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Query: UNIPROT:P15088 (mast cell)
 14,925 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Rat peritoneal mast cells (RPMC) and rat basophilic leukemia (RBL) cells are representative of connective tissue-type (CTMC) and mucosal-type (MMC) mast cells, respectively. Using polyethylene glycol, we have fused RPMC with 6-thioguanine resistant, HAT (hypoxanthine, aminopterin, thymidine) sensitive RBL-CA10.7 or RBL-CK2 cells, yielding several hybrid rat mast cell lines (HRMC). The hybridomas exhibited different size and cytoplasmic granularity when compared with parental cell lines. Analysis of both high (Fc epsilon RI) and low affinity (Fc epsilon RL) receptors for IgE revealed that the hybrid lines had more variable receptor patterns than the parent lines. Three hybridoma lines were chosen for further study. Differential histochemical staining with alcian blue and safranin O dyes indicated the hybrids to be predominantly of the MMC type: however, a few cells of one of these uncloned hybridomas were found to be of the CTMC type. Attempts to isolate the CTMC hybridomas yielded one culture which was predominantly of the CTMC phenotype and in a number of other cultures, cells were found expressing simultaneously both the CTMC and the MMC phenotype. After 3 weeks in culture, however, all hybridomas, including those which were cloned further, expressed only the MMC histochemical phenotype. This was found to correlate with the presence of rat mast cell protease II (RMCPII) and the absence of RMCPI in all hybridomas, as detected by Western blot analysis. In addition, the histamine content of all cells was significantly lower than that of the parent RPMC. Most hybrid mast cells expressed both Fc epsilon RI and Fc epsilon RL which in some cases exhibited significant variations in the Mr. These results indicate that somatic cell hybrids expressing the MMC and CTMC phenotype can be produced by the fusion of RBL and RPMC. The CTMC phenotype, however, is unstable, and possible reasons for this are discussed.
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PMID:Establishment and characterization of hybrid rat mast cells. 182 10

In this study, galactose dehydrogenase (EC 1.1.1.48) was chosen as a prototype target protein to investigate the capability of metal affinity precipitation to facilitate the purification of genetically engineered proteins. A DNA fragment encoding five histidine residues was fused to the 3'-terminal end of the galactose dehydrogenase gene from Pseudomonas fluorescens and thereafter expressed in Escherichia coli. The additional five histidines functioned as an affinity tail and the modified enzyme could be purified using metal affinity precipitation when the metal-chelate complex with ethylene glycol-bis-(beta-aminoethyl ether) N,N,N',N'-tetra-acetic acid, EGTA(Zn)2, was added to the protein solution. The affinity tail could also be applied for the purification of the fusion protein utilising immobilised metal affinity chromatography. After purification, the pentahistidine affinity tail could be removed enzymatically by carboxypeptidase A. Furthermore, growth rate experiments demonstrated that the expression of the metal-binding affinity tail in E. coli cells enhanced the tolerance to zinc ions when added to the growth medium.
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PMID:Metal affinity precipitation of proteins carrying genetically attached polyhistidine affinity tails. 190 25

The activity of polyethylene glycol 400, a widely used drug solvent, was tested on the release of histamine induced by adriamycin in vitro on peritoneal rat mast cells and in vivo in a mouse model. Preincubation of mast cells with high (10 and 5%) concentrations of polyethylene glycol 400 significantly inhibited the important histamine release induced by 100 micrograms/ml of adriamycin; furthermore, polyethylene glycol 400 (3.45 g/kg; 0.345 g/ml) pretreatment almost completely abolished the peritoneal and pericardial mast cell degranulation and the cardiac toxicity caused by an intraperitoneal injection of 15 mg/kg of adriamycin. This effect of polyethylene glycol 400 on adriamycin-induced histamine release could explain the protective action exhibited in vivo on adriamycin treated animals, therefore confirming that adriamycin cardiotoxicity could be related to the release of histamine and other vasoactive substances.
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PMID:Effect of polyethylene glycol 400 on adriamycin induced histamine release. 242 41

1. Stimulation of mast cells by externally applied secretagogues activated a slowly developing membrane current. With high external and low internal chloride (Cl-) concentrations, the current reversed at about -40 mV, but when external Cl- was made equal to internal Cl-, the reversal potential shifted to about 0 mV, demonstrating that the current carrier was Cl-. 2. In addition to external agonists, internally applied cyclic AMP and high concentrations of intracellular calcium [Ca2+]i could also activate the Cl- current. However, elevated [Ca2+]i produced only slow and incomplete activation. This suggests that the Cl- current is not directly Ca2+ activated. Also, activation of Cl- current by external agonists and by cyclic AMP was unimpaired when [Ca2+]i was clamped to low levels with internal ethylene glycol bis-N,N,N',N'-tetraacetic acid (EGTA), indicating that elevated [Ca2+]i is not necessary for activation of the Cl- current. Although activation by cyclic AMP was faster than that produced by elevated [Ca2+]i, it still required tens of seconds; thus the effect of cyclic AMP was also likely to be indirect. 3. Internal guanosine 5'-O-(3-thiotriphosphate) (GTP-gamma-S) could also activate the Cl- current, suggesting the involvement of a G protein in the control of the current. 4. The variance associated with the Cl- current was small, and noise analysis gave a lower limit of about 1-2 pS for the single-channel conductance. The Cl- current was reduced by 4,4'-diisothiocyano-2,2'-stilbenedisulphonate (DIDS), and during DIDS blockade, the variance of the current increased. This suggests that DIDS enters and blocks the open channel. 5. Activation of the Cl- current would make the membrane potential negative following stimulation of a mast cell, thus providing a driving force for entry of external calcium via the stimulation-induced influx pathways described in the preceding paper (Matthews, Neher & Penner, 1989).
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PMID:Chloride conductance activated by external agonists and internal messengers in rat peritoneal mast cells. 255 69

The presynaptically active snake venom neurotoxin beta-bungarotoxin (beta-Butx) is known to affect neurotransmitter release by binding to a subtype of voltage-activated K+ channels. Here we show that mast cell degranulating (MCD) peptide from bee venom inhibits the binding of 125I-labeled beta-Butx to chick and rat brain membranes with apparent Ki values of 180 nM and 1100 nM, respectively. The mechanism of inhibition by MCD peptide is noncompetitive, as is inhibition of 125I-beta-Butx binding by the protease inhibitor homologue from mamba venom, toxin I. Beta-Butx and its binding antagonists thus bind to different sites of the same membrane protein. Removal of Ca2+ by ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid inhibits the binding of 125I-beta-Butx by lowering its affinity to brain membranes.
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PMID:Inhibition of beta-bungarotoxin binding to brain membranes by mast cell degranulating peptide, toxin I, and ethylene glycol bis (beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. 313 91

IgE anti-DNP antibody-secreting hybridomas were obtained by polyethylene glycol fusion of murine myeloma cells with spleen cells from mice, primed with DNP-KLH and challenged with DNP-Nippostrongylus brasiliensis extract. The in vivo and in vitro continuously growing hybridomas are producing high amounts of monoclonal IgE anti-DNP antibodies, which are heat-labile, hapten-specific and have rat mast cell sensitizing capacity.
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PMID:Production of monoclonal mouse IgE antibodies with DNP specificity by hybrid cell lines. 615 72

We have previously shown that the weakly fluorescent cationic dye, berberine, forms a strongly fluorescent complex with the heparin of fixed mast cells and can be used for cytofluorometric measurement of heparin in both mast cells and individual mast cell granules. Now, we report on the use of berberine as a vital stain, demonstrating the secretory activity of mast cells. At a dye concentration of 0.025%, and after membrane stabilization with polyethylene glycol, normal mast cells exclude the dye while mast cells stimulated to secretion with polymyxin B show a strongly fluorescent dye binding to individual cytoplasmic granules. The mean fluorescence intensity (reflecting the number of stained granules) of the cell populations increased with increasing polymyxin B concentrations up to 2.0 micrograms/ml, thereafter remaining constant up to 10 micrograms/ml. Fluorescence intensity after vital staining with berberine was compared both to release of heparin measured by berberine binding to fixed cells (reflecting exocytosis of mast cell granules) and also to histamine release. The results strongly suggest that berberine specifically stains secreting granules that are located within the domains of the cells but may have released histamine. Vital berberine staining combined with histamine assays and cytofluorometric measurements of heparin is therefore of great potential interest for analyzing the dynamics of mast cell secretion.
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PMID:Fluorescent berberine binding as a marker of secretory activity in mast cells. 619 Jul 61

The fluorescent cationic dye berberine in combination with histamine release studies have been used to explore the different steps of the mast cell secretory process. We have previously shown that quantitation of heparin release by the binding of berberine to fixed mast cells can be used as a direct measure of release of granules. This report summarizes recent work using berberine as a vital stain demonstrating the secretory activity of mast cells. After membrane stabilization with polyethylene glycol (PEG) normal mast cells exclude the dye while mast cells stimulated to secretion with polymyxin B show a strongly fluorescent dye binding to individual cytoplasmic granules. The mean fluorescence intensity of the cell populations after vital berberine staining was compared both to heparin and histamine release. The results strongly suggest that berberine, under the vital staining conditions used, is a marker of intracellular granules that have released histamine. The vital staining method was also used to study membrane events following a polymyxin B-induced secretion. The mean fluorescence intensity decreased by 75% during the first hour after the termination of a polymyxin B stimulation while the mast cell content of histamine and heparin remained constant. The findings support the idea that the membranes are rapidly restored after mast cell secretion, permitting a selective amine release without accompanying release of heparin or other matrix components of the granules.
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PMID:The dynamics of mast cell secretion studied by vital berberine staining. 620 57

The specific IgE response that appears in subjects immunized with tetanus toxoid does not induce hypersensitivity reactions at subsequent immunizations. The type I immune response, therefore, was studied both in vivo and in vitro, in 11 subjects who had specific IgE antibodies for tetanus toxoid. The results showed that: 1. the specific IgE antibodies are heterogeneous regarding their affinity for the mast cell and basophil receptors; 2. the specific IgG antibodies for tetanus toxoid, at serum concentrations, are not able to interfere with the in vitro specific basophil degranulation; 3. in the PEG precipitate there are aggregates of specific IgE antibodies for tetanus toxoid. In vitro, these molecular aggregates are not able to sensitize the basophil cells.
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PMID:Functional characterization of specific IgE antibodies for tetanus toxoid. 662 28

Rat peritoneal mast cells and 6-thioguanine-resistant rat basophilic leukemia cells, representative of connective tissue-type (CTMC) and mucosal (MMC) mast cells, respectively, were fused using polyethylene glycol. Four out of 14 primary hybrid mast cell lines contained more than 50% of CTMC as demonstrated by histochemical staining. Two cell lines, one predominantly of the CTMC and the other of the MMC phenotype, were selected for further study. Among these, the phenotype was also confirmed by analysis for rat mast cell protease I and by mediator release triggered by compound 48/80 and ionophore A23187. The CTMC phenotype disappeared after culturing cells for 2 weeks. The change in phenotype did not significantly alter the mediator release due to calcium ionophore A23187. Repeated cloning of cells bearing the CTMC phenotype did not yield a cloned line of cells expressing the CTMC phenotype only, although it prolonged the persistence of this phenotype. During the period of CTMC phenotype loss, a drop in cellular DNA content occurred, suggesting that chromosome instability may, at least partially, have been responsible for the phenotypic changes.
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PMID:Phenotypic changes among hybrid rat mast cells. 758 Feb 87


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