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:4.2.2.7 (heparinase)
1,270 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A protein kinase capable of phosphorylating basic fibroblast growth factor (FGF) can be localized on the outer cell surface of human hepatoma cells (SK-Hep cells). The addition of [gamma-32P]ATP, but not H3(32)PO4, results in a rapid (less than 10 min) incorporation of 32P into exogenously added basic FGF. The reaction is time and concentration dependent (apparent Km, 170 nM) and is stimulated by the addition of cAMP (EC50, 0.5 microM), but not the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate. There is also no tyrosine protein kinase detected on the cell surface. The inhibition of basic FGF binding to its low and/or high affinity sites decreases the phosphorylation of basic FGF by the ecto-protein kinase. Accordingly, pretreatment of cells with heparinase for 30 min or coincubation with heparin (0.1-10 micrograms/ml) decreases phosphorylation in a dose-dependent manner. Furthermore, the addition of a nonphosphorylatable peptide analog of basic FGF ([Val112] basic FGF-(106-146)NH2) that can compete with basic FGF binding to cells prevents the phosphorylation of basic FGF. Together, these observations suggest that 1) exogenous basic FGF must associate with its low and/or high affinity binding sites to be phosphorylated, and 2) the kinase is cAMP dependent and associated with the outer cell surface, and support the hypothesis that phosphorylation may regulate the activity and/or bioavailability of the growth factor.
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PMID:Phosphorylation of basic fibroblast growth factor by a protein kinase associated with the outer surface of a target cell. 165 31

Acidic oligosaccharides derived from glycosaminoglycan heparin were separated by polyacrylamide gradient gel electrophoresis (PAGE). The gel could be visualized using Alcian Blue dye to give a pattern of highly resolved, well defined bands. The particular banding pattern obtained was the result of a heparinase catalyzed depolymerization which afforded oligosaccharide products that differed in size by one disaccharide unit. The separated oligosaccharides could be recovered prior to staining by electroelution onto a positively charged nylon membrane by a semi-dry transfer procedure. Subsequent elution and quantitative recovery of individual oligosaccharides from the membrane was achieved. By using multiple membrane layers a second separation dimension was obtained, resulting in increased oligosaccharide purity proportional to transfer depth. Preparative gradient polyacrylamide gel electrophoresis followed by semi-dry electro-transfer and recovery represents a novel method for the preparation of homogeneous acidic oligosaccharides.
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PMID:Isolation and recovery of acidic oligosaccharides from polyacrylamide gels by semi-dry electrotransfer. 169 Jun 41

Basic fibroblast growth factor (bFGF) was radiolabeled and used in axonal transport studies to determine whether certain neuronal populations express functional receptors for bFGF. Unlike 125I-NGF, 125I-bFGF was not retrogradely transported in the adult rat sciatic nerve or from iris to trigeminal ganglion or superior cervical ganglion. However, after intraocular injection of 125I-bFGF into the posterior chamber of the eye of adult rats, radioactivity was detected within the retinal ganglion cell projections. This radioactivity was localized to the ipsilateral optic nerve and in the contralateral lateral geniculate body and the contralateral superior colliculus by using autoradiographic techniques. Direct measurement of the radioactivity in dissected brain regions was used to study the process of 125I-bFGF uptake and transport by retinal ganglion cells. The uptake and transport were specific for biologically active bFGF since neither denatured, biologically inactive 125I-bFGF nor 125I-NGF was taken up and transported. The uptake and transport of 125I-bFGF were saturable phenomena since they were blocked in the presence of excess, unlabeled bFGF. Wheat germ agglutinin, but not heparinase, blocked uptake and transport of 125I-bFGF, a finding that is consistent with the uptake being mediated by high-affinity bFGF receptors. Radioactivity from 125I-bFGF was transported in retinal ganglion cell axons in an anterograde direction at a maximum rate in excess of 1.7 mm/hr. No specific retrograde transport of bFGF to the retina was detected after 125I-bFGF was injected into the superior colliculus. The radioactivity from 125I-bFGF that accumulated in the superior colliculus was lost from this tissue with a half-life of about 22 hr. Autoradiography of proteins separated by SDS-PAGE demonstrated that 125I-bFGF was not substantially degraded in the retina after internalization within retinal ganglion cells. During anterograde transport, however, 125I-bFGF underwent limited proteolytic cleavage resulting in 3 prominent 125I-bFGF derivatives of molecular weights greater than 7000 Da. Although these were the major radioactive species recovered from the superior colliculus after intraocular injection, some intact 125I-bFGF was also detected within the innervated target. These results indicate that retinal ganglion cells express high-affinity receptors for bFGF, that these receptors mediate the internalization of bFGF, that internalized bFGF undergoes limited proteolytic cleavage, and that bFGF and its derivatives are anterogradely transported to the lateral geniculate body and the superior colliculus. These data raise the possibility that bFGF or its derivatives may act as an anterograde trophic factor in the visual system, a system that is known to undergo anterograde transneuronal cell death.
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PMID:Basic fibroblast growth factor: receptor-mediated internalization, metabolism, and anterograde axonal transport in retinal ganglion cells. 169 44

Org 10172 provided adequate anticoagulation for this patient. An excellent correlation between anti-factor Xa activity and ACT was observed at the doses used for CPB. If high-dose Org 10172 is used, these data suggest that it may be possible to circumvent the measurement of anti-factor Xa activity by using the ACT as an index of this heparinoid's anticoagulant effect. Because postoperative bleeding may be excessive, however, development of a method of reversal of Org 10172 is desirable. Although the optimal ACT, dose, plasma concentration, and means of reversal (e.g., protamine vs. heparinase) remains to be determined, heparinoids provide an alternate means of anticoagulation for CPB in patients unable to receive standard heparin.
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PMID:"Heparin-free" cardiopulmonary bypass: first reported use of heparinoid (Org 10172) to provide anticoagulation for cardiopulmonary bypass. 169 48

The effect of specific glycosaminoglycan-hydrolyzing enzymes on the ruthenium red staining of pig spermatozoa was studied. Washed spermatozoa were incubated at 35 degrees C in buffer or with neuraminidase 0.5 units/ml, heparinase 0.2 mg/ml, or chondroitinase ABC 2.0 units/ml. After incubation sperm cells were washed, stained with ruthenium red and studied under the electron microscope. Anionic sites in the surface of untreated spermatozoa follow regularly the plasma membrane, but present are numerous processes constituting what has been defined as the glycocalyx. Neuraminidase did not affect the distribution of ruthenium red on the surface of the spermatozoa, but eliminated almost completely the processes of the glycocalyx. Heparinase caused loss of the ruthenium red-stained sites on the membrane surface of pig spermatozoa with less influence on the dense processes of the glycocalyx. A similar loss of ruthenium red-stained sites was observed with nitrous acid treatment. A striking effect of treatment with chondroitinase ABC was the production of a typical acrosome reaction.
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PMID:Glycosaminoglycan-sulfate as plasma membrane component of pig spermatozoa. 169 1

It has been reported that small cytoplasmic RNA (scRNA) from human placenta inhibits translation of most mRNAs in both wheat germ extracts and reticulocyte lysates (Lorberboum, H., Digweed, M., Erdmann, V. A., Servadio, Y., Weinstein, D., De Groot, N., and Hochberg, A. A. (1986) Eur. J. Biochem. 155, 279-287). We have investigated the mechanism by which scRNA preparations inhibit mRNA translation in vitro. We demonstrate that the inhibitory agent(s) is not sensitive to treatment with various ribonucleases but that translational inhibition is sensitive to incubation with 1 N NaOH at 95 degrees C or to treatment with heparinase. Based on these findings and on the ability of heparin to inhibit cell-free translation by itself, we conclude that the presence of heparin in preparations of scRNA from human placenta is responsible for effects which have previously been attributed to inhibitory RNA molecules. Since heparin is also frequently used in the isolation of translationally inhibitory scRNAs from other sources, we suggest that the sensitivity of these preparations to ribonucleases and heparinase should be examined.
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PMID:Effect of heparin contained in preparations of small cytoplasmic RNAs on cell-free translation. 170 59

Pentosanpolysulfate (PPS) represents the product obtained after sulfation of xylan and is composed of beta 1----4-D-xylopyranose residues sulfated at C2 and C3. Studies have shown that this compound can often be effective in relieving the symptoms of interstitial cystitis (IC). In order to elucidate the mode of action of PPS in IC, a sensitive and reliable assay was needed. To this end we prepared an immunogenic form of PPS by coupling it to methylated bovine serum albumin (MBSA). This complex was used to immunize NZW rabbits (1 mg, IM). Four of five animals responded with anti-PPS antibodies, three of which had high titer (greater than 1/2000) as measured by an enzyme-linked immunosorbent assay (ELISA). All sera were routinely absorbed with an MBSA-Sepharose immunoadsorbent to remove anti-MBSA antibodies. ELISA inhibition tests were used to determine the sensitivity and specificity of the sera. At least 50 ng/ml of PPS could be routinely detected by this assay. A number of naturally occurring proteoglycans, polysaccharides, monosaccharides and disaccharides were examined for reactivity with the antibodies but only heparin was an effective inhibitor. Absorption with heparin immunoadsorbents reduced, but did not eliminate, the ability of heparin to inhibit anti-PPS binding. This activity could be destroyed by treatment with heparinase without affecting PPS inhibition. Normal urine did not affect the ELISA or ELISA inhibition tests and thus allowed the determination of PPS levels in IC patient urines. Initial analysis of seven IC patients receiving oral PPS revealed urine concentration of 0.8-16.0 micrograms/ml. No inhibition could be detected in pre-treatment urine samples.
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PMID:The production of antibodies to pentosanpolysulfate (ELMIRON, SP-54). 170 2

The effect of nerve growth factor (NGF) on proliferation/differentiation of mast cells was investigated in vitro. Although NGF alone neither supported colony formation of bone marrow-derived cultured mast cells (BMCMC) nor induced development of mast cell colonies from nonadherent bone marrow cells (NBMC), addition of NGF to the suboptimal dose of interleukin 3 (IL-3) significantly increased the numbers of mast cell colonies produced by BMCMC or NBMC in methylcellulose. When stimulated by IL-3 alone, cells in mast cell colonies were not stained by berberine sulfate, a fluorescent dye. In contrast, mast cells developing in methylcellulose cultures obtaining both IL-3 and NGF were stained by berberine sulfate. The fluorescence was abolished by the treatment of heparinase but not of chondroitinase ABC, suggesting that mast cells stimulated by IL-3 and NGF produced and stored heparin proteoglycan. The histamine content of BMCMC maintained by IL-3 was also increased by addition of NGF. Since BMCMC showed mucosal mast cell-like phenotype, NGF appeared to induce the phenotypic change to connective tissue-type mast cells (CTMC). In the culture containing BMCMC, 3T3 fibroblasts, and IL-3, the phenotypic change of BMCMC to CTMC was observed as well. Since NGF was detected in this coculture and since addition of anti-NGF monoclonal antibody suppressed the phenotypic change, NGF produced by fibroblasts appeared to induce the phenotypic change. Neither BMCMC alone nor IL-3 alone increased the concentration of NGF. Therefore, there is a possibility that BMCMC stimulated by IL-3 may induce the production and/or release of NGF by fibroblasts.
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PMID:Nerve growth factor induces development of connective tissue-type mast cells in vitro from murine bone marrow cells. 171 69

Conditioned medium from Sertoli cells, prepared from testes of 20-day-old rats, contains component(s) that inhibit the incorporation of [3H]-thymidine into DNA of peritubular myoid cells (PMC) and inhibit the proliferation of PMC. These components are trypsin-resistant, heat-stable compounds having a molecular weight less than 30,000. The active inhibitory components in Sertoli cell conditioned medium are inactivated by treatment with heparinase, but not by treatment with hyaluronidase or chondroitin sulfate lyases. Addition of heparin or heparan sulfate results in inhibition of DNA synthesis by PMC in a dose-dependent manner, whereas other glycosaminoglycans (GAGs) examined (hyaluronic acid, keratan sulfate, and chondroitin sulfate) have no detectable effects. Heparin and heparan sulfate are unique among GAGs tested in inhibiting the characteristic multilayer growth pattern of PMC following the attainment of confluence in serum-rich medium. On the basis of these and other data presented, it is concluded that heparin and other heparin-like GAGs synthesized by Sertoli cells are implicated in the modulation of growth of PMC in vitro during co-culture. It is postulated that heparin may play a similar role in maintaining the quiescent peritubular myoid cell phenotype in vivo.
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PMID:Sertoli cells in culture secrete paracrine factor(s) that inhibit peritubular myoid cell proliferation: identification of heparinoids as likely candidates. 171 60

Glycosaminoglycans have been demonstrated throughout the cutaneous BMZ at the ultrastructural level. Colloidal iron and cationised ferritin proved of limited value, whilst staining with Alcian blue and application of the critical electrolyte concentration principle has provided evidence for the presence of sulphated GAGs at the lamina lucida and lamina reticularis. Digestions with chondroitin ABC lyase and heparin lyase have confirmed the existence of chondroitin and/or dermatan sulphates and heparan sulphates, although the results obtained with hyaluronate lyase have indicated that hyaluronates are also present.
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PMID:Ultrastructural localisation of anionic sites at the dermo-epidermal junction in normal human skin. 171 20


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