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Biomedical subjects

W A Simpson

Publications and source records attributed to W A Simpson.

At least 55 records · Page 3Linked to original sources
PubMed1988

Paget's disease, melanocytic neoplasia and hidradenoma papilliferum of the vulva.

We present a case of invasive melanoma of the vulva with contiguous changes including melanocytic atypia showing, in part, the features of melanocytic dysplasia and, in part, an in situ malignant component and extramammary Paget's disease. A benign hidradenoma papilliferum was also present. We can find no other reported cases of this combination of lesions. The possible histogenesis is discussed and an origin from a common local stem cell is suggested.

Adenoma, Sweat Gland↗
PubMed1988

Adherence of Streptococcus sanguis to conformationally specific determinants in fibronectin.

The adherence of Streptococcus sanguis to specific receptors exposed or deposited at the site of endothelial damage may play an important role in the development of infective endocarditis. Adherence of the Challis strain of S. sanguis to gelatin (or collagen) and gelatin-binding components of plasma was examined with an enzyme-linked immunosorbent assay. S. sanguis adhered poorly to immobilized gelatin and to molecular or fibrillar collagen. However, in the presence of fresh human plasma, the adherence of S. sanguis to all three substrates increased as much as eightfold. Removal of gelatin-binding proteins eliminates the ability of plasma to enhance adherence of S. sanguis to the substrates. Addition of purified human plasma fibronectin (Fn) to the absorbed plasma restored the adherence-promoting ability in a dose-dependent manner. A similar dose-dependent increase in S. sanguis adherence was observed when increasing concentrations of Fn alone were added to the gelatin-coated assay wells. S. sanguis adherence to immobilized fibronectin could not be inhibited by preincubating either the bacteria or the gelatin-coated assay wells with Fn or by including excess soluble Fn in the assay mixture. Studies with peptides purified from trypsin digests of Fn indicated that the 160- to 180-kilodalton (kDa) fragments which retain both the gelatin-binding and the cell-binding regions of the intact molecule support adherence of S. sanguis to gelatin. The 160- to 180-kDa fragments inhibited the interaction of S. sanguis with immobilized Fn. In contrast, intact Fn and the 31-kDa amino-terminal fragment were unable to inhibit the adherence when used in equivalent or greater molar amounts. These in vitro results suggest that in the presence of whole plasma, S. sanguis binds to immobilized gelatin or collagen via Fn bound to the immobilized substrates. Our finding that adherence of S. sanguis to immobilized Fn can occur in the presence of large concentrations of Fn, whether in plasma or purified, indicates that a S. sanguis-binding domain is cryptic in the Fn molecule while in solution and is exposed by a conformational change when the Fn becomes bound to gelatin-coated plastic. The ability of peptide fragments of Fn to inhibit S. sanguis adherence is consistent with this hypothesis.

Bacterial Adhesion↗
PubMed1987

Hybridoma antibodies to the lipid-binding site(s) in the amino-terminal region of fibronectin inhibits binding of streptococcal lipoteichoic acid.

In this report, we present evidence to suggest that streptococci and lipoteichoic acid (LTA) interact with a fatty acid binding site located near the NH2-terminus of fibronectin. The evidence is based on the following observations. Antibodies directed against a synthetic peptide (residues 1-30 of the amino-terminus of fibronectin) reacted with the two thermolysin-generated peptides (24 and 28 kilodaltons [kDa]) that were adsorbed by and eluted from streptococci. The adsorption of the 24- and 28-kDa peptides to streptococci was inhibited by LTA. The two monoclonal antibodies that inhibited the binding of LTA to fibronectin reacted only with the 24- and 28-kDa fragments of fibronectin. Conversely, LTA, as well as lauric acid and oleic acid, blocked the binding of the same monoclonal antibodies to fibronectin. LTA had no effect on the binding of hybridoma antibodies directed against the collagen or cell-binding domain.

Animals↗
PubMed1987

Phenotypic variation of Staphylococcus epidermidis slime production in vitro and in vivo.

Clinical studies performed by us and others have found an association between slime production and strains of coagulase-negative staphylococci that infect indwelling medical devices. By serial low-speed centrifugation of broth cultures we have isolated a stable, weakly adherent strain (RP62A-NA) from a strongly adherent, slime-producing, pathogenic strain of Staphylococcus epidermidis sensu stricto (RP62A, ATCC 35984). We obtained a second strain from RP62A-NA (RP62A-NAR) by serial subculture of glass-adherent cells of RP62A-NA. All three strains had the same pattern of biochemical reactions, antimicrobial susceptibilities, and plasmid analysis. Transmission electron micrograph sections stained with the mucopolysaccharide-specific stain alcian blue demonstrated that the adherent strains RP62A and RP62A-NAR were covered with an extracellular coat of polysaccharide-rich material. In contrast, the nonadherent RP62A-NA strain lacked this external coat. All three strains were used in a mouse model of foreign body infection and a rat model of catheter-induced infective endocarditis. The adherence characteristics of isolates of RP62A and RP62A-NA recovered from experimental animals were relatively stable, although we noted a slight but a significant increase in the adherence of RP62A-NA isolates recovered from the foreign body model. The adherence characteristics of RP62A-NAR isolates recovered from infected animals were variable; in general these isolates were less adherent than the laboratory strain of RP62A-NAR. In both models the 50% infective dose (calculated by the Reed and Muench method) was three times greater for the RP62A-NA strain than for the RP62A strain. The phenotypic expression of slime production is subject to both in vitro and in vivo variation and could play a role in the pathogenesis of foreign body infection.

Animals↗
PubMed1987

Effects of fibronectin and other salivary macromolecules on the adherence of Escherichia coli to buccal epithelial cells.

The effect of saliva and fibronectin (Fn) on the adherence of a type 1 fimbriated strain of Escherichia coli to human buccal epithelial cells was studied. Saliva pretreatment of epithelial cells led to a dose-dependent increase in adherence that was inhibited by alpha-methyl mannoside, which is typical of a type 1 fimbria-mediated event. The molecules responsible for affecting this increased adherence were nondialyzable and were recovered after lyophilization. E. coli adherence was stimulated by individual saliva samples from each of 11 volunteers. Fn inhibited E. coli adherence to saliva-treated buccal cells by more than 60%. Biotinylated E. coli and Fn were reacted with Western blots of whole saliva to identify the receptors that might explain the phenomenon described above. Both E. coli and Fn bound to 57- and 62-kilodalton (kDa) protein bands in Western blots of sodium dodecyl sulfate gels of whole saliva. The binding of E. coli to these bands was inhibited by pretreatment with unlabeled Fn. To study these salivary components, samples of saliva were electrophoresed on sodium dodecyl sulfate gels, strips corresponding to the appropriate molecular weights were cut out, and the proteins were eluted electrophoretically. Material that eluted from strips at 57 and 62 kDa, but not that from a control strip, stimulated E. coli adherence to buccal cells. Alternatively, saliva was fractionated over 100- and 50-kDa cutoff filters. Of the three fractions obtained, only the fraction passing through the 100-kDa filter and retained by the 50-kDa filter stimulated E. coli adherence to buccal cells. This fraction also increased the binding of Fn to buccal cells. These observations suggest the possibility that one or more salivary components bind to the surface of buccal cells and serve as receptors for type 1 fimbriated E. coli. Fn also binds to this isolated material; and it is apparently by these interactions, at least in part, that saliva stimulates and Fn inhibits E. coli adherence. The way in which these interactions may affect bacterial adherence in vivo remains to be elucidated.

Bacterial Adhesion↗
PubMed1986

Isolation and characterization of a 60-kilodalton salivary glycoprotein with agglutinating activity against strains of Streptococcus mutans.

A bacterial agglutinin specific for strains of Streptococcus mutans was isolated from human saliva. Physiochemical analyses showed the agglutinin to be a glycoprotein with a molecular weight of 60,000. The agglutinin aggregated four of the eight strains of Streptococcus mutans tested but did not aggregate the strains of Streptococcus salivarius, Streptococcus sanguis, and Streptococcus mitis tested. Chemical modification of carbohydrate moieties of the agglutinin with sodium metaperiodate had no effect on aggregation, whereas modification of the polypeptide portion with trypsin abolished aggregating activity. A set of five murine hybridoma antibodies was employed to further analyze the agglutinin. Two carbohydrate-specific antibodies, directed against D-mannose and N-acetylgalactosamine moieties, respectively, failed to block agglutinin- or whole saliva-mediated aggregation of S. mutans cells. In contrast, two antibodies directed against pronase-sensitive antigenic sites blocked both agglutinin- and saliva-mediated aggregation of S. mutans cells. Western blot analysis with the agglutinin-specific hybridoma antibodies demonstrated the agglutinin in whole saliva and in artificial tooth pellicles formed on hydroxyapatite beads incubated with saliva. These results suggest that a 60-kilodalton glycoprotein of human saliva is a bacterial agglutinin with specificity for certain strains of S. mutans. They further suggest that aggregation is mediated by polypeptide rather than carbohydrate determinants of the glycoprotein.

Agglutination↗
PubMed1986

Relationship of critical micelle concentrations of bacterial lipoteichoic acids to biological activities.

The critical micelle concentration (CMC) of lipoteichoic acid (LTA) was investigated with two dyes, rhodamine 6G and Coomassie brilliant blue R-250. Both dyes gave similar values for the CMC of LTA. The CMC of LTA from several species of bacteria ranged from 28 to 60 micrograms/ml in phosphate-buffered saline. The CMC values for the LTAs are in the range expected for an amphiphile containing a single, 16-carbon fatty acid residue. Formation of micelles was not detected with deacylated LTA. Salt decreased the CMC of LTA from 380 micrograms/ml in distilled water to 37 micrograms/ml in 0.5 M NaCl. At concentrations greater than the CMC, LTA induced the lysis of sheep erythrocytes and was cytotoxic for Girardi heart cells. The data suggest that LTA in the micellar state may cause disruption of the erythrocyte membrane and may be cytotoxic for cells in culture.

Animals↗
PubMed1986

Release of fibronectin-lipoteichoic acid complexes from group A streptococci with penicillin.

Fibronectin binds to Streptococcus pyogenes, and this binding is inhibited by lipoteichoic acid (LTA). Previous studies have shown that LTA can be released from S. pyogenes by treatment with penicillin. Penicillin released LTA from both S. pyogenes and Staphylococcus aureus; however, the binding of fibronectin correlated with the amount of LTA released only in the case of S. pyogenes. Contrarily, clindamycin decreased the ability of S. aureus to bind fibronectin without affecting the binding of fibronectin to S. pyogenes. Further studies indicated that LTA does not inhibit the binding of fibronectin to S. aureus. Fibronectin bound to S. pyogenes could be released from the cell surface by penicillin. Immunological analysis of the released fibronectin indicated that LTA was the only surface component which could be detected as a soluble complex with the released fibronectin. These studies suggest that LTA plays a central role in the binding of fibronectin to S. pyogenes and is not involved in the binding of fibronectin to S. aureus.

Adhesiveness↗
PubMed1986

Inhibition of the interaction of Streptococcus sanguis with hexadecane droplets by 55- and 60-kilodalton hydrophobic proteins of human saliva.

The effect of salivary secretions on the hydrophobicity of Streptococcus sanguis was investigated. Pretreatment of the bacteria with paraffin-stimulated whole saliva resulted in a 79% inhibition of adhesion to hexadecane droplets. Column chromatography on Sepharose 4B and sodium dodecyl sulfate gel electrophoretic analysis indicated that the inhibitory activity of saliva resided in a fraction containing material of approximately 60,000 molecular weight. The active components, which we have termed the hydrophobic components (HC), bind to octyl-Sepharose beads. Pretreatment of S. sanguis with HC resulted in a dose-dependent inhibition of the streptococcus-hexadecane interaction that reached a maximum of 85%. Furthermore, HC effectively blocked the ability of S. sanguis to adhere to hydroxyapatite beads coated with either whole saliva or HC. Sodium dodecyl sulfate-polyacrylamide gel analysis indicated that HC eluted from octyl-Sepharose consisted primarily of two proteins (60 kDa and 55 kilodaltons) which could be resolved by high-pressure liquid chromatography. Both of these proteins were able to inhibit the binding of S. sanguis to hexadecane in a dose-dependent manner; however, the 60-kilodalton molecule was slightly more effective in this assay. Amino acid analysis of these proteins showed that both proteins contained a high percentage of nonpolar amino acids. These findings suggest that certain components of saliva influence the interaction of S. sanguis with hydrophobic surfaces.

Adhesiveness↗
PubMed1986

Binding of Streptococcus pyogenes to soluble and insoluble fibronectin.

The interaction of soluble and insoluble fibronectin with Streptococcus pyogenes was investigated. Soluble fibronectin bound to S. pyogenes in a dose-dependent and irreversible manner. Lipoteichoic acid competitively inhibited the binding of fibronectin to S. pyogenes but had little effect on the binding of fibronectin to staphylococci or pneumococci. The phase of growth of the streptococci had a slight effect on binding of fibronectin, with optimal binding occurring in the late log phase. S. pyogenes cells bound to fibronectin immobilized on microtiter plates in a dose-dependent and saturable manner. Both soluble fibronectin and lipoteichoic acid inhibited the binding of streptococci to immobilized fibronectin, suggesting that streptococci interact with soluble and insoluble fibronectin in a similar manner. Antibodies to fibronectin blocked the attachment of streptococci to immobilized fibronectin, whereas normal serum had no effect. Adherence of streptococci to buccal epithelial cells was inhibited by antibodies to fibronectin, but not by normal sera or by antibodies to buccal epithelial cells. The data suggest that lipoteichoic acid on the surface of S. pyogenes binds to fibronectin exposed on the host cell and that such binding mediates the attachment of streptococci to host cells.

Adhesiveness↗
PubMed1986

Immunochemical and ultrastructural mapping of the gelatin-binding and cell-attachment regions of human plasma fibronectin with monoclonal antibodies.

Monoclonal antibodies against fibronectin were used to locate the gelatin-binding and cell-attachment regions of plasma fibronectin at an ultrastructural level. A total of 23 hybridomas were generated using mice immunized with either intact fibronectin or a 40 000 Mr gelatin-binding fibronectin fragment. One of these antibodies (D9b) strongly inhibited the interaction of radio-labelled fibronectin with gelatin. Another antibody (IB10) inhibited the attachment of Chinese hamster ovary (CHO) cells to a fibronectin substratum by 99%. Both of these antibodies were purified by affinity chromatography on columns of fibronectin-Sepharose and were then incubated with soluble fibronectin to form antigen-antibody complexes. The complexes were separated from free antibody on a column of Sephadex G-200 and were prepared for electron-microscopic examination by spraying on mica discs and rotary shadowing with platinum. As determined by this method, the fibronectin molecules measured 124 +/- 1.7 nm in length. Monoclonal antibody IB10 was visualized as a globular projection 40 +/- 1.4 nm from one end of the fibronectin filament. Monoclonal antibody D9b, on the other hand, was visualized as a globular projection at or near one or both ends of the molecule. These data provide the first morphological localization of the gelatin-binding and cell-attachment regions of fibronectin and indicate that further studies using monoclonal antibodies directed toward other epitopes should shed light not only on function but also on the tertiary and quaternary structure of the fibronectin molecule.

Animals↗
PubMed1985

Human plasma fibronectin inhibits adherence of Streptococcus pyogenes to hexadecane.

The effect of human plasma fibronectin on the adherence of Streptococcus pyogenes to hexadecane droplets was investigated. Fibronectin blocked the adherence of streptococci to hexadecane in a dose-dependent manner. The inhibitory effect resulted from the binding of fibronectin to the streptococcal cells; radiolabeled fibronectin failed to bind to the hexadecane but bound readily to untreated streptococci. Chemical treatments of streptococci that decreased streptococcal binding of fibronectin also decreased their binding to hexadecane. Pretreatment of fibronectin with lipoteichoic acid blocked the binding of fibronectin to streptococci and abolished its ability to inhibit streptococcal adherence to hexadecane in a dose-related manner. In contrast, wheat germ agglutinin, which binds to N-acetylglucosamine on the surface of S. pyogenes cells, failed to alter hexadecane adherence. The data suggest that fibronectin binds to lipoteichoic acid on the surface of the streptococci, thereby preventing lipoteichoic acid from interacting with the hexadecane phase.

Alkanes↗
PubMed1985

Role of fibronectin in attachment of Streptococcus pyogenes and Escherichia coli to human cell lines and isolated oral epithelial cells.

We studied the binding of cells of Streptococcus pyogenes and mannose-sensitive Escherichia coli to human fibroblast cell lines and isolated buccal epithelial cells in relation to the cell-associated endogenous or exogenous fibronectin of the host cells. The degree of bacterial binding to cell lines correlated directly with the content of endogenous fibronectin on the surface of the cultured cells, although the correlation was better with S. pyogenes than with E. coli. The addition of exogenous plasma fibronectin to the cell lines or oral epithelial cells enhanced binding of S. pyogenes but suppressed binding of mannose-sensitive E. coli. These findings are consistent with the notion that exogenously acquired fibronectin on the surface of host cells modulates bacterial adherence by providing attachment sites for certain pathogens, such as S. pyogenes, and by blocking receptors for others, such as mannose-sensitive E. coli.

Adhesiveness↗
PubMed1985

Binding of fibronectin to human buccal epithelial cells inhibits the binding of type 1 fimbriated Escherichia coli.

The interaction of purified human plasma fibronectin (FN) with human buccal epithelial cells was studied. Maximal binding of FN occurred at pH 5. The majority of the binding was specific and reversible. The binding of FN to buccal cells was saturable, reaching a maximum when 10(5) buccal cells were incubated with approximately 200 micrograms of radiolabeled protein per ml. The adherence of a type 1 fimbriated strain of Escherichia coli to buccal epithelial cells was inhibited by the addition of FN in a dose-related manner. Our results indicate that exogenous FN can bind to human buccal epithelial cells and block the attachment of a type 1 fimbriated strain of E. coli.

Adhesiveness↗
PubMed1985

Protection against Escherichia coli-induced urinary tract infections with hybridoma antibodies directed against type 1 fimbriae or complementary D-mannose receptors.

Hybridoma antibodies directed against quaternary structural epitopes of the type 1 fimbrial adhesin of Escherichia coli or against D-mannose, the sugar determinant in the complementary host cell receptor, prevented the attachment of mannose-sensitive E. coli to various eucaryotic cells. Passive intraperitoneal administration of the fimbria-specific or D-mannose-specific antibodies protected mice against retrograde colonization with mannose-sensitive E. coli instilled into their urinary bladders. Monoclonal antibodies directed against fimbrial subunits rather than quaternary structural epitopes or against N-acetylgalactosamine rather than D-mannose residues lacked protective activity. These studies provide evidence that bacterial colonization can be blocked or interrupted by antibodies directed against either the adhesin or the complementary host cell receptor of pathogenic microorganisms.

Adhesiveness↗
PubMed1985

Adherence of coagulase-negative staphylococci to plastic tissue culture plates: a quantitative model for the adherence of staphylococci to medical devices.

The adherence of coagulase-negative staphylococci to smooth surfaces was assayed by measuring the optical densities of stained bacterial films adherent to the floors of plastic tissue culture plates. The optical densities correlated with the weight of the adherent bacterial film (r = 0.906; P less than 0.01). The measurements also agreed with visual assessments of bacterial adherence to culture tubes, microtiter plates, and tissue culture plates. Selected clinical strains were passed through a mouse model for foreign body infections and a rat model for catheter-induced endocarditis. The adherence measurements of animal passed strains remained the same as those of the laboratory-maintained parent strain. Spectrophotometric classification of coagulase-negative staphylococci into nonadherent and adherent categories according to these measurements had a sensitivity, specificity, and accuracy of 90.6, 80.8, and 88.4%, respectively. We examined a previously described collection of 127 strains of coagulase-negative staphylococci isolated from an outbreak of intravascular catheter-associated sepsis; strains associated with sepsis were more adherent than blood culture contaminants and cutaneous strains (P less than 0.001). We also examined a collection of 84 strains isolated from pediatric patients with cerebrospinal fluid (CSF) shunts; once again, pathogenic strains were more adherent than were CSF contaminants (P less than 0.01). Finally, we measured the adherence of seven endocarditis strains. As opposed to strains associated with intravascular catheters and CSF shunts, endocarditis strains were less adherent than were saprophytic strains of coagulase-negative staphylococci. The optical densities of bacterial films adherent to plastic tissue culture plates serve as a quantitative model for the study of the adherence of coagulase-negative staphylococci to medical devices, a process which may be important in the pathogenesis of foreign body infections.

Animals↗
PubMed1983

Antiadhesive properties of a quaternary structure-specific hybridoma antibody against type 1 fimbriae of Escherichia coli.

The relationship between the structure and biological function of type 1 fimbriae of Escherichia coli was investigated using a set of monoclonal antibodies directed against conformation-specific antigenic determinants. Of three monoclonal antibodies tested, only one (clone CD3) prevented adhesion of the vaccine strain to epithelial cells or guinea pig erythrocytes. The antibody produced by CD3, but not that produced by the other two hybridoma clones (AA8 and GG1), precipitated isolated fimbriae by double diffusion in agar gel and was shown to bind in a highly discrete, periodic manner along the length of each of the fimbriae by immunoelectron microscopy. Immunoelectroblots of type 1 fimbrial subunits and polymers electrophoresed in SDS-gels indicated that the antibodies in AA8 and GG1 reacted only with fimbrial monomers (mol wt 17,000), whereas the antibody in CD3 reacted only with polymers of mol wt 102,000 (hexamers) or higher. ELISA inhibition assays demonstrated that dissociated fimbrial subunits lost their reactivity with antibody CD3 but gained reactivity with antibodies AA8 and GG1. Conversely, when allowed to reassemble in vitro in the presence of 5 mM MgCl2, the reassembled fimbriae lost their reactivity with antibodies AA8 and GG1 but regained reactivity with antibody CD3. These results demonstrated that certain antigenic epitopes are dependent on quaternary structural determinants, whereas others are independent of quaternary fimbrial structure and also are inaccessible for antibody binding in fimbriae once they have been assembled. These monoclonal antibodies should prove useful in studies of the structural determinants of the biological function of type 1 fimbriae as well as in studies of fimbrial synthesis, transport, and assembly.

Adhesiveness↗