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Results for “Functional characterization”

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PubMed1996

Cloning and functional characterization of human heteromeric N-methyl-D-aspartate receptors.

Human cDNAs encoding N-methyl-D-aspartate receptor type (NMDAR)1A, NMDAR2A and NMDAR2B subunits were cloned and receptors encoded by these cDNAs were functionally expressed by injection of the respective mRNAs in Xenopus oocytes. The pharmacological properties of recombinant human N-methyl-D-aspartate (NMDA) receptors were characterized by profiling two agonists and four antagonists at both the NMDA and glycine sites in voltage-clamped oocytes. NMDA, glycine and D-serine were significantly more potent at human NMDAR (hNMDAR)1A/2B receptors than at nNMDAR1A/2A, whereas there was no detectable subtype-dependent difference in the potency of glutamate. Of the NMDA-site antagonists tested, CGP 43487 and 3-(2-carboxypiperazin-4-yl) propyl-1-phosphonate exhibited 5.8- and 3.9-fold greater potency, respectively, at hNMDAR1A/2A receptors than at hNMDAR1A/2B. Of the four glycine-site competitive antagonists tested, L-689,560 displayed 5-fold greater potency at hNMDAR1A/2A, whereas 5,7-dichlorokynurenic acid, HA-966 and CGP 58411 did not discriminate between hNMDAR1A/2A and hNMDAR1A/2B. Receptors resulting from injection of hNMDAR1A, hNMDAR2A and hNMDAR2B transcripts in a 1:1:1 ratio were indistinguishable from hNMDAR1A/2B receptors in terms of their sensitivity to NMDA, glycine, D-serine, CGS 19755 and CGP 40116. Ifenprodil was approximately 350-fold more potent at hNMDAR1A/2B than at hNMDAR1A/2A receptors. Ifenprodil sensitivities of receptors formed in oocytes injected with a constant amount of hNMDAR1A mRNA but varying ratios of hNMDAR2A or hNMDAR2B mRNAs were compared. The receptors expressed at a 10:1 ratio of 2A:2B transcripts displayed an ifenprodil sensitivity that would be predicted for a population in which 51% was represented by hNMDAR(1A)2(2A)3 complexes. Our results underscore the need for subtype-selective compounds acting at novel sites to sufficiently probe the pharmacological differences between NMDA receptor subtypes formed by different subunit combinations.

Animals↗

PubMed2006

Expression and functional characterization of the putative protein 8b of the severe acute respiratory syndrome-associated coronavirus.

SARS 8b is one of the putative accessory proteins of the severe acute respiratory syndrome-associated coronavirus (SARS-CoV) with unknown functions. In this study, the cellular localization and activity of this estimated 9.6 kDa protein were examined. Confocal microscopy results indicated that SARS 8b is localized in both nucleus and cytoplasm of mammalian cells. Functional study revealed that overexpression of SARS 8b induced DNA synthesis. Coexpression of SARS 8b and SARS 6, a previously characterized SARS-CoV accessory protein, did not elicit synergistic effects on DNA synthesis.

Animals↗

PubMed2004

Functional characterization of the human SOX3 promoter: identification of transcription factors implicated in basal promoter activity.

SRY-related HMG-box genes (Sox genes) constitute a large family of developmentally regulated genes involved in the decision of cell fates during development and implicated in the control of diverse developmental processes. Sox3, an X-linked member of the family, is expressed in the central nervous system (CNS) from the earliest stages of development. It is considered to be one of the earliest neural markers in vertebrates playing the role in specifying neuronal fate. The aim of this study has been to determine and characterize the promoter of the human SOX3 gene and to elucidate molecular mechanisms underlying the regulation of its expression. In this study, we have isolated and performed the first characterization of the human SOX3 promoter. We have identified the transcription start point (tsp) and carried out the structural and functional analysis of the regulatory region responsible for SOX3 expression in NT2/D1 cell line. Using promoter-reporter constructs, we have determined the minimal SOX3 promoter region that confers the basal promoter activity, as well as two regulatory elements which have positive effects on the promoter activity. We have investigated in detail the functional properties of three conserved motifs within the core promoter sequence that bind transcription factors specificity protein 1 (Sp1), upstream stimulatory factor (USF) and nuclear factor Y (NF-Y). By mutational analysis, we have shown that all three sites are of functional relevance for constitutive SOX3 expression in NT2/D1 cells. We have also shown that, besides the TATA motif, at least one other essential regulatory element is required for the basal transcription of the human SOX3. Taken together, data presented in this paper suggest that transcription factors such as Sp1, USF and NF-Y could function as key regulators for the basal activation of the human SOX3 gene.

Animals↗

PubMed2000

Molecular and functional characterization of porcine LFA-1 using monoclonal antibodies to CD11a and CD18.

We describe in this report the production and characterization of monoclonal antibodies (mAb) to the swine homologues of CD11a and CD18 antigens, and their use for phenotypic and functional analysis of porcine leukocytes. Monoclonal antibodies BL1H8 and BL2F1 precipitated two bands of approximately 170 and 95 kDa, whereas mAb BA3H2 brought down three bands of 170, 155 and 95 kDa, from alveolar macrophage lysates. Clearance of macrophage lysates with mAbs BL1H8 and BL2F1 resulted in complete removal of the 170-kDa band. The cell distribution of the molecules recognized by these mAbs was similar to that of human LFA-1. It was found on all leukocytes, although its expression varied among the different leukocyte subpopulations, with monocytes, granulocytes and a subset of CD8+ cells expressing the highest levels. Cross-blocking studies showed that these antibodies recognize different epitopes on porcine LFA-1. Both anti-LFA-1 mAbs strongly inhibited the mitogenic response of PBMC to ConA, whereas the anti-CD18 mAb had no effect. These anti-LFA-1 mAbs also inhibited the mixed lymphocyte reaction (MLR) and the NK cell-mediated lysis of K-562 cells.

Animals↗

PubMed2000

Functional characterization of a mammalian Sac1 and mutants exhibiting substrate-specific defects in phosphoinositide phosphatase activity.

The Saccharomyces cerevisiae SAC1 gene was identified via independent analyses of mutations that modulate yeast actin function and alleviate the essential requirement for phosphatidylinositol transfer protein (Sec14p) activity in Golgi secretory function. The SAC1 gene product (Sac1p) is an integral membrane protein of the endoplasmic reticulum and the Golgi complex. Sac1p shares primary sequence homology with a subfamily of cytosolic/peripheral membrane phosphoinositide phosphatases, the synaptojanins, and these Sac1 domains define novel phosphoinositide phosphatase modules. We now report the characterization of a rat counterpart of Sac1p. Rat Sac1 is a ubiquitously expressed 65-kDa integral membrane protein of the endoplasmic reticulum that is found at particularly high levels in cerebellar Purkinje cells. Like Sac1p, rat Sac1 exhibits intrinsic phosphoinositide phosphatase activity directed toward phosphatidylinositol 3-phosphate, phosphatidylinositol 4-phosphate, and phosphatidylinositol 3,5-bisphosphate substrates, and we identify mutant rat sac1 alleles that evoke substrate-specific defects in this enzymatic activity. Finally, rat Sac1 expression in Deltasac1 yeast strains complements a wide phenotypes associated with Sac1p insufficiency. Biochemical and in vivo data indicate that rat Sac1 phosphatidylinositol-4-phosphate phosphatase activity, but not its phosphatidylinositol-3-phosphate or phosphatidylinositol-3, 5-bisphosphate phosphatase activities, is essential for the heterologous complementation of Sac1p defects in vivo. Thus, yeast Sac1p and rat Sac1 are integral membrane lipid phosphatases that play evolutionary conserved roles in eukaryotic cell physiology.

Amino Acid Sequence↗

PubMed2003

Molecular and functional characterization of SLC26A11, a sodium-independent sulfate transporter from high endothelial venules.

Lymphocyte emigration from the blood into most secondary lymphoid organs and chronically inflamed tissues occurs at the level of high endothelial venules (HEV). A unique characteristic of HEV endothelial cells (HEVEC) is their capacity to incorporate large amounts of sulfate into sialomucin-type counter-receptors for the lymphocyte homing receptor L-selectin. We have previously shown that sulfate uptake into HEVEC is mediated by two distinct functional classes of sulfate transporters: Na+-coupled transporters and sulfate/anion exchangers. Here, we report the molecular characterization from human HEVEC of SLC26A11, a novel member of the SLC26 sulfate/anion exchanger family. Functional expression studies in COS-7 and Sf9 insect cells revealed that SLC26A11 is targeted to the cell membrane and exhibits Na+-independent sulfate transport activity, sensitive to the anion exchanger inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). Northern blot analysis showed the highest SLC26A11 transcript levels in placenta, kidney, and brain. The SLC26A11 gene mapped to human chromosome 17q25, very close to the hereditary hearing loss diseases loci DFNA20, DFNA26, and USH1G. RT-PCR analysis of SLC26 sulfate transporters in human HEVEC revealed coexpression of SLC26A11 with SLC26A2/DTDST and lack of SLC26A1/SAT1, SLC26A3/DRA, and SLC26A8/TAT1. Together, our results indicate that SLC26A11 is a novel Na+-independent sulfate transporter that may cooperate with SLC26A2 to mediate DIDS-sensitive sulfate uptake into HEVEC.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1995

Functional characterization of lymphoid cells generated in serum-deprived culture stimulated with stem cell factor and interleukin 7 from normal and autoimmune mice.

We have investigated the phenotypic and functional characteristics of murine pre-B cells obtained in semisolid and liquid culture with stem cell factor (SCF) and interleukin 7 (IL-7). Both serum-supplemented and serum-deprived culture conditions were used. The source of bone marrow cells was either normal mice (CD1 and C3H) or the lupus strain of mice MRL/lpr and its congenic strain MRL/+. SCF (100 ng/ml) and IL-7 (250 ng/ml) supported murine B cell proliferation in vitro from all the murine strains analyzed both in serum-supplemented and serum-deprived conditions. Maximal colony growth was observed in both cases when the factors were used in combination. The growth factors alone induced some colony growth in serum-supplemented cultures but were either ineffective or had modest activity in serum-deprived cultures. Cells harvested from the colonies or generated in liquid cultures and stimulated with SCF + IL-7 in the absence of serum had almost exclusively a pre-B cell phenotype (BP-1+, B220+, slg-, CD4-, CD8-, Mac-1-, RB-6-). Both the maximal colony growth in semisolid culture and the maximal number of cells in liquid culture were observed at day 12-14. At this time, the pre-B cells failed to differentiate further and started to die. Pre-B cells generated in vitro were, however, capable of differentiating in vivo. SCID mice injected with 2 x 10(6) pre-B cells had readily detectable serum levels of IgM (54 +/- 26 micrograms/ml) and IgG (60 +/- 95 micrograms/ml) at 4 weeks and 6 weeks posttransplantation, respectively. Mature B and T cells of the donor major histocompatibility complex type were detected in the SCID mice at sacrifice 14 weeks posttransplantation. These data indicate that purified (> 80% BP-1+) populations of functional pre-B cells can be grown from murine bone marrow of normal mice as well as of lupus mice in serum-deprived cultures stimulated with SCF and IL-7. These cultures, therefore, provide a highly enriched source of pre-B cells but also contain T cell precursors that differentiate upon adoptive transfer into SCID mice.

Animals↗

PubMed2000

Expression and functional characterization of human protein X variants in SV40-immortalized protein X-deficient and E2-deficient human skin fibroblasts.

To gain further insight into the nature and function of the domains of the human protein X (a pyruvate dehydrogenase complex component also known as the E3-binding protein), we expressed the wild-type as well as two artificially created variants, K37E and S422H, in SV40-immortalized protein X-deficient and E2-deficient human skin fibroblasts. The former mutant does not carry the lipoic acid moiety, the latter mutant was designed to investigate the possibility that protein X could exhibit an intrinsic acetyltransferase activity and use either its own catalytic center or the catalytic center of E2. Similar experiments have been performed in the past using the Saccharomyces cerevisiae expression system. However, lack of sequence similarity between the mammalian and the yeast protein X homologues suggests they are not biochemically equivalent. Mutant cells transfected with the wild-type gene for protein X produced a PDH complex that exhibited about 50% overall activity of the control cells. None of the expressed protein X variants had an effect on the specific activity of the PDH complex, suggesting that the human protein X plays a purely structural role in the functioning of the pyruvate dehydrogenase complex.

Amino Acid Sequence↗

PubMed2001

Structural and functional characterization of human NAD kinase.

NADP is essential for biosynthetic pathways, energy, and signal transduction. Its synthesis is catalyzed by NAD kinase. Very little is known about the structure, function, and regulation of this enzyme from multicellular organisms. We identified a human NAD kinase cDNA and the corresponding gene using available database information. A cDNA was amplified from a human fibroblast cDNA library and functionally overexpressed in Escherichia coli. The obtained cDNA, slightly different from that deposited in the database, encodes a protein of 49 kDa. The gene is expressed in most human tissues, but not in skeletal muscle. Human NAD kinase differs considerably from that of prokaryotes by subunit molecular mass (49 kDa vs 30-35 kDa). The catalytically active homotetramer is highly selective for its substrates, NAD and ATP. It did not phosphorylate the nicotinic acid derivative of NAD (NAAD) suggesting that the potent calcium-mobilizing pyridine nucleotide NAADP is synthesized by an alternative route.

Animals↗

PubMed2003

Functional characterization of IL-13 receptor alpha2 gene promoter: a critical role of the transcription factor STAT6 for regulated expression.

Interleukin (IL)-4 and IL-13 are two structurally and functionally related cytokines that have overlapping but also distinct biological activities. One of the components of the IL-13 receptor, the alpha2 chain (IL-13Ralpha2), has been reported to downregulate the cell responsiveness to IL-13, without affecting IL-4 signaling. Here, we report that TNFalpha synergizes with either IL-4 or IL-13 in inducing the IL-13Ralpha2 chain at both the mRNA and protein levels in the HaCaT human keratinocyte cell line. Further studies by 5'RACE identified as yet undescribed exonic sequences of the IL-13Ralpha2 5'UTR, provided evidence for the expression of alternatively spliced IL-13Ralpha2 transcripts and defined the transcription start of the IL-13Ralpha2 gene. A 1.5 kb region upstream of the first exon of the IL-13Ralpha2 gene displayed basal promoter activity when inserted in a reporter plasmid and transiently transfected in HaCaT cells. This promoter activity was further increased in response to IL-4 and IL-13. Furthermore, by electrophoretic mobility shift assay and site-directed mutagenesis, we showed that the IL-4/IL-13-induced promoter activity depended upon a positively acting STAT6 response element. Finally, TNFalpha was shown to potentiate IL-4/IL-13-induced IL-13Ralpha2 promoter activity when the same reporter construct was studied in stably but not in transiently transfected cells. These results suggest that the synergistic effect of TNFalpha on IL-4/IL-13-induced IL-13Ralpha2 expression is dependent upon chromatin re-modeling events.

5' Flanking Region↗

PubMed2000

Functional characterization of yeast mitochondrial release factor 1.

The yeast Saccharomyces cerevisiae mitochondrial release factor was expressed from the cloned MRF1 gene, purified from inclusion bodies, and refolded to give functional activity. The gene encoded a factor with release activity that recognized cognate stop codons in a termination assay with mitochondrial ribosomes and in an assay with Escherichia coli ribosomes. The noncognate stop codon, UGA, encoding tryptophan in mitochondria, was recognized weakly in the heterologous assay. The mitochondrial release factor 1 protein bound to bacterial ribosomes and formed a cross-link with the stop codon within a mRNA bound in a termination complex. The affinity was strongly dependent on the identity of stop signal. Two alleles of MRF1 that contained point mutations in a release factor 1 specific region of the primary structure and that in vivo compensated for mutations in the decoding site rRNA of mitochondrial ribosomes were cloned, and the expressed proteins were purified and refolded. The variant proteins showed impaired binding to the ribosome compared with mitochondrial release factor 1. This structural region in release factors is likely to be involved in codon-dependent specific ribosomal interactions.

Amino Acid Sequence↗

PubMed2005

Functional characterization of human monocarboxylate transporter 6 (SLC16A5).

Human monocarboxylate transporter 6 (MCT6) has recently been isolated, and its tissue distribution has been established at the mRNA level, but its functional properties remain unknown. The aim of this study is to investigate the transport properties of MCT6. When expressed in Xenopus laevis oocytes, MCT6 transported [3H]bumetanide in a pH- and membrane potential-sensitive but not proton gradient-dependent manner, with the K(t) value of 84 microM. Furthermore, MCT6 transported various drugs such as probenecid and nateglinide. Neither [14C]L-lactic acid nor [3H]L-tryptophan, typical substrates of other MCT isoforms, was transported by MCT6. Four loop diuretics, i.e., furosemide, piretanide, azosemide, and torasemide, thiazides, probenecid, glibenclamide, and nateglinide inhibited the MCT6-mediated uptake of [3H]bumetanide. In contrast, short-chain carboxylic acids, such as L-lactic acid and succinic acid did not inhibit the MCT6-mediated uptake of bumetanide. These results suggest that the substrate specificity of MCT6 is distinct from those of other MCTs. Bumetanide would be a good tool for investigating the functional properties of MCT6. It is probable that MCT6 is involved in the disposition of various drugs, including bumetanide.

Animals↗

PubMed2003

Functional characterization of polymorphisms in DNA repair genes using cytogenetic challenge assays.

A major barrier to understanding the role of polymorphic DNA repair genes for environmental cancer is that the functions of variant genotypes are largely unknown. Using our cytogenetic challenge assays, we conducted an investigation to address the deficiency. Using X-rays or ultraviolet (UV) light, we irradiated blood lymphocytes from 80 nonsmoking donors to challenge the cells to repair the induced DNA damage, and we analyzed expression of chromosome aberrations (CA) specific to the inducing agents. We have genotyped polymorphic DNA repair genes preferentially involved with base excision repair (BER) and nucleotide excision repair (NER) activities (XRCC1, XRCC3, APE1, XPD) corresponding to the repair of X-ray- and UV light-induced DNA damage, respectively. We expected that defects in specific DNA repair pathways due to polymorphisms would cause corresponding increases of specific CA. From our data, XRCC1 399Gln and XRCC3 241Met were associated with significant increases in chromosome deletions compared with the corresponding homozygous wild types (18.27 1.1 vs 14.79 1.2 and 18.22 0.99 vs 14.20 1.39, respectively); XPD 312Asn and XPD 751Gln were associated with significant increases in chromatid breaks compared with wild types (16.09 1.36 vs 11.41 0.98 and 16.87 1.27 vs 10.54 0.87, respectively), p < 0.05. The data indicate that XRCC1 399Gln and XRCC3 241Met are significantly defective in BER, and the XPD 312Asn and XPD 751Gln are significantly defective in NER. In addition, the variant genotypes interact significantly, with limited overlap of the two different repair pathways.

Adult↗

PubMed1991

Primary structure and functional characterization of a human 5-HT1D-type serotonin receptor.

We describe the nucleic acid sequence encoding a human 5-hydroxytryptamine1D (5-HT1D) serotonin receptor and some of the functional characteristics of the gene product. The receptor gene was isolated by hybridization to a probe based on a canine thyroid cDNA (called RDC4) previously isolated by others and believed to encode a heretofore undetermined member of the guanine nucleotide-binding protein (G protein)-linked receptor family. The human clone we isolated, called MA6A, contains an apparently intronless open reading frame encoding a 377-amino acid polypeptide with the seven hydrophobic domains characteristic of G protein-linked receptors. The MA6A deduced amino acid sequence is 88% identical to that for RDC4 and 43% identical to that for the human 5-HT1A receptor. Expression of the human gene product in transfected cell lines results in the appearance of saturable high affinity 5-HT1D-type [3H]5-HT binding. The expressed receptor exhibits features indicative of coupling to Gi proteins, i.e., robust inhibition of forskolin-stimulated cAMP accumulation and formation of a pertussis toxin-sensitive high agonist affinity binding state. These findings may help clarify several ambiguities in the classification and action of serotonin receptor subtypes.

Amino Acid Sequence↗

PubMed1993

Functional characterization of peripheral blood neutrophils in patients with primary hypothyroidism.

The clinical observations of increased risk of bacterial infection in patients with severe hypothyroidism motivated investigations of the biological function of neutrophils in peripheral blood of 30 women with primary hypothyroidism. The following functions of neutrophils were examined: phagocytosis, adherence to fibers, as well as spontaneous migration of leukocytes. The total number of leukocytes and neutrophils was also determined. The results were compared with those in a control group formed by 20 healthy women. The data show that in patients with primary hypothyroidism, the adherence of neutrophils was increased (63 +/- 8% in patients and 53 +/- 11% in control subjects) and spontaneous migration decreased (45 +/- 20 cm2 in patients and 65 +/- 30 cm2 in the control group). However, there were no significant deviations of phagocytic activity of neutrophils in hypothyroid individuals as compared to euthyroid controls.

Adult↗

PubMed2001

Expression and functional characterization of the mt1 melatonin receptor from rat brain in Xenopus oocytes: evidence for coupling to the phosphoinositol pathway.

Melatonin-sensitive receptors were expressed in Xenopus laevis oocytes following an injection of mRNA from rat brain. The administration of 0.1-100 micromol/L melatonin to voltage-clamped oocytes activates calcium-dependent chloride currents via a pertussis toxin-sensitive G protein and the phosphoinositol pathway. To determine which melatonin receptor type (mt1, MT2, MT3) is functionally expressed in the Xenopus oocytes, we used (i) agonists and antagonists of different receptor types to characterize the pharmacological profile of the expressed receptors and (ii) a strategy of inhibiting melatonin receptor function by antisense oligonucleotides. During pharmacological screening administration of the agonists 2-iodomelatonin and 2-iodo-N-butanoyl-5-methoxytryptamine (IbMT) to the oocytes resulted in oscillatory membrane currents, whereas the administration of the MT3 agonist 5-methoxycarbonylamino-N-acetyltryptamine (GR135,531) exerted no detectable membrane currents. The melatonin response was abolished by a preceding administration of the antagonists 2-phenylmelatonin and luzindole but was unaffected by the MT3 antagonist prazosin and the MT2 antagonist 4-phenyl-2-propionamidotetralin (4-P-PDOT). In the antisense experiments, in the control group the melatonin response occurred in 45 of 54 mRNA-injected oocytes (83%). Co-injection of the antisense oligonucleotide, corresponding to the mt1 receptor mRNA, caused a marked and significant reduction in the expression level (13%; P < 0.001). In conclusion, the results demonstrate that injection of mRNA from rat brain in Xenopus oocytes induced the expression of the mt1 receptor which is coupled to the phosphoinositol pathway.

Animals↗

PubMed2002

Electrochemical and functional characterization of the proline dehydrogenase domain of the PutA flavoprotein from Escherichia coli.

The multifunctional PutA flavoprotein from Escherichia coli is a peripherally membrane-bound enzyme that has both proline dehydrogenase (PDH) and Delta(1)-pyrroline-5-carboxylate dehydrogenase (P5CDH) activities. In addition to its enzymatic functions, PutA displays DNA-binding activity and represses proline catabolism by binding to the control region DNA of the put regulon (put intergenic DNA). Presently, information on structure-function relationships for PutA is derived from primary structure analysis. To gain further insight into the functional organization of PutA, our objective is to dissect PutA into different domains and to characterize them separately. Here, we report the characterization of a bifunctional proline dehydrogenase (PutA(669)) that contains residues 1-669 of the PutA protein. PutA(669) purifies as a dimer and has a PDH specific activity that is 4-fold higher than that of PutA. As anticipated, PutA(669) lacks P5CDH activity. At pH 7.5, an E(m) (E-FAD/E-FADH(-)) of -0.091 V for the two-electron reduction of PutA(669)-bound FAD was determined by potentiometric titrations, which is 15 mV more negative than the E(m) for PutA-bound FAD. The pH behavior of the E(m) for PutA(669)-bound FAD was measured in the pH range 6.5-9.0 at 25 degrees C and exhibited a 0.03 V/pH unit slope. Analysis of the DNA and membrane-binding properties of PutA(669) shows that it binds specifically to the put intergenic control DNA with a binding affinity similar to that of PutA. In contrast, we did not observe functional association of PutA(669) with membrane vesicles. We conclude that PutA(669) has FAD-binding and DNA-binding properties comparable to those of PutA but lacks a membrane-binding domain necessary for stable association with the membrane.

Bacterial Proteins↗

PubMed1987

Mast cells from the human intestinal lamina propria. Isolation, histochemical subtypes, and functional characterization.

With the use of a collagenase dispersion technique, cells were isolated from the lamina propria of the human small and large intestine. The cell suspensions contained 8% mast cells, which on average contained 1 to 2 pg of histamine/cell. With the use of histochemical procedures based upon fixative sensitivity and dye binding, which identify functionally distinct mast cell subtypes in the rat, dispersed human intestinal mast cells contained approximately equal proportions of two histochemical subtypes analogous to those in the rat. Whether these are functionally distinct as in the rat remains to be determined. The histochemically mixed mast cell populations from the human intestinal mucosa secreted histamine in a dose- and energy-dependent manner in response to anti-IgE and A23187, but not 48/80. Theophylline, doxantrazole, quercetin, and salbutamol all significantly inhibited anti-IgE-induced histamine secretion by human intestinal mast cells, but cromolyn sodium and the experimental antisecretory drugs, nedocromil sodium and FPL 52694, did not inhibit histamine secretion by the mast cell mixture to a statistically significant extent. Cromolyn sodium inhibited histamine secretion by 15 to 30%, and whether this reflected inhibition of one of the two histochemical mast cell subtypes to a greater extent than the other or all the cells to a minimal degree remains to be established. Control investigations of the intestinal cell isolation procedure indicated that these qualities did not reflect effects of the cell dispersal procedure. Further characterization and analysis of intestinal mast cells is essential to determine if functionally distinct mast cell subtypes exist in human tissues.

Antibodies, Anti-Idiotypic↗