Biomedical subjects

V L Murty

Publications and source records attributed to V L Murty.

At least 37 records · Page 2Linked to original sources

PubMed1993

Characterization of gastric mucosal mucin receptor.

Mucin receptor was isolated from gastric epithelial cell membrane by a procedure involving membrane solubilization with octylglucoside followed by affinity chromatography on Sepharose-bound wheat germ agglutinin. The receptor protein yielded on SDS-PAGE a single 97kDa band and displayed specific affinity, in a concentration-dependent manner, towards the mucin-coated surface. The receptor showed requirement for carbohydrate chains in mucin for binding, as their removal caused a marked (87%) reduction in binding capacity. Scatchard analysis revealed a linear plot with a single class of high affinity binding (Kd = 43.8 nM; Bmax = 140 pmol/mg protein) sites. The results demonstrate for the first time the presence in gastric mucosa of a specific receptor for mucin.

Animals↗

PubMed1993

Salivary mucin modulation of buccal mucosal calcium channel activity.

The effect of the low and high molecular weight salivary mucins on the activity of calcium channel isolated from buccal epithelial cell membranes was investigated. The 45Ca2+ uptake into the vesicle-reconstituted channels, while only moderately (15%) affected by the intact mucin forms, was significantly inhibited (60%) by the acidic mucin fractions. This effect was associated with the sialic acid and sulfate ester groups of the glycoproteins. The channel complex in the presence of epidermal growth factor (EGF) and ATP responded by an increase in protein tyrosine phosphorylation, and the vesicles containing the phosphorylated channels showed a 46% increase in 45Ca2+ uptake. The phosphorylation process was inhibited by the acidic mucins, which also interfered with the binding of EGF to the channel protein. The inhibitory effect was dependent upon the presence of sulfate ester and sialic acid groups, and the loss of the inhibitory capacity occurred following their removal. The results demonstrate the salivary mucins actively participate in the modulation of buccal mucosal calcium channel activity.

Adenosine Triphosphate↗

PubMed1993

Glycosulfatase activity of Porphyromonas gingivalis a bacterium associated with periodontal disease.

Porphyromonas gingivalis, a bacterium implicated in the pathogenesis of periodontal disease, was found to elaborate an extracellular glycosulfatase enzyme. Upon purification by low temperature acetone fractionation, an active enzyme at 60% acetone was obtained which on SDS-PAGE gave a protein band of 37kDa. The glycosulfatase effectively caused desulfation of galactosyl- and lactosylceramide sulfates (pH 5.0) which contain the sulfate ester groups at C-3 of galactose, a well as proteoglycans (pH 5.7-6.2) of gingival tissue which are rich in N-acetylgalactosamine-4-sulfate, but not the sulfated salivary mucin with the sulfate groups at C-6 of galactose and C-6 of N-acetylglucosamine. The results demonstrate for the first time that P. gingivalis displays glycosulfatase activity and that the disruptive action of this enzyme may be a major factor in the etiology of periodontal disease.

Culture Media↗

PubMed1993

Salivary and plasma insulin levels in man.

Changes in plasma and salivary immunoreactive insulin were studied in saliva and blood of 9 healthy subjects following either oral glucose or a test meal ingestion at 30 min intervals for up to 3h. The levels of plasma immunoreactive insulin reached maximum at 60 min with a value of 62.77 +/- 6.52 microU/ml after the glucose load and 62.62 +/- 6.52 microU/ml after the test meal. In the case of salivary immunoreactive insulin levels, the maximum was attained at 90 min with a value of 18.47 +/- 2.68 microU/min after the glucose load and 22.40 +/- 2.01 microU/min after the test meal. Positive linear correlation (r = 0.74) was found between the salivary and plasma immunoreactive insulin levels.

Administration, Oral↗

PubMed1993

Glycosulfatase activity of Helicobacter pylori towards gastric sulfomucin: effect of nitecapone.

A glycosulfatase activity towards human gastric sulfomucin was identified in the extracellular material elaborated by H. pylori, a pathogen implicated in the etiology of gastric disease. The purified enzyme exhibited maximum activity at pH 5.7 in the presence of 0.3% Triton X-100 and 100mM CaCl2, and displayed on SDS-PAGE an apparent molecular weight of 30kDa. The H. pylori glycosulfatase effectively caused desulfation of N-acetylglucosamine-6-sulfate and galactose-6-sulfate of the carbohydrate chains of mucins, as well as that of glucose-6-sulfate of glyceroglucolipids, but was ineffective towards galactosyl -and lactosylceramide sulfates which contain galactose-3-sulfate. The glycosulfatase activity towards human gastric sulfomucin was inhibited by an antiulcer agent, nitecapone, which at its optimal concentration (100 micrograms/ml) caused a 61% decrease in mucin desulfation. The results show that H. pylori through its glycosulfatase activity causes desulfation of sulfated mucins and glyceroglucolipids of the protective mucus layer, and that nitecapone is able to interfere with this detrimental action.

Anti-Ulcer Agents↗

PubMed1992

Glycosulfatase activity of Helicobacter pylori toward gastric mucin.

A glycosulfatase activity toward sulfated gastric mucus glycoprotein was identified in the extracellular material elaborated by H. pylori, a bacteria implicated in the etiology of gastric disease. Upon acetone precipitation, an active enzyme fraction at 64% acetone was obtained which on SDS-PAGE gave a major 30kDa protein band. The H. pylori glycosulfatase exhibited maximum activity (314.8 pmol/mg protein/h) at pH 5.7 in the presence of Triton X-100 and CaCl2, and was capable of removal of the sulfate ester groups situated at C-6 of N-acetylglucosamine, galactose and glucose. However, the enzyme was ineffective toward galactosylceramide and lactosylceramide sulfates which contain the sulfate ester group on C-3 of galactose. The results suggest that H. pylori is capable of overcoming the interference by sulfated mucus glycoprotein with its colonization of gastric mucosa.

Gastric Mucins↗

PubMed1992

Activation of dihydropyridine-sensitive parotid salivary gland calcium channels by epidermal growth factor.

The calcium channel complex of the parotid was isolated from solubilized acinar-cell membranes by affinity chromatography on wheatgerm agglutinin. The channel, after labelling the calcium antagonist-receptor site with [3H]-PN200-100, was reconstituted into phosphatidylcholine vesicles that exhibited active 45Ca2+ uptake. This uptake was independent of sodium and potassium gradients, indicating its electroneutrality. The channels responded in a dose-dependent manner to the dihydropyridine calcium antagonist, PN200-110, which at 0.4 microM exerted a maximal inhibitory effect of 75% on 45Ca2+ uptake; a 46% enhancement in 45Ca2+ uptake occurred with a specific calcium-channel activator, BAY K8644. On epidermal growth-factor (EGF) binding in the presence of ATP, there was an increase in tyrosine phosphorylation of 55 and 170 kDa calcium-channel proteins. Such phosphorylated channels, after reconstitution into vesicles, displayed a 61% greater 45Ca2+ uptake, indicating the involvement of tyrosine kinase in EGF-dependent activation of the calcium channel. The results point towards the importance of EGF in the regulation of calcium homeostasis in salivary gland.

Animals↗

PubMed1992

Mechanism of ebrotidine protection against gastric mucosal injury induced by ethanol.

1. The gastroprotective properties of a new H2-receptor antagonist, ebrotidine, against ethanol-induced mucosal injury was investigated. 2. Groups of rats, with and without indomethacin pretreatment, received intragastrically either a dose of ebrotidine or vehicle only, followed by ethanol given at various intervals up to 4 hr. The gastric mucosa, 30 min after the ethanol challenge, was then subjected to macroscopic and histologic examination, and physicochemical measurements. 3. Ebrotidine at doses of 50 mg and higher per kg body wt effectively prevented the alcohol-induced mucosal injury, even in the presence of indomethacin. The protective effect was demonstrable already at 30 min, reached maximum at 1 hr, and persisted up to 3 hr. 4. Physicochemical analyses established that ebrotidine elicited 30% increase in mucus gel dimension, caused 19-20% increase in glycolipids and phospholipids, and evoked 21% increase in sulfomucin and 18% in sialomucins. As a consequence, the mucus gel viscosity increased by 1.4-fold, H+ retardation capacity by 16%, and hydrophobicity by 65%. 5. The results demonstrate that ebrotidine is a unique H2-antagonist endowed with a remarkable mucosal strengthening capability.

Animals↗

PubMed1992

Identification of human salivary protease activity toward mucin: differences with caries.

A protease activity directed toward high molecular weight salivary mucus glycoprotein was identified in the secretion of human submandibular salivary gland. The protease exhibited maximum activity at pH 7.0-7.4, and following ammonium sulfate fractionation yielded an active enzyme at 60% saturation which on SDS-PAGE gave 48 and 53kDa protein bands. The enzyme exhibited serine-protease properties by showing susceptibility to phenyl methyl sulfonyl fluoride, alpha 1-antitrypsin, and egg white and soybean inhibitors. The protease activity in submandibular saliva of caries-resistant subjects was found to be 3.8-fold greater than that in saliva of caries-susceptible individuals, thus suggesting that the enzyme expression may be linked to the resistance to caries.

Adult↗

PubMed1992

Expression of salivary mucin bacterial aggregating activity: difference with caries.

The low and high molecular weight mucin forms were isolated from saliva of individuals with different caries status and assessed for their bacterial aggregating potential towards S. mutans and S. sanguis. The high molecular weight mucin from both groups exhibited similar protein and carbohydrate content, but the level of covalently bound fatty acids was lower in the caries-resistant group. The mucin from caries-resistant group showed only a weak inhibitory potential, while no inhibitory activity was observed with the mucin of caries-susceptible group. The low molecular weight mucins from both groups, while displaying compositional similarities, showed a marked variation in the bacterial aggregating activity, and the titer of the mucin from caries-resistant group was at least 128-fold greater than that of caries-susceptible group. The results demonstrate that the bacterial aggregating epitope of salivary mucins is expressed to a greater extent in caries-resistant individuals, and that this epitope is apparently more accessible to bacteria in the low molecular weight mucin form.

Dental Caries↗

PubMed1992

Glycosulfatase activity of H. pylori toward human gastric mucin: effect of sucralfate.

Colonization of gastric mucosa by Helicobacter pylori, a bacterium implicated in the etiology of gastric disease, involves the cell surface sulfated glycosphingolipid receptors for the attachment. Evidence has also been obtained recently that sulfated mucus glycoproteins have the ability to interfere with this process. Here, we show that H. pylori displays glycosulfatase activity, and report the specificity of this enzyme toward gastric mucosal sulfated glycoproteins and glycolipids. With 35S-labeled human gastric sulfated mucin as substrate, the enzyme activity was identified in the extracellular material elaborated by the bacterium. The glycosulfatase exhibited maximum activity at pH 5.7 in the presence of Triton X-100 and CaCl2, and gave on SDS-PAGE a protein band of 30 kDa. Specificity studies revealed that the enzyme effectively caused desulfation of N-acetylglucosamine-6-sulfate and galactose-6-sulfate present in carbohydrate chains of gastric mucins, as well as that of glucose-6-sulfate, a constituent of mucus glyceroglucolipids. However, the H. pylori glycosulfatase was ineffective toward galactosyl- and lactosylceramide sulfates which serve as receptors for this bacterium attachment and contain the sulfate ester group at C-3 of galactose. The glycosulfatase activity toward human sulfated gastric mucin was inhibited by sucralfate. The inhibitory effect was proportional to the concentration of sucralfate up to 120 micrograms/ml, at which a 78% decrease in mucin desulfation occurred. The results demonstrate that H. pylori, through its glycosulfatase activity, affects the sulfated mucin and glyceroglucolipid content of the protective mucus layer, and that antiucler drug sucralfate is able to counteract the detrimental action of this enzyme.

Gastric Mucins↗

PubMed1992

Inhibition of Helicobacter pylori glycosulfatase activity toward gastric sulfomucin by nitecapone.

A glycosulfatase activity toward gastric sulfomucin was identified in the extracellular material elaborated by H. pylori. The enzyme exhibited maximum activity at pH 5.7 in the presence of Triton X-100 and CaCl2, and displayed on SDS-PAGE an apparent molecular weight of 30kDa. The H. pylori glycosulfatase effectively caused desulfation of N-acetylglucosamine-6-sulfate and galactose-6-sulfate of the carbohydrate chains of mucins, as well as that of glucose-6-sulfate of glyceroglucolipids, but was ineffective towards galactosyl- and lactosylceramide sulfates which contain galactose-3-sulfate. The glycosulfatase activity towards human gastric sulfomucin was affected by an antiulcer agent, nitecapone, which at its optimal concentration (100 micrograms/ml) caused a 61% inhibition. The results show that H. pylori through its glycosulfatase activity causes desulfation of sulfated mucins and glyceroglucolipids of the protective mucus layer, and that nitecapone is able to interfere with this detrimental action.

Anti-Ulcer Agents↗

PubMed1991

Adrenergic and cholinergic regulation of phospholipid release in sublingual salivary gland in vitro.

1. The role of adrenergic and cholinergic mediators in the regulation of salivary phospholipid secretion was investigated using rat sublingual acinar cells maintained in the presence of [3H]choline. 2. The release of [3H]choline containing phospholipids was enhanced by beta-adrenergic agonist, isoproterenol to a greater extent than the cholinergic agonist, pilocarpine. 3. The effect of isoproterenol on phospholipid release was observed even after 5 min and by 30 min a 1.7-fold increase in secretion occurred, whereas pilocarpine evoked 1.2-fold increase by 30 min with no discernible effect in 5 min. The isoproterenol effect was blocked by alprenolol, and an inhibition of pilocarpine effect was observed with atropine. 4. In the absence of mediators, 82% of secreted labeled phospholipids were represented by phosphatidylcholine (PC), 4.3% by lysophosphatidylcholine (LPC) and 12.1% by sphingomyeline (Sph), those secreted in the presence of isoproterenol showed significantly lower (37%) content of LPC, while those secreted in response to pilocarpine were substantially richer in LPC (2.5 times) and Sph (27%), and contained less (17%) PC. 5. The results provide first evidence for the involvement of adrenergic and cholinergic mediators in the regulation of salivary phospholipid secretion.

Alprenolol↗

PubMed1991

GM1 ganglioside protection against ethanol-induced gastric mucosal injury.

The gastroprotective properties of GM1-ganglioside, an indigenous component of epithelial cell membrane, was investigated. The experiments were conducted with groups of rats with and without indomethacin pretreatment. The animals received intragastrically either a dose of GM1 as emulsion in 5% gum arabic or vehicle alone, followed by ethanol given at various time intervals up to 3 h after the GM1. The animals were sacrificed 30 min after the ethanol dose and their gastric mucosa subjected to macroscopic and histologic assessment, and physicochemical measurements. In the absence of GM1, ethanol caused extensive gastric hemorrhagic lesions which were significantly reduced by pretreatment with GM1 at dose as low as 70 micrograms/100 g body weight. Removal of sialic acid from GM1 led to the loss of gastroprotection. Furthermore, the effect of GM1 was not thwarted by indomethacin. The maximal protection was achieved 1 h following GM1 dose and this protective effect persisted at least 2.5 hr. The results of physicochemical measurements revealed that GM1 was capable of preventing the detrimental effect of indomethacin on the adherent mucus gel dimension, and on its content of sulfo- and sialomucins, protein, and phospholipids. The effects brought by GM1 were also accompanied by a significant (40-60%) increase in mucus gel viscosity, hydrogen ion retardation capacity (35-46%) and hydrophobicity (70-94%). The results indicate that the gastroprotective action of GM1 occurs through the enhancement of the physicochemical characteristics of the mucus layer, and does not appear to be mediated by endogenous prostaglandins.

Animals↗

PubMed1991

Role of sulfation in the processing of gastric mucins.

The role of sulfation in the processing of mucus glycoprotein in gastric mucosa was investigated. Rat gastric mucosal segments were incubated in MEM at various medium sulfate concentrations in the presence of [35S]Na2SO4, [3H]glucosamine and [3H]proline, with and without chlorate an inhibitor of PAPS formation. The results revealed that the mucin sulfation attained maximum at 300 microM medium sulfate concentration. Introduction of chlorate into the incubation medium, while having no effect on the protein synthesis as evidenced by [3H]proline incorporation, caused at its optimal concentration of 2 mM a 90% decrease in mucin sulfation and a 40% drop in mucin glycosylation. Evaluation of mucin molecular forms distribution indicated the predominance of the high molecular mucin form in the intracellular fraction and the low molecular mucin from in the extracellular fraction. Increase in medium sulfate caused an increase in the high molecular weight mucin form in both fractions, and this effect was inhibited by chlorate. Also, higher medium sulfate concentrations led to a higher degree of sulfation in the high molecular weight mucin form, the effect of which was inhibited by chlorate. The results suggest that the sulfation process is an early event taking place at the stage of mucin subunit assembly and is required for mucin polymer formation. Hence, the disturbances in mucin sulfation process could be detrimental to the maintenance of gastric mucus coat integrity.

Animals↗

PubMed1991

Identification of gastric mucosal mucus glycoprotein sulfotransferase.

Sulfation of mucus glycoproteins, reaction catalyzed by Golgi resident sulfotransferase, is an important event in posttranslational processing of gastric mucins. Here we report the purification of mucus glycoprotein sulfotransferase enzyme from the microsomal fraction of rat gastric mucosa. The enzyme was released from the membrane with 0.5% Triton X-100 and precipitated from the 100,000xg supernatant with 90% ice-cold acetone. The enzyme activity (44.7 pmol/mg/45 min) in the precipitate was enriched nearly 10-fold compared to Triton X-100 extract of microsomal membrane (4.2 pmol/mg/45 min). On SDS-PAGE, the enzyme gave a single 43 kDa protein band, which was active towards mucin, but did not catalyze the sulfation of galactosylceramide. The study is the first to report the characteristics of a sulfotransferase enzyme specific for gastric mucin.

Animals↗

PubMed1991

Inhibition of Helicobacter pylori colonization by sulfated gastric mucin.

The role of human gastric mucin in mucosal protection against Helicobacter pylori colonization was investigated. H. pylori cells were incubated with purified intact mucin or its acidic fractions and then examined for their inhibitory capacity of H. pylori attachment to erythrocytes. Titration data established that the inhibitory activity of mucin was associated with its acidic component as the fraction enriched in sialic acid and sulfate showed 16-fold higher inhibitory titer than that of the intact mucin. While the inhibitory titer of acidic mucin fraction was not affected by the removal of sialic acid, the desulfation led to a complete loss of its inhibitory activity, thus pointing towards the importance of sulfate ester groups in this process. The results for the first time point towards the involvement of sulfomucins in the protection of gastric mucosa against colonization by H. pylori.

Animals↗

PubMed1990

The relationship of prostaglandins to cAMP, IgG, IgM and alpha-2-macroglobulin in gingival crevicular fluid in chronic adult periodontitis.

Gingival crevicular fluid, collected from 8 patients with chronic adult periodontitis before and 21 days after root planing and scaling, was analysed for prostaglandin E2, 6KPGF1 alpha, cAMP, IgG, IgM and alpha-2-macroglobulin, and their inter-relationship evaluated. There was a significant decrease in the levels of prostaglandin E2, IgG, IgM and alpha-2-macroglobulin after treatment, whereas the levels of 6KPGF1 alpha and cAMP remained essentially unchanged. The level of prostaglandin E2 decreased by 35%, IgG by 32%, IgM by 90%, and alpha-2-macroglobulin by 79%. There was a significant degree of correlation between prostaglandin E2 and 6KPGF1 alpha and cAMP before treatment but not after, but no correlation between prostaglandin E2 and IgG, IgM and alpha-2-macroglobulin either before or after. This correlation pattern indicates the involvement of E2, prostaglandin 6KPGF1 alpha and cAMP in inflammation in the periodontium. The changes in IgG, IgM and alpha-2-macroglobulin reflect yet another mechanism of host response which appears to be independent of prostaglandins.

6-Ketoprostaglandin F1 alpha↗