAF-antagonist. In the bronchospasm induced by PAF in guinea pigs, epinastine was more effective than ketotifen in inhibiting the bronchoconstriction, while it showed no remarkable effect on the hypotension induced by PAF. Epinastine caused a potent antagonistic effect on LTC4-induced contraction of isolated guinea pig trachea. In conclusion, the potent anti-histamine, anti-PAF and anti-LT effects of epinastine may significantly contribute to its antiallergic activity.

Animals↗

PubMed1991

Involvement of the microtubular system in the endothelin-1 secretion from porcine aortic endothelial cells.

The effects of certain microtubule-disrupting agents on endothelin-1 (ET-1) secretion from porcine aortic endothelial cells were studied. When endothelial cells were treated with thrombin (1 unit/mL), a significant increase in ET-1 secretion was detected in the incubation medium, while ET-1 secretion in the medium was diminished when the cells were treated simultaneously with either colchicine or vinblastine (10(-8)-10(-6) M). In such cases, however, the ET-1 content detected in the cells increased dose-dependently in accordance with the concentrations of the microtubule-disrupting agents. The intracellular accumulation of ET-1 was observed both in mitochondrial and microsomal fractions. On the other hand, thrombin produced a significant increase in polymerized tubulin content without affecting the total tubulin content. A thrombin-induced increase in the intracellular Ca2+ concentration of endothelial cells was inhibited by treatment with either colchicine or vinblastine. These results seem to indicate that the microtubular system may play an important role in ET-1 secretion from endothelial cells.

Animals↗

PubMed1991

Comparison of the amino acid and nucleotide sequences between human and two guinea pig major basic proteins.

By means of reverse-phase HPLC, 2 different proteins were obtained from apparently purified pig eosinophil major basic protein (MBP) and these proteins were named GMPB1 and GMBP2. It was revealed that these 2 components of MBP have similar molecular weights and pI values, although the amino acid compositions were slightly different. In the previous study, we cloned and sequenced GMPB1 cDNA. Here we obtained another clone by plaque hybridization using a screening probe synthesized by means of polymerase chain reaction. After sequencing, it became apparent that this clone corresponded to GMBP2. As in the case of GMBP1, the cDNA of GMBP2 encoded pre-proGMBP2 with 3 domains; signal peptide, acidic pro-portion, and mature GMBP2. By comparing the sequences of GMBP1 and GMBP2, it was revealed that the proteins were quite similar to each other. In addition, their sequences also resembled those of human MBP, especially in the basic domain of mature protein; but no such similarity existed in the pro-portion. Although the molecular weights determined by SDS-PAGE of guinea pig and human MBPs were 11,000 and 9,300, respectively, the calculated molecular weights of these 3 MBPs were all 13.8 kDa. The calculated pI values of GMBP1, GMBP2 and human MBP were 11.7, 11.3 and 11.6, respectively. By means of Harr plot analysis, it was revealed that the amino acid sequences, not only in signal peptides but also in the basic domains of mature proteins, were well conserved between guinea pig and human MBPs.

Amino Acid Sequence↗

PubMed1991

Sequencing and cloning of the cDNA of guinea pig eosinophil major basic protein.

Major basic protein (MBP) purified from guinea pig eosinophils elicited histamine release from rat peritoneal mast cells at concentrations higher than 3 micrograms/ml both in the presence and in the absence of extracellular Ca2+. After reverse-phase high-performance liquid chromatography, it was revealed that MBP was composed of two different proteins with quite similar molecular weights and pI values, although the amino acid compositions were slightly different. The partial amino acid sequence of one of these MBPs was determined and the primers for the polymerase chain reaction (PCR) were synthesized according to the partial amino acid sequence. Using these primers and the cDNAs obtained from guinea pig eosinophils, the PCR was carried out in order to synthesize the hybridization probe of MBP for screening the cDNA library. After screening with 8 x 10(5) clones, a positive clone, which encoded a full length of pre-proMBP, was obtained. According to the sequencing data of this clone, it was revealed that pre-proMBP was composed of 3 domains; signal peptide, acidic domain and mature MBP. The predicted pI value of mature MBP was 11.7, though that of proMBP was 7.8. The homology in the amino acid sequence between guinea pig proMBP and human proMBP was 49.4%, while guinea pig mature MBP was more homologous (58%) to human mature MBP.

Amino Acid Sequence↗

PubMed1991

Role of the cytoskeleton in Ca2+ release from the intracellular Ca store of rat peritoneal mast cells.

In order to study the role of the cytoskeleton in histamine release from mast cells, the effects of cytochalasin D, cholchicine and vinblastine on Ca2+ release from the intracellular Ca store induced by compound 48/80 were investigated by means of a video-intensified microscopy system. When the quin 2-loaded mast cells were stimulated by 0.35 micrograms/ml of compound 48/80, a rapid increase in intracellular Ca2+ was observed. At concentrations higher than 10(-6) M, both colchicine and vinblastine pretreatments significantly inhibited the increase in intracellular Ca2+ concentrations caused by compound 48/80, although cytochalasin D had no effect. When permeabilized mast cells were exposed to potassium-antimonate solution, microtubules became attached to the endoplasmic reticulum, where many dots of Ca-antimonate were observed; in some areas, the microtubules interconnected the endoplasmic reticulum and granules in the mast cells. From the results of the present study, it was assumed that microtubules play some important role in the processes leading to Ca2+ release from the intracellular Ca store.

Aminoquinolines↗

PubMed1991

The existence of filaments connecting the granules and the cell membrane in rat peritoneal mast cells.

SEM images of rat peritoneal mast cells showed that microtubule-like filaments seemed to penetrate the granules and project several branches just beneath the granular surface. Each granule appeared to be a cluster of microgranules. TEM observations frequently revealed microtubules connecting the granule surface to the subplasmalemmal network or to the adjacent granule surface and on some occasions, microtubules also appeared to penetrate the granule membranes. Moreover, TEM revealed that individual granules seemed to consist of a mass of microgranules, each 10-15 nm in diameter; freeze-fracture images showed similar configurations. These findings suggest that the microtubule-like filamentous structures in mast cells play a role corresponding to that of the open canalicular system in platelets.

Animals↗

PubMed1991

Role of cytoskeletons on Ca2+ release from the intracellular Ca store of rat peritoneal mast cells.

Cytochalasin D, colchicine or vinblastine effectively inhibited both histamine release and 45Ca uptake induced by compound 48/80 in rat mast cells. The inhibitory effects of cytochalasin D or colchicine on histamine release were exerted more remarkably when permeabilized mast cells were stimulated with either Ca2+ or inositol-1,4,5-trisphosphate (IP3). Since colchicine, vinblastine or cytochalasin D were not effective in inhibiting IP3 formation, it was assumed that microtubules or microfilaments may not participate in the initial stages of the membrane events leading to histamine release. By contrast, in Ca2+ release from the intracellular Ca store both colchicine and vinblastine (but not cytochalasin D) were effective in inhibiting Ca2+ mobilization, indicating that microtubules, rather than microfilaments, are intimately related to Ca2+ release from the endoplasmic reticulum (ER). By means of a fluorescence microscope, it was revealed that colchicine decreased the fluorescence intensity of FITC-labeled anti-tubulin antibody in the mast cells, while the amount of tubulin polymer in mast cells increased after exposure to compound 48/80. The findings indicate that colchicine simply suppressed polymerization of tubulin, while the rearrangement of microtubules so as to increase the polymerization took place after exposure to compound 48/80. Using an electron microscope in combination with potassium antimonate technique, Ca-antimonate dots were clearly observed in a cluster on the surface of the ER and a distinct connection between the ER and microtubules was also observed. It was concluded that microtubules play an important role in the processes leading to Ca2+ release from the intracellular Ca store and in subsequent histamine release.

Animals↗

PubMed1991

Inhibitory effect of MY-1250 on histamine release from rat peritoneal mast cells and guinea pig lung fragments: the elucidation of the mechanism.

When the effect of MY-1250 (5,6-dihydro-7,8-dimethyl-4,5-dioxo-4 H-pyrano [3,2-c] quinoline-2-carboxylic acid) on histamine release from rat peritoneal mast cells induced by compound 48/80 was studied, MY-1250 caused a significant inhibition of histamine release at concentrations higher than 3 microM. Furthermore, the compound inhibited not only 45C a uptake into the mast cells but also Ca2+ release from the intracellular Ca store at a concentration of 10 microM in both cases. By contrast, MY-1250 did not affect either histamine release from permeabilized mast cells induced by TPA, IP3 and GTP gamma S or Ca2+ release from the endoplasmic reticulum induced by IP3. In the chopped lung preparations, MY-1250 at doses of 10 and 100 microM caused a significant inhibition in histamine release from the pieces of actively sensitized guinea pigs exposed to antigen and simultaneously prevented a decrease in intracellular cAMP contents taking place in association with antigen-antibody reaction. No significant changes were effected by MY-1250 in alpha-chymotrypsin activity and phospholipase A2 activity. Also, no antagonistic effects on LTD4 and PAF were observed.

Animals↗

PubMed1991

Antiallergic effects of major metabolites of astemizole in rats and guinea pigs.

Antiallergic effects of astemizole (CAS 68844-77-9) and its metabolites were studied using rats and guinea pigs. All the metabolites of astemizole tested i.e., desmethylastemizole, 6-hydroxydesmethylastemizole and norastemizole, were more active than astemizole in inhibiting the contraction of the ileum as well as the bronchoconstriction induced by histamine in guinea pigs. Desmethylastemizole was about the same as the parent compound in inhibiting the 3H-mepyramine binding in guinea pig cerebellum. In heterologous passive cutaneous anaphylaxis (PCA) and homologous PCA, the metabolites caused almost equipotent inhibition to that seen in astemizole. On the other hand, in the studies of histamine release from rat peritoneal mast cells induced by compound 48/80 or from lung fragments in actively sensitized guinea pigs, desmethylastemizole, 6-hydroxydesmethylastemizole and norastemizole were much less potent than was astemizole. No H2-antagonistic activity was observed with either astemizole or desmethylastemizole.

Animals↗

PubMed1991

Comparative study of the adverse effects of various radiographic contrast media, including ioversol, a new low-osmolarity medium. II. The complement system and endothelial cells.

The effects of ioversol, iohexol, iopamidol and meglumine sodium amidotrizoate (MSA) on the complement system and endothelial cells were investigated. The protein bindings of the radiographic contrast media (RCM), each tested with guinea pig plasma, were less than 1%. When guinea pig serum was incubated with any of the RCM, activation of the complement system, which leads to hemolysis, was not influenced by the nonionic agents, ioversol, iohexol or iopamidol. However, MSA, an ionic agent, significantly reduced hemolytic activity at 370 mgI/ml. Perfusion of the abdominal aorta with nonionic agents did not elicit significant endothelial damage; ioversol induced the least damage among the nonionic RCM, while MSA caused remarkable endothelial damage. Although MSA caused a marked release of endothelin-1 from cultured endothelial cells obtained from porcine aorta, nonionic RCM did not induce significant endothelin-1 release; no influence was elicited by ioversol and iohexol caused a weak suppression, while iopamidol had the opposite effect. These results indicate that ioversol could be used as a safe contrast medium in intravascular administration.

Animals↗