Biochemical changes in (glyco)proteins of saliva of epileptic patients.
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Biochemical Phenomena↗
Biomedical subjects
E C Veerman
Publications and source records attributed to E C Veerman.
Adherence of Streptococcus gordonii HG 222 in the presence of saliva.
The influence of the presence of saliva from different salivary glands on the adherence of Streptococcus gordonii strain HG 222 to saliva-coated polystyrene surfaces was tested. In the presence of undiluted parotid saliva or diluted whole, submandibular and sublingual saliva the adherence of HG 222 was enhanced by the formation of small aggregates on the attachment surface. In the presence of undiluted whole, submandibular and sublingual saliva large aggregates were formed and the adherence to saliva-coated polystyrene surfaces was inhibited. Adherence in the presence of whole saliva compared to adherence in buffer was decreased when lower densities of bacterial suspension were used, although in this case in the presence of whole saliva smaller bacterial aggregates were formed. In conclusion, these results suggest that the presence of saliva in solution may both enhance and decrease the adherence of S. gordonii HG 222 to saliva-coated polystyrene surfaces, partly depending on the size of bacterial aggregates that are formed in the presence of saliva.
Adult↗
Protein, albumin and cystatin concentrations in saliva of healthy subjects and of patients with gingivitis or periodontitis.
Salivary protein, albumin and cystatin concentrations were investigated in subjects with a healthy periodontium and in patients with gingivitis or periodontitis. Protein and albumin concentrations in saliva of individuals with gingivitis or periodontitis were significantly increased compared with healthy subjects. Salivary protein and albumin appeared to be positively correlated in all the groups, which suggests that the increase in salivary protein concentration in subjects with gingivitis or periodontitis is caused by leakage of plasma proteins. Cystatin concentrations in saliva of subjects with periodontitis were significantly increased when compared with the healthy group and the gingivitis group (p < 0.01). In the gingivitis and periodontitis group, salivary cystatin was only weakly correlated with albumin concentrations, which suggests that the increased salivary cystatin activity found in subjects with gingivitis and periodontitis is derived from sources other than plasma.
Adolescent↗
Interaction of the salivary glycoprotein EP-GP with the bacterium Streptococcus salivarius HB.
The interaction of the human salivary glycoprotein EP-GP with a number of oral bacterial species, following incubation with human whole saliva, has been investigated. EP-GP could be detected with a specific monoclonal antibody, by means of ELISA or by electrophoresis in combination with Western Transfer. The results indicated that EP-GP is bound only by Streptococcus salivarius, and not by the other tested strains of bacteria, Actinomyces viscosus, A. naeslundii, Actinobacillus actinomycetemcomitans, Bacteroides fragilis, S. gordonii, S. oralis, S. sanguis, S. mitis, S. mutans, S. sobrinus, S. rattus, S. constellatus, and S. anginosus. Binding of EP-GP to S. salivarius is mediated by a protein-protein interaction, which was found to be pH-dependent with a maximum binding between pH 5 and 6. For further characterization of the binding of EP-GP to S. salivarius, four mutants were tested, each of them lacking different cell wall antigens. EP-GP was bound to all mutants in amounts comparable with the wildtype, in spite of the different surface antigen compositions. We were able to identify a 27-kD EP-GP binding protein, by extraction of S. salivarius-cell wall antigens and electrophoretic techniques. In addition to EP-GP, S. salivarius also bound two other salivary proteins, namely, secretory IgA and low-molecular-weight mucin (MG-2).
Animals↗
[Saliva and saliva substitutes].
Saliva is a mixture of secretions of both the large salivary glands (glandula sublingualis submandibularis and parotis) and the minor salivary glands of the palate, tongue, lips and cheeks. The rheological flow of the separate glandular salivas differs remarkably. Sublingual saliva is both more viscous and elastic than the other glandular salivas. The viscosity of parotid saliva, which is of importance during eating (digestion), is equal to the viscosity of water. Utilization of the combination of viscosity and elasticity may improve new saliva substitutes and of frequent applications of saliva substitutes by xerostomia patients may reduce. Besides, the utilization of correct visco-elastic properties of saliva substitutes may lead to better moistening and protective properties.
Humans↗
Isolation of different high-Mr mucin species from human whole saliva.
By using CsCl-density-gradient ultracentrifugation, two high-Mr mucin species were isolated from human whole saliva, having buoyant densities in 0.2 M-guanidinium chloride of approx. 1.56 g/ml (pool IA) and 1.48 g/ml (pool IIA). Analytical density-gradient centrifugation of submandibular, sublingual, labial and palatal saliva, followed by immunochemical analysis with anti-mucin monoclonal antibodies, indicated immunochemical and physicochemical similarities between the high-density mucins of pool IA and mucins from palatal salivary glands. Chemical analysis indicated that the putative palatal mucin was rich in sulphate, but poor in sialic acid. The lower-density mucins of pool IIA equated with the high-Mr mucins of submandibular-sublingual saliva, both immunochemically and physicochemically (buoyant density).
Antibodies, Monoclonal↗
Changes in saliva of epileptic patients.
Unstimulated whole saliva samples of 27 indoor epileptic patients were studied on their protein composition using biochemical and immunochemical methods. A number of salivary proteins appeared at least partially to be hydrolyzed. In a number of saliva samples the concentration of carbohydrate-containing isoenzymes of amylase was reduced. In addition, the concentration of the 20 kD glycoprotein EP-GP was reduced by 60%. Sialic acid, the terminal sugar of the glycoproteins and mucins, was released for about 50% and in three salivas even nearly completely. Moreover, sialic acid- and fucose-containing epitopes could hardly be detected by monoclonal antibodies to human salivary mucins. As a consequence of this hydrolytic breakdown the saliva mediated aggregation of two S. sanguis strains had been reduced. In contrast, the aggregation of S. oralis had been maintained.
Adolescent↗
Influence of saliva on aggregation and adherence of Streptococcus gordonii HG 222.
The influence of saliva on the aggregation and adherence of Streptococcus gordonii HG 222 was studied. The aggregation was measured spectrophotometrically, and the adherence of S. gordonii to microtiter plate wells was measured in an enzyme-linked immunosorbent assay system. The aggregation of HG 222 was induced primarily by mucous saliva, whereas the adherence of HG 222 to microtiter plates was mediated by both mucous and serous saliva. Fractions of submandibular saliva, obtained by gel filtration and containing low-molecular-weight mucins (MG-2), induced both bacterial aggregation and adherence. Purified MG-2 induced aggregation and promoted adherence, whereas high-molecular-weight mucins (MG-1) did not. After incubating clarified human whole saliva with HG 222, only MG-2, and not MG-1, was bound by the bacteria. Proline-rich proteins (PRPs) and proline-rich glycoprotein (PRG) promoted the adherence of HG 222. These proteins in solution bound to HG 222 but did not induce aggregation of the bacterial cells. PRPs and PRG in solution were not able to inhibit adherence to microtiter plate wells coated with the same components. Purified alpha-amylase hardly promoted adherence to microtiter plates but, in the soluble state, readily bound to HG 222. In conclusion, these results indicate that the aggregation of S. gordonii HG 222 is mediated primarily by MG-2. These mucins also promote adherence. Several other salivary components, such as PRPs and PRG, are also involved in the adherence of HG 222.
Adult↗
Immunochemical analysis of high molecular-weight human salivary mucins (MG1) using monoclonal antibodies.
Using four Mabs with different specificities for salivary mucins, an ELISA has been developed in which human whole saliva, glandular salivas, salivary protein fractions and purified, high molecular-weight, mucin fractions (MG1) isolated from human submandibular and sublingual glandular tissues have been immunochemically analysed. All four Mabs reacted with MG1s. Three of them reacted with the purified, low molecular-weight salivary mucins (MG2). None was reactive with parotid saliva. MG1 preparations isolated from submandibular and sublingual glandular tissues of one and the same individual displayed different patterns of reactivity with these Mabs, indicating that they differ immunochemically. Analysis of the MG1s in salivas derived from individual salivary glands showed differences in immunochemical composition. These results indicate that the MG1 fraction in human whole saliva consists of several immunochemically different species.
ABO Blood-Group System↗
Detection of proteins related to a salivary glycoprotein (EP-GP). Concentrations in human secretions (saliva, sweat, tears, nasal mucus, cerumen, seminal plasma).
With a highly specific monoclonal antibody against a previously isolated and characterized human salivary 19-20-kDa glycoprotein, designated as extra-parotid glycoprotein [Rathman et al. (1989) J. Biol. Buccale 17, 199-208], a common epitope was detected on proteins in several excretory human body fluids. With a quantitative ELISA the EP-GP epitope was measured in widely different concentrations in several secretory human body fluids in the descending order of seminal plasma much greater than tears approximately nasal mucus approximately sweat much greater than saliva. Crossreactivity was also observed in cerumen but not in milk, cerebrospinal fluid, blood plasma and urine. The relative amount of EP-GP in the positively reacting secretions was however, in the same order in each fluid per mg of protein on an average of 1% of the total protein amount. The EP-GP-epitope bearing proteins found in the various human secretions were further characterized by means of electrophoresis and immunoblotting. The molecular masses and the isoelectric points of the proteins in the different secretions display strong resemblance to values found for the salivary glycoprotein EP-GP (molecular masses 19 and 20 kDa; pI values between 4.8 and 5.4). All these findings point to the presence of proteins related to EP-GP in human secretions other than saliva.
Antibodies, Monoclonal↗
Adsorption to hydroxyapatite of partially deglycosylated human salivary mucins in competition with phosvitin and phytate.
The effect of phosvitin and phytate on the binding of native as well as partially deglycosylated human whole salivary mucins (HWSM) to hydroxyapatite was studied. Native HWSM preadsorbed onto hydroxyapatite was completely desorbed in the presence of greater than 500 micrograms/ml phosvitin. In contrast, in similar experiments, asialo-HWSM was desorbed approximately 10%. Desorption of preadsorbed asialo-afuco-HWSM in the presence of 1 mg/ml phosvitin was approximately 20%. Further deglycosylation of HWSM resulted in preparations which, after preadsorption to hydroxyapatite, were not desorbed upon subsequent incubation with phosvitin. With phytate, a less effective competitor of HWSM for the hydroxyapatite surface, essentially the same results were obtained, i.e. increase in deglycosylation of HWSM was concomitant with decrease in desorption by phytate. Using other incubation conditions (preadsorption of a phosphocompound, and simultaneous incubation of HWSM and phosphocompounds) essentially the same conclusion was obtained. The data indicate that the ability of salivary mucins to absorb to hydroxyapatite in competition with phosphocompounds appears to be enhanced by deglycosylation.
Asialoglycoproteins↗
Influence of the blood group reactive substances in saliva on the aggregation of Streptococcus rattus.
The interaction of blood group reactive substances in saliva with bacteria was investigated by testing saliva from persons with different blood groups in a bacterial aggregation assay with Streptococcus rattus HG 59, originally S. rattus BHT. For blood group A, saliva from 10 persons out of 11 aggregated S. rattus and for blood group O, saliva from 10 persons out of 16 aggregated S. rattus. For blood group B, saliva from 6 persons out of 8 aggregated S. rattus weakly and the average aggregation activity of blood group B was much lower than for blood group A or O. In addition, saliva from 3 non-secretors did not aggregate S. rattus. The role of blood group antigens in bacterial aggregation was confirmed by inhibition studies with blood group specific sugars and various other sugars. GalNAc, specific for blood group A, inhibited bacterial aggregation by saliva whereas D-galactose, specific for blood group B, and D-fucose, specific for blood group O, did not. In addition, sialic acid, a major terminal sugar residue in mucins, also inhibited the bacterial aggregation. This study shows that the blood group and secretor status of a person may influence the interaction of saliva with bacteria in the oral cavity.
Acetylgalactosamine↗
Saliva-induced aggregation of oral streptococci and the influence of blood group reactive substances.
Aggregation of strains of Streptococcus rattus, Strep. mutans and Strep. salivarius by saliva from individuals of blood groups A, B and O was investigated. Blood group A salivas had a significantly higher aggregation activity with Strep. rattus than blood group B salivas (P less than 0.05). However, Strep. mutans and Strep. salivarius were better aggregated by blood group B saliva and this was significant for Strep. mutans (P less than 0.05). For all three strains, the variance within blood group O was too large to give significant differences with either blood group A or B. The blood group A-specific carbohydrate, N-acetyl-D-galactosamine, inhibited aggregation of Strep. rattus, but not of the other strains. The blood group B-specific carbohydrate, D-galactose, inhibited aggregation of Strep. mutans but not of Strep. rattus or Strep. salivarius. L-Fucose, specific for blood group O failed to inhibit aggregation of any of the three strains. These findings suggest that blood group-specific substances may be involved in bacterial aggregation.
ABO Blood-Group System↗
Coadsorption onto hydroxyapatite of human salivary mucins and phosvitin, a phosphoprotein from egg yolk.
The adsorption of human salivary mucins (HWSM, 0.4 mg/ml) onto hydroxyapatite (HAP) was studied in the presence of varying amounts of the phosphoprotein phosvitin by three different procedures. a. Preadsorption of HWSM onto HAP for 20 h, followed by 4 h coadsorption with phosvitin, resulted in a decrease of 50% in HWSM binding to HAP with 0.3 mg/ml phosvitin and a complete desorption with 1.0 mg/ml phosvitin. b. Preincubation of HAP with phosvitin for 20 h, followed by 4 h coadsorption with HWSM, resulted in decrease of 50% in HWSM binding to HAP with 0.15 mg/ml phosvitin and the adsorption of HWSM was prevented completely with 1.0 mg/ml phosvitin. c. Simultaneous incubation of HWSM and phosvitin gave the least adsorption of HWSM to HAP: a decrease of 50% with as little as 0.025 mg/ml phosvitin and a nearly complete desorption with 0.3 mg/ml phosvitin. Similarly, the adsorption of phosvitin was strongly inhibited by HWSM after either simultaneous adsorption or preadsorption with HWSM. However, after preincubation of HAP with phosvitin, desorption of phosvitin by HWSM was not achieved. Release of phosphate increased by preadsorption with HWSM followed by incubation with phosvitin, but was lowered by about 50% after preadsorption with phosvitin. After simultaneous incubation of HAP with both species, the adsorption did not result in release of phosphate ions. Calcium release was only substantial when phosvitin was in excess in solution. The smallest release of calcium ions was observed when HAP was preincubated with phosvitin, followed by coadsorption with HWSM.
Adsorption↗
Characterization of monoclonal antibodies to human salivary (glyco) proteins. Cellular localization of mucin, cystatin-like 14 kD protein and 20 kD glycoprotein in the human submandibular gland.
Using the hybridoma technique, monoclonal antibodies (Mabs) have been produced against three different types of human salivary proteins: high molecular weight mucin, a 20 kD glycoprotein and a 14 kD protein, identified as a member of the cystatin family. The Mabs appeared to be highly specific to their antigen in Elisa and immunoblotting tests. The Mabs were of the IgG-1 (against 20 kD glycoprotein) and IgM (against 14 kD protein and mucin) type. For the 14 kD protein and the 20 kD glycoprotein it was demonstrated that they are present mainly in submandibular-sublingual saliva. None of the antigens studied could be localized distinctly in the human parotid gland. In the submandibular gland, the three proteins have a different pattern of localization. The mucins have been detected particularly in the apical part of the mucous acinar cells, the 20 kD glycoprotein mainly in the serous acinar cells and the 14 kD protein in both serous acinar cells and striated duct cells.
Antibodies, Monoclonal↗
Comparison of a salivary 14 kD protein displaying cysteine proteinase inhibitory activity with other salivary cystatins.
An acidic protein with a molecular weight of 14 kD and a cysteine proteinase inhibitory activity was isolated from human whole salive. It was classified therefore as a cystatin. Fractionation of whole saliva by FPLC ion-exchange chromatography, followed by immunochemical analysis with monoclonal antibodies, indicated that this protein, designated 14 kD/E8B, eluted in a single peak, which comprised 6% of the total salivary cystatin-activity. The 14 kD/E8B cystatin had nearby 5 times less apparent affinity for papain than chicken egg cystatin. In addition to 14 kD/E8B cystatin, four other cystatins were separated, having pI values of 4.5 and 4.7. One of these appeared together with amylase. Furthermore a protein having a pI of 6.7 was isolated, which accounted for approximately 50% of the total cystatin activity present in whole saliva. The total cystatin activity in unstimulated whole saliva showed at least a three fold difference between individuals. Specific cystatin activity, based on protein content, was highest in saliva derived from the submandibular gland. In addition, cystatin activity could be detected in sublingual and labial saliva, whereas parotid saliva displayed little activity and palatinal saliva no activity at all.
Chemical Fractionation↗
Isolation and characterization of three non-mucinous human salivary proteins with affinity for hydroxyapatite.
The isolation and chemical analysis of three human salivary proteins is reported. Using cation-exchange, FPLC anion-exchange chromatography and gel filtration, three human salivary proteins with affinity for hydroxyapatite were isolated, having molecular weights of 60 kD, 20 kD and 14 kD, respectively. The 60 kD protein was identified as albumin, and the 14 kD protein as a member of the cystatin family. Isoelectric focusing of the 14 kD protein revealed a single band, having an isoelectric point (pl) of 4.7. The third protein, not described yet, is appearing as a 20 kD doublet on SDS-PAGE. Isoelectric focusing resolved the 20 kD protein into four bands having pl's of 4.8, 5.0, 5.2 and 5.4, respectively. All bands were recognized by monoclonal antibodies to the 20 kD protein indicating that these four protein bands share the same epitope. The 20 kD protein is a glycoprotein with a carbohydrate content of 13% and a molar ratio of Fuc: Man: Gal: GlcNAc: NeuAc = 3.4:2.6:2.9:4.0:0.4. All three proteins bind strongly to a hydroxyapatite-based HPHT column at pH 6.0. With increasing pH, the binding diminished, especially of the 14 kD protein.
Albumins↗
Viscosity of human salivary mucins: effect of pH and ionic strength and role of sialic acid.
The viscosity of isolated human salivary mucins has been studied as a function of shear rate, mucin concentration, pH, and ionic strength. At neutral pH, viscosity increased proportionally with mucin concentrations between 0 and 14 mg/ml. Increasing the ionic strength from 35 to 235 mM resulted in an approximately 50% decrease in specific viscosity. A part from the ionic strength effect, no specific effect of calcium ions was observed. Under low ionic strength conditions, viscosity of mucin solutions reached a maximum value at pH 4.2. The extent of the viscosity increase was dependent on ionic strength, mucin concentration and shear rate. Increase of the ionic strength up to 200 mM almost completely abolished the pH-optimum, suggesting that electrostatic interactions underlie the pH-dependent behaviour. The position of the pH-optimum was not changed upon desialization of the mucin, indicating that terminal sialic acid residues do not determine the pH-dependence of human salivary mucin viscosity.
Humans↗