Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:4.2.2.7 (heparinase)
 1,270 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have previously reported that exposing cultured Madin Darby canine kidney (MDCK) cells to the polycation protamine (PRO) results in increased short-circuit current and decreased barrier integrity as measured by mannitol permeability and transepithelial electrical resistance. To further investigate the interaction of PRO with the surface of epithelial cells, we labeled PRO with [14C] with use of reductive alkylation. [14C]PRO bound to the cells in a biphasic pattern. Approximately 10% of the [14C]PRO was bound to the cells in the first 5 min, followed by an additional 10% that was bound over the next 25 min. No additional [14C]PRO bound to the cells after the initial 30 min. Binding of [14C]PRO was inhibited by "cold" PRO, which suggested specificity. Binding was also inhibited by polyanions, serum, and albumin, agents previously found to protect MDCK cells from PRO-induced injury. The binding of PRO to MDCK cells was not inhibited by incubation of the MDCK cells with neuraminidase, to remove surface sialic acid residues, or with heparinase, to remove surface heparan sulfate, even though metabolic labeling experiments demonstrated that neuraminidase decreased cell sialic acid and heparinase decreased cell heparan sulfate. Neuraminidase and heparinase offered no protection from PRO injury and had no effect themselves on mannitol permeability. Incubation of the cells with trypsin, however, blunted both the binding of PRO to the cells and the increase in mannitol permeability after exposure of the cells to PRO.(ABSTRACT TRUNCATED AT 250 WORDS)
 ...
PMID:Protamine interaction with the epithelial cell surface. 153 21

To evaluate the effect of biochemical modifications not possible in vivo, filters of dog glomerular basement membrane (GBM) were constructed in ultrafiltration cells in vitro. The sieving coefficients (SCs) of three protein markers of differing size and charge (native, anionic bovine albumin-BSA; cationized BSA-cBSA; and immunoglobulin G-IgG) were determined using filters of differing amounts of control GBM, and under varying transmembrane pressures (delta P). Flow rates did not increase proportionately with increasing delta P, indicating filter compressibility. Protein SCs did not change with changing delta P, but did decrease with increasing filter thickness. Control filters showed a small but definite charge selectivity (SCcBSA++ - SCBSA greater than 0); a much greater degree of size selectivity (SCcBSA - SCIgG) was observed. Hexadimethrine (HDM), a polycation which causes proteinuria in vivo, led to marked increases in protein SCs. In contrast, removal of the major population of intrinsic GBM negative charges by carboxyl group methylation only produced a small increase in the filtration of BSA, with no change in filtration of cBSA or IgG. Other biochemical modifications (heparinase or neuraminidase treatment) had no effect on filter permselectivity. Carboxyl group methylation essentially abolished filter binding of cationized ferritin, which showed substantial binding to control filters. These in vitro studies provide confirmatory evidence for a direct effect of HDM on the permselective properties of GBM. In addition, biochemical modification studies suggest a fundamental difference between the binding of an exogenous polycation to GBM anionic sites and the removal of intrinsic charges.
 ...
PMID:Macromolecular sieving by glomerular basement membrane in vitro: effect of polycation or biochemical modifications. 207 68

Physiological stimuli induce rapid and unexplained increases in the number of red blood cells within capillaries of skeletal muscle. We hypothesized that such alterations in intracapillary red cell numbers might be due to an undefined interaction between one or more components of blood and the luminal surface of the capillary. This proposition was tested by in situ microperfusion of capillaries with enzymes directed against macromolecules likely to be expressed on the surface of endothelial cells. The instantaneous fractional volume of red blood cells within a capillary (tube hematocrit) was used as an index of a capillary's response to enzyme microperfusion. Five to 8 min of perfusion with enzyme vehicle (0.25% albumin-Ringer solution) produced no significant alteration in capillary tube hematocrit. Perfusion with solutions containing heparinase raised the tube hematocrit at least twofold (P less than 0.05) without a significant change in red cell velocity. Heat-denatured heparinase and other enzymes such as neuraminidase, hyaluronidase, papain, pronase E, and clostripain had no detectable effect on the tube hematocrit (P greater than 0.05). After enzyme treatment, application of adenosine (10(-4) M) or oxygen caused brisk vasomotor responses in arterioles feeding perfused capillary units, but the usual changes in the tube hematocrit were not observed. Thus heparinase treatment results in a sustained elevation in the capillary tube hematocrit and a dissociation of the typical relationship between vasomotor changes and red cell distribution in capillaries. These findings suggest that physiological stimuli which alter the number of red blood cells within capillaries may operate by modifying interactions between plasma and one or more components on the luminal surface of capillaries.
 ...
PMID:Heparinase treatment suggests a role for the endothelial cell glycocalyx in regulation of capillary hematocrit. 231 79

We undertook studies in the isolated perfused rat lung to determine 1) the effects of endothelial charge neutralization with the polycation protamine sulfate on microvascular permeability, lung water, and anionic ferritin binding to the endothelium and 2) the role of heparan sulfate and hyaluronate, negatively charged cell surface glycosaminoglycans, on permeability. Capillary permeability was determined by tissue 125I-albumin accumulation in isolated perfused rat lungs. In control lungs the 5-min albumin uptake was 0.50 +/- 0.05 cm3.s-1.g dry tissue-1 X 10(-3). It was increased by 132 +/- 7.8% (P less than 0.001) by protamine (0.08 mg/ml) and 65 +/- 12% (P less than 0.01) by heparinase (5 U/ml), whereas hyaluronidase (25 NFU/ml) was without effect. In control lungs total water was 4.83 +/- 0.15 ml g/dry tissue. Protamine increased lung water 12 +/- 2% (P less than 0.05). Heparinase caused a 9 +/- 3% increase (P less than 0.05), and hyaluronidase had no effect. Electron microscopy demonstrated that protamine increased anionic ferritin binding to the surface of endothelial cells. We conclude that protamine sulfate neutralization of negative charge in the pulmonary microcirculation leads to increased microvascular permeability. Heparin sulfate may be responsible for this charge effect.
 ...
PMID:Effects of protamine, heparinase, and hyaluronidase on endothelial permeability and surface charge. 369 32

Alterations in the permeability of the glomerular basement membrane (GBM) towards native ferritin (NF) and iodinated albumin (125I-BSA) following removal of the major glycosaminoglycans (GAGs) of the GBM, heparan sulfate (HS) and hyaluronic acid (HA), were assessed utilizing the techniques of routine electron microscopy and autoradiography, respectively. Kidneys were incubated with heparinase (to degrade the GAGs of the GBM) and subsequently perfused with either NF or 125I-BSA. Control kidneys, which were not treated with heparinase, showed a low permeability to both tracers, with NF being confined to the lamina rara interna and 125I-BSA exhibiting a low level of passage into the urinary spaces (as indicated by a low density of autoradiographic grains over the urinary spaces). After heparinase treatment there was an increase in the permeability of the GBM such that both NF and 125I-BSA passed through the GBM in larger quantities and entered the urinary spaces. Perfusion of cationized ferritin (CF) into control kidneys revealed this probe to bind to the HS-rich anionic sites present within the GBM. Treatment with heparinase resulted in an abolition of the CF binding thereby indicating that the sites are composed mainly of HS and that HS plays a key role in establishing the permeability properties of the GBM. The changes in the pattern of distribution and density of the anionic sites of the GBM following induction of nephrosis was also studied. Animals were rendered nephrotic by subcutaneous injections of an aminonucleoside of puromycin and their kidneys subsequently perfused with either CF or cationized cytochrome c. No difference in either the pattern of distribution on density of the anionic sites in the GBM of nephrotic kidneys was observed when compared to nonnephrotic controls; thus indicating that the proteinuria associated with aminonucleoside nephrosis might be due to changes in components of the glomerular capillary wall other than the anionic sites.
 ...
PMID:Glycosaminoglycans of the glomerular basement membrane in normal and nephrotic states. 730 62

The binding of albumin to the glomerular capillary wall was studied using albumin-gold in perfused kidneys, the interaction of [3H]albumin with isolated glomeruli at 37 degrees C and 4 degrees C and the interaction at [3H]albumin with purified basement membrane. The albumin-gold was found to bind predominantly to the basement membrane and this interaction could be dissociated with high concentrations of albumin. There was binding of albumin to isolated rat glomeruli which exhibited temperature dependence. Glomeruli exhibited a binding site at both 37 degrees C and 4 degrees C with an association constant in the range of 1 to 3 x 10(4) M-1 that bound 7 x 10(13) molecules/glomerulus. At 37 degrees C, however, there was anomalous Scatchard binding behaviour at relatively higher concentrations of albumin (30 to 50 mg/ml) which could be due to either glomerular cell uptake or the appearance of multiple binding sites or both. The binding of albumin to isolated glomeruli and the glomerular albumin levels in isolated kidney perfusion could largely be accounted for by the binding of albumin to the glomerular basement membrane. The albumin binding to glomeruli at 37 degrees C was enhanced by Pronase digestion and heparinase digestion, but remained unchanged following trypsin treatment or neuraminidase treatment. Similarly, albumin was shown to bind to purified basement membrane preparations. This binding was also enhanced (approximately 80 times) by heparinase digestion but remained unchanged after digestion with chondroitinase ABC or hyaluronidase.(ABSTRACT TRUNCATED AT 250 WORDS)
 ...
PMID:Albumin interaction with the glomerular capillary wall in vitro. 778

A method to provide near-constant sustained release of high molecular weight, water-soluble proteins from polyanhydride microspheres is described. The polyanhydrides used were poly(fatty acid dimer) (PFAD), poly(sebacic acid) (PSA), and their copolymers [P(FAD-SA)]. P(FAD-SA) microspheres containing proteins of different molecular sizes--lysozyme, trypsin, heparinase, ovalbumin, albumin, and immunoglobulin--were prepared by a solvent evaporation method using a double emulsion. The microspheres containing proteins were spherical, with diameters of 50-125 microns, and encapsulated more than 80% of the protein, irrespective of the protein used. Enzymatic activity studies showed that encapsulation of enzymes inside polyanhydride microspheres can protect them from activity loss. When not placed inside polyanhydride microspheres, trypsin lost 80% of its activity in solution at 37 degrees C at pH 7.4 in 12 hr, whereas inside the polyanhydride microspheres the activity loss was less than 10% under these conditions. About 47% of the enzymatic activity of heparinase encapsulated in the microspheres was lost at 37 degrees C in 24 hr, while in solution it lost over 90% of its activity. The protein-loaded microspheres displayed near-zero-order erosion kinetics over 5 days as judged by the release of sebacic acid (SA) from the microspheres. The microspheres degraded to form SA and FAD monomers. All proteins were released at a near-constant rate without any large initial burst, irrespective of polymer molecular weight and protein loading. The period of protein release was longer than that of SA and continued protein release was observed even after the microsphere matrix had completely degraded. Differential scanning calorimetric studies demonstrated an interaction between protein and the FAD monomers produced with microsphere degradation. It is likely that the protein interaction with FAD monomers permits formation of water-insoluble protein aggregates which slowly dissolve and diffuse out of the matrix, leading to delayed protein release. For trypsin-loaded microspheres, trypsin lost 40% of its activity during microsphere preparation. Activity studies demonstrated that the sonication process was primarily responsible for activity loss. A reduction in the period of ultrasound exposure decreased the loss of protein activity to around 20%.
 ...
PMID:Controlled delivery systems for proteins using polyanhydride microspheres. 848 30

Concentrations of up to 1.5 milliunits/ml xanthine oxidase (XO) (1.1 micrograms/ml) are found circulating in plasma during diverse inflammatory events. The saturable, high affinity binding of extracellular XO to vascular endothelium and the effects of cell binding on both XO catalytic activity and differentiated vascular cell function are reported herein. Xanthine oxidase purified from bovine cream bound specifically and with high affinity (Kd = 6 nM) at 4 degreesC to bovine aortic endothelial cells, increasing cell XO specific activity up to 10-fold. Xanthine oxidase-cell binding was not inhibited by serum or albumin and was partially inhibited by the addition of heparin. Pretreatment of endothelial cells with chondroitinase, but not heparinase or heparitinase, diminished endothelial binding by approximately 50%, suggesting association with chondroitin sulfate proteoglycans. Analysis of rates of superoxide production by soluble and cell-bound XO revealed that endothelial binding did not alter the percentage of univalent reduction of oxygen to superoxide. Comparison of the extent of CuZn-SOD inhibition of native and succinoylated cytochrome c reduction by cell-bound XO indicated that XO-dependent superoxide production was occurring in a cell compartment inaccessible to CuZn-SOD. This was further supported by the observation of a shift of exogenously added XO from extracellular binding sites to intracellular compartments, as indicated by both protease-reversible cell binding and immunocytochemical localization studies. Endothelium-bound XO also inhibited nitric oxide-dependent cGMP production by smooth muscle cell co-cultures in an SOD-resistant manner. This data supports the concept that circulating XO can bind to vascular cells, impairing cell function via oxidative mechanisms, and explains how vascular XO activity diminishes vasodilatory responses to acetylcholine in hypercholesterolemic rabbits and atherosclerotic humans. The ubiquity of cell-XO binding and endocytosis as a fundamental mechanism of oxidative tissue injury is also affirmed by the significant extent of XO binding to human vascular endothelial cells, rat lung type 2 alveolar epthelial cells, and fibroblasts.
 ...
PMID:Binding of xanthine oxidase to vascular endothelium. Kinetic characterization and oxidative impairment of nitric oxide-dependent signaling. 998 43

We have investigated the effect of soluble or extracellular-matrix (ECM) -bound heparin in conjunction with various second messenger pathways on cell proliferation and tissue-specific gene expression in primary cultures of hepatocytes. None of the combinations of heparin and second messenger stimulators or inhibitors had an effect on hepatocyte proliferation. Soluble heparin enhanced albumin expression in hepatocytes. Activation of protein kinase C, as well as an increase in intracellular cAMP, abolished this increase in albumin expression in the presence of heparin. When hepatocytes were plated on hepatocyte-derived ECM, containing highly sulfated heparan sulfate chains, activation of protein kinase C and an increase in intracellular cAMP strongly reduced albumin expression in hepatocytes. When heparan sulfate chains were removed from the ECM by heparinase treatment, activation of protein kinase C and increased cAMP were less inhibitory for albumin expression in hepatocytes. Inhibition of tyrosine kinases did not affect the induction of albumin mRNA by heparin. We conclude that heparin induces albumin expression in hepatocytes and activation of protein kinase C or increased intracellular cAMP antagonize this effect. ECM-bound heparan sulfates do not act in the same manner as soluble heparin.
 ...
PMID:Synergies of heparin and second messengers pathways involved in tissue-specific gene expression in hepatocytes. 1134 47

We explored the effect of extracellular matrix (ECM) produced by fetal and adult hepatocytes on tissue-specific gene expression and proliferation of fetal and adult hepatocytes. Adult hepatocytes ECM strongly induced expression of both albumin and HNF-4 in adult hepatocytes. In contrast, fibroblast ECM reduced the expression of mRNAs for albumin and alpha-fetoprotein in fetal hepatocytes. Adult hepatocytes ECM also increased the activity of liver-specific enzymes of adult hepatocytes (DPP IV and glucose-6-phosphatase) in both fetal and adult hepatocytes, while fetal hepatocyte-derived ECM increased activity of the fetal hepatocyte enzyme GGT in fetal hepatocytes. Fibroblast ECM was inhibitory for the activity of all enzymes assayed. Removal of heparin chains from the various matrices by pretreatment of the ECM with heparinase resulted in reduction of glucose-6-phosphatase and DPP IV in adult hepatocytes. Removal of chondroitin sulfate chains from fetal hepatocyte-derived ECM resulted in loss of induction of GGT in the fetal cells. Fetal hepatocytes proliferated best on adult hepatocyte-derived ECM. Adult hepatocytes showed only modest proliferation on both fetal and adult hepatocytes ECM and their growth was inhibited by fibroblast ECM. In conclusion, adult hepatocyte ECM better supports the expression of adult genes, whereas fetal hepatocyte ECM induced expression of fetal genes. Fibroblast derived-ECM was inhibitory for both proliferation and tissue-specific gene expression in fetal and adult hepatocytes. The data support a role for heparan sulfate being the active element in adult ECM, and chondroitin sulfate being the active element in fetal ECM.
 ...
PMID:The role of fetal and adult hepatocyte extracellular matrix in the regulation of tissue-specific gene expression in fetal and adult hepatocytes. 1189 78


1 2 Next >>