Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.11.1.7 (peroxidase)
65,474 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Direct histocytochemical staining methods on undisrupted tissues, stabilized by chemical fixation, potentially offer perhaps the most reliable approach to the study of the enzymes of the cell with relation to its ultrastructure. The atoms which, for the most part, comprise the biomacromolecules and enzymes of cells and tissues contribute little to their inherent electron opacity or ability to scatter electrons differentially. The latter property of a substance is responsible for its observation with the electron microscope. Since the introduction of osmiophilic reagents into cytochemistry (HANKER et al. 1964), the selective deposition of relatively large amounts of polymeric osmium black reaction products at the subcellular sites of insoluble or immobilized enzymes or biomacromolecules has facilitated their demonstration with the light and electron microscopes. Perhaps the most widely employed osmiophilic reagent in histocytochemistry has been DAB which was introduced by GRAHAM and KARNOVSKY (1966a, b). Although it receives its widest use for demonstrating the sites to which the exogenous ultrastructural tracer horseradish peroxidase (HRP) is transported in vertebrate tissues, it is also widely employed for the demonstration of catalase in peroxisomes with the media of FAHIMI (1969) or of NOVIKOFF and GOLDFISCHER (1969), and for the demonstration of cytochrome oxidase with the medium of SELIGMAN et al. (1968a). The importance of this reagent lies in its ability to undergo oxidative polymerization forming an insoluble osmiophilic melanin-like product (HANKER et al. 1972a) which comforms well to ultrastructure, at the sites of enzymic or nonenzyme proteins which catalyze its oxidation. In the past few years, studies in our laboratory have shown that a rational approach to the histocytochemical demonstration of enzymes could be devised. It is based on the selective deposition of transition metal compounds at the sites of enzymes that resemble hemoproteins in their ability to catalyze the oxidative polymerization of DAB. The most useful of these compounds, cupric ferrocyanide (Hatchett's brown) was also introduced into cytochemistry by Karnovsky's laboratory (KARNOVSKY 1964; KARNOVSKY and ROOTS 1974). By the use of natural substrates, when available, or synthetic substrates which liberate or form a reducing agent at the sites of the enzymatic activity, many diverse types of enzymes have been demonstrated by methods depending on this principle known as catalytic osmiophilic polymer generation. DAB has probably been the most useful histocytochemical reagent of the past decade. Yet its borderline carcinogenicity and the frequent interruption of a supply of good quality DAB have encouraged research into a substitute reagent. A new substitute for DAB has resulted from the study of artificial melanins in our laboratory for several years. It consists of a mixture of p-phenylenediamine and pyrocatechol and is much better than DAB for the demonstration of HRP used as a cytochemical tracer...
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PMID:Osmiophilic reagents in electronmicroscopic histocytochemistry. 9 99

Catechol estrogens (CE) are among the major metabolites of estrone (E1) and 17 beta-estradiol (E2). Oxidation of these metabolites to semiquinones and quinones could generate ultimate carcinogenic forms of E1 and E2. The 2,3- and 3,4-quinones of E1 and E2 were synthesized by MnO2 oxidation of the corresponding CE, following the method for synthesizing E1-3,4-quinone [Abul-Hajj (1984) J. Steroid Biochem. 21, 621-622]. Characterization of these compounds was accomplished by UV, nuclear magnetic resonance, and mass spectrometry. The relative stability of these compounds was determined in DMSO/H2O (2:1) at room temperature, and the 3,4-quinones were more stable than the 2,3-quinones. The four quinones directly reacted with calf thymus DNA to form DNA adducts analyzed by the 32P-postlabeling method. The adducts were compared to those formed when the corresponding CE were activated by horseradish peroxidase (HRP) to bind to DNA. The E1- and E2-2,3-quinones formed much higher levels of DNA adducts than the corresponding 3,4-quinones. In addition, many of the adducts (70-90%) formed by the E1- and E2-2,3-quinones appeared to be the same as those formed by activation of 2-OHE1 or 2-OHE2 by HRP to bind to DNA. Little overlap was observed between the adducts formed by E1- and E2-3,4-quinones and HRP-activated 4-OHE1 and 4-OHE2. These results suggest that semiquinones and/or quinones are ultimate reactive intermediates in the peroxidatic activation of catechol estrogens.
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PMID:Synthesis and characterization of estrogen 2,3- and 3,4-quinones. Comparison of DNA adducts formed by the quinones versus horseradish peroxidase-activated catechol estrogens. 133 90

In routine guaiacol assays for thyroid peroxidase and lactoperoxidase employing a newly purchased bottle of guaiacol from Aldrich Chemical Co., we were surprised to find the formation of a blue color instead of the expected amber color classically associated with this assay. This was observed also with horseradish, myelo-, and cytochrome c peroxidase. The blue color (Amax approximately 650 nm) was not formed with guaiacol reagents obtained from two other chemical companies, nor was it seen with a bottle of old Aldrich guaiacol that had been in use in the laboratory for more than 10 years. In the present investigation we provide evidence that formation of the blue color is closely associated with the presence of a low concentration of catechol (approximately 0.5 mol%) in the new Aldrich guaiacol reagent. Catechol itself, even in much higher concentration, is a very weak donor for peroxidase, forming a light pink color. The blue color in Aldrich new guaiacol is not formed to the exclusion of 470-nm-absorbing product(s). Formation of the latter is, however, inhibited, and use of Aldrich new guaiacol for assay leads to low values for peroxidase activity. Other dihydroxyphenols (resorcinol and hydroquinone) do not mimic the action of catechol in formation of the blue color. Resorcinol is a very potent inhibitor of peroxidation of guaiacol. Possible schemes are proposed for formation of the products that may be associated with the amber and blue colors.
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PMID:An unexpected side reaction in the guaiacol assay for peroxidase. 144 72

The coadministration of phenol and hydroquinone has been shown to produce myelotoxicity in mice similar to that observed following benzene exposure. One explanation of this phenomenon may be that phenol enhances the peroxidase-dependent metabolic activation of hydroquinone in the mouse bone marrow. Here we report that radiolabeled [14C]hydroquinone and [14C]phenol bind covalently to tissue macromolecules of blood, bone marrow, liver and kidney, when administered intraperitoneally to the mouse in vivo. Substantially more radiolabeled hydroquinone was covalently bound 18 h after administration as compared with that bound after 4 h. Phenol, when administered together with [14C]hydroquinone, significantly stimulated the covalent binding of [14C]hydroquinone oxidation products to blood (P less than 0.001) and bone marrow (P less than 0.05) macromolecules, but had no significant effect on covalent binding of [14C]hydroquinone oxidation products to liver and kidney macromolecules (P greater than 0.05). Catechol, on the other hand, had no effect on the binding of [14C]hydroquinone oxidation products in either bone marrow, kidney or liver (P greater than 0.05). When hydroquinone was administered together with [14C]phenol, a stimulation of the covalent binding of phenol oxidation products to bone marrow macromolecules also occurred (P less than 0.05). In addition, hydroquinone co-administration increased the covalent binding of [14C]phenol oxidation products in kidney and blood (P less than 0.05), but significantly decreased the covalent binding in liver (P less than 0.05). These results suggest that altered pharmacokinetics play a major role in the hydroquinone-dependent stimulation of covalent binding of [14C]phenol oxidation products to extrahepatic tissue macromolecules in vivo. The mechanism underlying the phenol-induced stimulation of binding of [14C]hydroquinone by phenol in blood and bone marrow remains unclear, but stimulation of peroxidase-mediated hydroquinone metabolism may be responsible. The latter may therefore play an important role in benzene-induced myelotoxicity.
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PMID:Phenol-induced stimulation of hydroquinone bioactivation in mouse bone marrow in vivo: possible implications in benzene myelotoxicity. 234 55

Because benzidine and its derivatives have possible carcinogenic activity, a safe method is needed to demonstrate endogenous peroxidase activity. Colonies derived from mouse bone marrow cells in plasma clot culture were classified as granulocyte (CFU-g) or macrophage (CFU-m) precursors by peroxidase and naphthol AS acetate (NASA) esterase staining, respectively. Endogenous peroxidase activity was measured using benzidine or p-phenylenediazine-pyrocatechol (PPD-PC). The effectiveness of peroxidase staining with both reagents was evaluated under several conditions, and the enzyme property was confirmed by inactivation with a variety of inhibitors. The level of peroxidase activity did not differ significantly between PPD-PC and benzidine. Colony number and number of cultured cells were strongly correlated (P greater than 0.983). We conclude that PPD-PC safely demonstrates peroxidase activity in cultured cells and is as accurate, reliable, and efficient as benzidine.
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PMID:Differential identification of mouse granulocyte (CFU-g) and macrophage (CFU-m) precursors in plasma clots. 240 88

Various chromogen protocols for visualizing peroxidase and alkaline phosphatase activity in immunoenzyme histochemistry were compared with respect to their sensitivity. They were tested on tissue sections of human skeletal muscle and in an antigen spot test using antibodies against slow skeletal muscle myosin. The chromogens included 3-amino-9-ethylcarbazole (AEC), 3, 3'-diaminobenzidine (DAB), p-phenylenediamine-pyrocatechol (PPD-PC) and 4-chloro-1-naphthol (CN) in peroxidase histochemistry, and 5-bromo-4-chloro-3-indolyl phosphate-nitro blue tetrazolium salt (BCIP-NBT), BCIP-tetra nitro blue tetrazolium salt (TNBT) and various combinations of substituted naphthol phosphate-diazonium salt in alkaline phosphatase histochemistry. DAB, CN, and PPD-PC were also employed with imidazole and DAB in addition to Co2+ and Ni2+ ions. The results indicate that DAB-imidazole and DAB-Co2+ and Ni2+ ions are the most sensitive chromogen protocols for visualizing peroxidase activity. Although no large differences were found between the various chromogen protocols for visualizing alkaline phosphatase activity, the protocol BCIP-TNBT is especially recommended. Furthermore, the various chromogen protocols were evaluated as to stability of chromogen solutions and final precipitates, background staining, localization properties, and enhancement of enzyme activity.
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PMID:Sensitivity of various visualization methods for peroxidase and alkaline phosphatase activity in immunoenzyme histochemistry. 241 13

Experiments were performed in order to investigate, at the light- and electron-microscopic levels, the transneuronal transport of wheatgerm agglutinin conjugated with horseradish peroxidase (WGA:HRP) in the somatosensory system of rats. In five adult albino rats, various amounts of WGA:HRP at different concentrations were bilaterally injected in the dorsal column nuclei (DCN). In one additional animal, WGA:HRP was injected in one side, whereas free HRP was injected in the contralateral DCN. In another five rats, WGA:HRP was injected in the first somatosensory cortex (SI). The postinjection survival time of the animals ranged from 30 to 48 hr. The histochemical visualization of the enzyme was performed using either paraphenylenediamine-pyrocatechol (PPD-PC) or tetraethylbenzidine (TMB) as chromogens on adjacent horizontal serial sections. All the reacted samples were studied at the light-microscopic level, and sections from four animals were processed for the ultrastructural investigation. After DCN injections, a massive anterograde labeling was always observed in nucleus ventralis posterolateralis (VPL) of the thalamus, where also labeled neurons and glial cells were detected at both the light- and the electron-microscopic levels. Labeled neurons and terminals in the lateral border of nucleus reticularis (Re) of the thalamus were also observed after either DCN or SI injection of WGA:HRP. Our results show that WGA:HRP is effectively transported not only anterogradely and retrogradely through the somatosensory system of the rat, but also transneuronally. The transneuronal transfer of the tracer seems to be mainly related to the postlabeling survival time of the animal, and it does not occur when free HRP is injected. In controlled experimental conditions, WGA:HRP can therefore be useful for tracing secondary projections. Moreover, in the present report, the existence of a mediolateral arrangement of the projections of the somatosensory-related area of Re toward VPL is directly demonstrated. As for the histochemical procedure employed, differences in sensitivity are shown between PPD-PC and TMB, although the same general pattern of labeling is present with both chromogens.
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PMID:Transneuronal transport of wheatgerm agglutinin conjugated with horseradish peroxidase in the somatosensory system of the rat: a light- and electron-microscopic study. 242 66

An electron-microscopic investigation of the synaptic organization of the rat's ventroposterolateral nucleus (VPL) and of a reticular thalamic nucleus (RTN) area related to somatosensory thalamic nucleus was performed. In a group of 11 rats, wheatgerm agglutinin conjugated to horseradish peroxidase (WGA:HRP) was injected either in the first somatosensory area of cortex (SI) or in the dorsal column nuclei (DCN). The retrogradely and/or anterogradely transported enzyme was visualized using paraphenylenediamine-pyrocatechol (PPD-PC) as substrate. In a second series of six experiments, an immunocytochemical procedure using a specific anti-gamma-aminobutyric acid (anti-GABA) was employed. Postembedding localization of GABA was performed for ultrastructural observation by means of the colloidal gold immunostaining procedure. Thin sections of recognized VPL and RTN areas from WGA:HRP-injected animals were further processed for immunocytochemistry in order to localize simultaneously, at the electron-microscopic level, the transported enzyme and GABA. The results obtained with this procedure demonstrated that HRP-labeled terminals from DCN contacted the soma and proximal dendrites of VPL neurons, while the terminals labeled after SI cortical injections were predominantly localized to the distal portion of the dendrites. The same cortical injection also determined the presence of labeled synaptic boutons contacting the soma, and both proximal and distal dendrites of RTN neurons. GABA-immunolabeled terminals were observed in VPL in a number larger than those observed with other methods, since not only typical F terminals were labeled but also terminals containing round and/or pleomorphic vesicles. GABA-ergic terminals contacted the soma and the proximal and distal dendrites of VPL neurons, while in RTN cells they made synaptic contact mainly with the soma and proximal dendrites. In the double-labeling experiments, terminals containing both HRP and specific immunogold GABA staining were never observed. The present data provide a direct demonstration of the presence of a strong inhibitory input from RTN upon VPL neurons and of the existence of autoinhibition within RTN neurons.
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PMID:The intrinsic organization of the ventroposterolateral nucleus and related reticular thalamic nucleus of the rat: a double-labeling ultrastructural investigation with gamma-aminobutyric acid immunogold staining and lectin-conjugated horseradish peroxidase. 328 95

Insulin-like immunoreactivity was localized in tissue sections and cell cultures of mouse seminal vesicle using the indirect technique of immunocytochemistry. Seminal vesicles were cut into fragments, fixed in 2.5% glutaraldehyde, embedded in epoxy resin, sectioned at 1 micron, and transferred to glass slides. Epithelial cell cultures of seminal vesicle were grown on coverslips in Dulbecco's Minimal Essential Medium for 4-6 days and fixed in 2.5% glutaraldehyde. Sections (etched with sodium ethanolate) or coverslips were incubated in guinea pig antiporcine insulin antiserum, in antiserum immunoabsorbed with porcine insulin, or in normal guinea pig serum. For indirect immunocytochemistry, incubation with primary antiserum was followed by treatment with rabbit anti-guinea pig immunoglobulin (Ig) G conjugated to peroxidase, or with protein A and then rabbit peroxidase anti-peroxidase (PAP). Finally, treated samples were incubated in phenylenediamine-pyrocatechol-H2O2 substrate mixture for 6-8 min at room temperature. Specific immunoreactivity to insulin antisera was confined to the epithelium of the seminal vesicle in tissue sections. No staining occurred in subepithelial connective tissue. Specific immunoreactivity was also observed in the cytoplasm of cultured seminal vesicle epithelial cells.
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PMID:Immunocytochemical localization of insulin-like immunoreactivity in mouse seminal vesicle. 354 7

p-Phenylenediamine/pyrocatechol mixture (PPD-PC) was evaluated as a reagent for the ultracytochemical demonstration of retrograde axonal transport of horseradish peroxidase (HRP). HRP crystals were applied to the proximal stumps of the severed infraorbital nerves in rats. After 48 h the rats were sacrificed by perfusion, and the trigeminal ganglia ipsilateral to the severed nerves were processed for HRP cytochemistry and then prepared for electron microscopy. PPD-PC was rapidly oxidized in HRP-labeled neurons to form a dark brown-black osmiophilic reaction product which was more readily visible than the DAB product in the sections. This facilitated selection by light microscopy of areas in the epoxy wafers for ultrathin sectioning. In thin sections viewed under the electron microscope, the osmicated electron opaque PPD-PC reaction product was present in membrane-bound structures including smooth endoplasmic reticulum and granules of various sizes. The PPD-PC reaction product formed after 10-min incubation appeared to be more electron opaque than the DAB reaction product formed after 20 min. PPD-PC was found to be much less readily oxidized than DAB by endogenous hemoproteins. This methodology facilitated the ultracytochemical localization of HRP in neurons following retrograde axonal transport.
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PMID:Facilitated ultracytochemical demonstration of retrograde axonal transport of horseradish peroxidase in peripheral nerve. 615 30


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