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Biomedical subjects

W A Simpson

Publications and source records attributed to W A Simpson.

At least 73 records · Page 4Linked to original sources
PubMed1983

Adherence of group A streptococci to fibronectin on oral epithelial cells.

The possibility that fibronectin on the surface of oropharyngeal cells may serve as a receptor for the binding of group A streptococci (Streptococcus pyogenes) was investigated. Purified human plasma fibronectin inhibited the adherence of group A streptococci to oral epithelial cells in a dose-dependent manner. The relative amounts of fibronectin available on oral epithelial cells correlated closely with the ability of these cells to bind streptococci. Group A streptococci agglutinated latex beads containing covalently linked fibronectin on their surface, and this agglutination could be inhibited by lipoteichoic acid, the adhesion that mediates attachment of group A streptococci to epithelial cells. Gelatin and the alpha 1 chain of type I collagen partially inhibited both the adherence of streptococci to oral epithelial cells and the binding of radiolabeled fibronectin to streptococci; however, the purified fibronectin-binding peptide of collagen, alpha 1 (I)CB7, inhibited neither. The binding of radiolabeled fibronectin to streptococci was inhibited by lipoteichoic acid. These results suggest that fibronectin on oral epithelial cells serves as a lipoteichoic acid-sensitive receptor for group A streptococci.

Agglutination↗
PubMed1983

Experimental foreign body infections in mice challenged with slime-producing Staphylococcus epidermidis.

The virulence of two previously described Staphylococcus epidermidis strains was examined in an experimental model of foreign body infection in mice. Animals challenged with the slime-producing strain developed three times as many infections as animals challenged with the strain that did not produce slime (P less than 0.001). Bacterial isolates recovered from the infected sites retained the characteristics of the inoculated strain. Animals without foreign bodies but challenged in a similar manner with either staphylococcal strain did not become infected. Thus, the presence of a foreign body predisposed the animals to S. epidermidis infection. These results indicate that the production of slime by S. epidermidis is a stable characteristic retained after animal passage and may be important in the pathogenesis of these infections.

Adhesiveness↗
PubMed1983

Interaction of human plasma fibronectin with cariogenic and non-cariogenic oral streptococci.

The interaction of purified human plasma fibronectin (Fn) with bacteria was studied with a variety of oral streptococci. Each of the strains of Streptococcus mutans, Streptococcus salivarius, Streptococcus sanguis, and Streptococcus mitis tested was aggregated by Fn to various degrees, depending on the concentration of Fn added to the test mixtures. Binding assays performed with radiolabeled Fn and various strains of streptococci demonstrated various capabilities to bind Fn, and the amount of Fn bound by each strain was paralleled by its Fn-induced aggregation, with S. mutans 6715 giving the highest values in both assays. Because of the avid binding of Fn by certain strains of potentially cariogenic streptococci, we investigated the possibility that Fn may be present in human saliva and may be adsorbed from saliva onto artificial tooth pellicles. Immunoreactive Fn was detected in paraffin-stimulated whole saliva by enzyme-linked immunosorbent assays of saliva adsorbed onto gelatin-coated cuvettes and by immunoelectroblots (Western blots) of salivary components separated on sodium dodecyl sulfate-polyacrylamide slab gels. Furthermore, immunoreactive Fn was found to be present in artificial tooth pellicles formed by incubating hydroxyapatite beads with whole human saliva. These results demonstrate that certain strains of oral streptococci bind to and are aggregated by Fn. The presence of Fn in artificial tooth pellicles suggests that this macromolecule may play a role in the attachment of potentially cariogenic and other oral streptococci to dental tissues.

Fibronectins↗
PubMed1983

Adherence of streptococcus pyogenes, Escherichia coli, and Pseudomonas aeruginosa to fibronectin-coated and uncoated epithelial cells.

The relationship between the variability in the fibronectin (Fn) content on human buccal epithelial cells and the capacity of the cells to bind gram-positive (Streptococcus pyogenes) or gram-negative (Escherichia coli or Pseudomonas aeruginosa) bacteria was investigated. Adhesion experiments performed with mixtures of epithelial cells and mixed suspensions of either S. pyogenes and E. coli or S. pyogenes and P. aeruginosa exhibited three major populations of buccal cells: one of these was able to bind S. pyogenes (gram positive) but neither of the gram-negative bacteria; a second population was able to bind the gram-negative but not the gram-positive bacteria; and a third was able to bind various numbers of both types of organisms. Further adhesion experiments performed with a mixture of epithelial cells and a mixed suspension of S. pyrogens, E. coli, and fluoresceinconjugated methacrylate beads coated with immune immunoglobulin G directed against Fn revealed that the epithelial cells recognizing the gram-positive bacteria were rich in Fn, whereas those recognizing the gram-negative organisms were poor in Fn. Immunoelectron microscopy confirmed that cells of S. pyogenes bound to epithelial cells coated with Fn, whereas cells of E. coli bound to epithelial cells lacking Fn. These results suggest that Fn on the surfaces of epithelial cells may modulate the ecology of the human oropharyngeal cavity, especially with respect to the colonization of these surfaces by pathogenic gram-negative or gram-positive bacteria.

Adsorption↗
PubMed1983

Binding of streptococcal lipoteichoic acid to fatty acid-binding sites on human plasma fibronectin.

The ability of Streptococcus pyogenes lipoteichoic acid and palmitic acid to bind to purified human plasma fibronectin was investigated. Initial studies indicated that intact fibronectin formed soluble complexes with lipoteichoic acid, resulting in a change in the mobility of fibronectin in an electrical field. Fibronectin covalently linked to agarose beads bound radiolabeled lipoteichoic acid in the acylated form but not in the deacylated form. An 18-M excess of fibronectin inhibited binding of lipoteichoic acid to the immobilized protein by 92%. Fibronectin-bound [(3)H]lipoteichoic acid could be specifically eluted with unlabeled lipoteichoic acid, as well as by fatty acid-free serum albumin. Serum albumin, which is known to contain fatty acid-binding sites capable of binding to the lipid moieties of lipoteichoic acid, inhibited the binding of lipoteichoic acid to fibronectin in a competitive fashion. The fibronectin-bound lipoteichoic acid could be eluted by 50% ethanol and various detergents but not by 1.0 M NaCl, various amino acids, or sugars. Similarly, radiolabeled palmitic acid adsorbed to fibronectin could be eluted with 50% ethanol but not with 1.0 M NaCl. Fibronectin adsorbed to a column of palmityl-Sepharose was eluted with 50% ethanol in 0.5% sodium dodecyl sulfate but not with 1.0 M NaCl or 1% sodium dodecyl sulfate alone. The binding of lipoteichoic acid to fibronectin followed first-order kinetics and was saturable. A Scatchard plot analysis of the binding data indicated a heterogeneity of lipoteichoic acid-binding sites similar to that previously found for serum albumin. Nevertheless, fibronectin contains at least one population of high-affinity binding sites for lipoteichoic acid. The binding affinity (nKa approximately 250 muM(-1)) is 2 orders of magnitude greater than the binding affinity of serum albumin. These data suggest that human plasma fibronectin contains specific binding sites for fatty acids and that lipoteichoic acid binds to these sites by way of its glycolipid moiety.

Binding Sites↗
PubMed1983

Characterization of clinically significant strains of coagulase-negative staphylococci.

On occasion, a patient may have two or more clinical cultures yielding a coagulase-negative staphylococcus If these multiple isolates have the same phenotype, one might conclude that the same strain was reisolated from the patient, indicating its persistent and pathological presence. We examined the validity of this conclusion when we applied a number of characterizing systems to a collection of 143 isolates of coagulase-negative staphylococci collected during an outbreak of intravascular catheter-associated sepsis. The probability of classifying two random isolates as the same phenotype or species was as follows: P = 0.356 for phage typing, P = 0.348 for Baird-Parker biotyping, P = 0.346 for the API STAPH-IDENT (Analytab Products) system, P = 0.327 for Bentley et al. biotyping, and P = 0.077 for antimicrobial susceptibility patterns. Although antimicrobial susceptibility patterns had the lowest probability, a variability in test results of 7.7% and a tendency for strains to have similar antibiograms effectively raised the probability to P = 0.897. The combination of the API STAPH-IDENT with antibiograms resulted in a probability of P = 0.037 to P = 0.147. When all of the above methods were used together a probability of P = 0.014 was achieved. Five patients had isolates from two or more blood cultures spaced more than 1 day apart that were identical by all of the above criteria, thus confirming prolonged bacteremia. The collection was also examined for the incidence of slime production. Slime production was not associated with any of the above groups, but was associated with symptomatic infections (P less than 0.05) and gentamicin resistance (P less than 0.01). Slime production was strain stable and was of assistance in typing strains of coagulase-negative staphylococci.

Bacteriological Techniques↗
PubMed1982

Hybridoma antibodies against protective and nonprotective antigenic determinants of a structurally defined polypeptide fragment of streptococcal M protein.

Hybridoma technology was used to produce a set of monoclonal antibodies against a purified polypeptide fragment of type 24 streptococcal M protein to delineate the protective determinants of M protein exposed on the surface of the virulent streptococci. Several hybridoma antibodies were found to be opsonic against the homologous type streptococci. At least two of these antibodies (IIC3.7 and IIC4.6) protected mice against challenge infections with the homologous, but not a heterologous, serotype of bacteria. One of the hybridoma antibodies that reacted in high dilution (1:204,800) with the isolated M protein failed to react with the M protein on the surface of type 24 streptococci, and thus did not opsonize the homologous organisms or protect mice against challenge infections. Because hybridoma antibodies are directed against a single distinct immunodeterminant, these results indicate that protective immunity may be directed at any one of several distinct antigenic determinants of M protein exposed on the surface of virulent group A streptococci.

Animals↗
PubMed1982

The adherence of group A streptococci to oropharyngeal cells: the lipoteichoic acid adhesin and fibronectin receptor.

The attachment of group A streptococci to oropharyngeal epithelial cells is mediated by adhesive molecules (adhesins) on the surfaces of the micro-organisms that interact with receptor molecules on the epithelial cells. The evidence that the adhesin is composed of lipoteichoic acid (LTA) complexed with bacterial cell surface proteins is as follows: (a) Among the purified cell wall substances tested, only LTA was able to inhibit attachment; (b) treatment of streptococci with anti LTA but not with antibody against other surface substances blocks attachment; (c) LTa forms complexes with purified M protein, the most abundant protein on the surface of virulent streptococci; (d) the lipid moieties of LTA, which mediate attachment, remain free in the M protein-LTA complexes to interact with receptor analogues, such as serum albumin. The evidence that the receptor for the LTA mediated binding of streptococci resides in fibronectin molecules on oropharyngeal cells is as follows: (a) the addition ot adhesion test mixtures of fibronection inhibits binding; (b) the number of streptococci capable of attaching is directly proportional to the amount of fibronectin present on epithelial cells; (c) purified fibronectin immobilized on latex beads agglutinates suspensions of streptococci; (d) radiolabeled fibronectin binds to group A streptococci; (e) both the agglutination of fibronectin-beads and the binding of fibronectin to streptococci is blocked by LTA, the streptococcal adhesin.

Adhesiveness↗
PubMed1982

Adherence of slime-producing strains of Staphylococcus epidermidis to smooth surfaces.

Slime production is not a generally recognized feature of Staphylococcus epidermidis. In a recent outbreak of S. epidermidis intravascular catheter-associated sepsis, we noted that 63% of clinically implicated strains grew as a slimy film coating the culture tube walls when propagated in tryptic soy broth. Only 37% of randomly collected blood culture contaminants and skin isolates demonstrated a similar phenomenon (p less than 0.05). Transmission electron micrographs of these coating bacteria showed them to be encased in an extracellular matrix that stained with alcian blue. Slime production was most evident in autoclaved media containing Casamino Acids and glucose supplementation (0.25% wt/vol). There were strain and media preparation variability of slime production in the presence of other carbohydrates. Some strains were not able to produce slime under any of the tested conditions. The production or nonproduction of slime did not influence growth rate. When grown in vitro, slime producers accumulated on the surface of intravascular catheters as macrocolonies, whereas non-slime, producers did not. Transmission and scanning electron micrographs showed slime producers to be encased in an adhesive layer on the catheter surface, whereas nonproducers were not encased. These results suggest that slime-mediated adherence may be a critical factor in the pathogenesis of S. epidermidis infections of medical devices.

Adhesiveness↗
PubMed1982

Fibronectin-mediated binding of group A streptococci to human polymorphonuclear leukocytes.

Human plasma was shown to promote the attachment of an avirulent strain of Streptococcus pyogenes to human polymorphonuclear leukocytes. Removal of the fibronectin by absorption of the plasma with agarose-immobilized gelatin abolished the attachment-promoting activity. The activity of the absorbed plasma was restored with purified human serum fibronectin. The fibronectin was found to promote the attachment of three M protein-negative, but not of three M protein-positive, strains of streptococci. Radiolabeled fibronectin was shown to bind in various degrees to all strains of S. pyogenes tested. Thus, in the absence of streptococci surface M protein, fibronectin promotes the association of group A streptococci with human polymorphonuclear leukocytes.

Antigens, Bacterial↗
PubMed1982

Formation of molecular complexes between a structurally defined M protein and acylated or deacylated lipoteichoic acid of Streptococcus pyogenes.

The orientation of lipoteichoic acid (LTA) molecules on the surface of bacterial cells undoubtedly is determined by the ability of the LTA, during its transit through the cell wall, to bind via its polyglycerophosphate backbone or its glycolipid moieties to other constituents of the cytoplasmic membrane and the cell wall. We have investigated the possibility that LTA may become anchored to the cell surface by binding through its polyanionic backbone to positively charged regions of cell wall proteins. LTA was found to prevent the precipitation of partially purified HCl extracts of several strains of streptococci as well as a structurally defined streptococcal M protein molecule (pep M24) in 83% solutions of ethanol. The formation of complexes between LTA and M protein was demonstrated further by immunoelectrophoresis of pep M24 protein with increasing concentrations of radiolabeled LTA and by using antiserum against pep M24 to develop precipitin arcs. Pep M24 electrophoresed alone produced a single precipitin arc close to the origin. In contrast, when electrophoresed as a mixture with LTA or deacylated LTA, the M protein produced a second precipitin arc toward the anode coinciding with the area of migration of the radioactive LTA. Increasing concentrations of LTA or deacylated LTA shifted increasing amounts of the pep M24 antigen to the region of the second arc. Maleylation of M protein to block the positively charged free amino groups before mixing it with LTA prevented the formation of complexes. The complexes formed by the M protein with LTA, but not with deacylated LTA, showed the capacity to bind bovine serum albumin; LTA had been shown previously to bind to the fatty acid binding sites on bovine serum albumin. These results indicate that the LTA molecule is able to bind via its polyanionic backbone to positively charged residues of surface proteins of cells of S. pyogenes. The implications of such interaction as to the orientation of LTA molecules on the surface of cells of S. pyogenes are discussed.

Acylation↗
PubMed1982

Cytotoxicity of the glycolipid region of streptococcal lipoteichoic acid for cultures of human heart cells.

The ability of LTA of Streptococcus pyogenes to stimulate cell division or to kill tissue culture cells derived from human heart was investigated. Initial studies indicated that at low concentrations, ranging from 0.01 to 1.0 micrograms/ml, LTA stimulated cell division, whereas at higher concentrations, ranging from 10 to 1000 micrograms/ml, it killed the cells. Deacylated lipoteichoic acid, which lacked cell membrane binding activity, similarly stimulated or killed the heart cells depending on the concentration added to the tissue cultures. Fractionation of LTA after mild ammonia hydrolysis yielded a polyglycerophosphate and polar lipid fraction, both of which retained the glucose from the glycolipid moiety of the LTA molecule, and a neutral lipid fraction that was devoid of phosphorus or glucose. Toxic activity was present only in the fractions containing glucose. Oxidation of LTA or its deacylated derivative with sodium metaperiodate reduced the assayable glucose content without destroying the polyglycerophosphate backbone and resulted in a parallel loss of cytotoxicity, strongly suggesting that the glucose moieties of S. pyogenes LTA must be intact for the toxic activity against human heart cells to be expressed.

Acylation↗
PubMed1981

Depth perception in linear and inverse perspective pictures.

The range of pictorial depth perception was tested with four pictures from the repertoire of European art, rather than the customary line drawings or photographs. These pictures included those rendered in linear perspective and inverse perspective, as well as those with different degrees of depth. Using Pandora' Box, the subjects were asked to place a lamp at the same apparent depth as objects in the pictures. The subjects did so without regard to the depiction technique. The results suggest that depth is seen in pictures both where the rules of linear perspective hold and where they have been violated.

Art↗
PubMed1981

Characterization of lipoteichoic acid binding to polymorphonuclear leukocytes of human blood.

Human polymorphonuclear leukocytes (PMN) were shown to possess specific binding sites for lipoteichoic acid (LTA). LTA binding was reversible and time and temperature dependent. Scatchard plot analysis revealed an apparently single population of 6.6 X 10(6) LTA binding sites per PMN with a dissociation constant of 5.6 microM. Attachment of an avirulent, unencapsulated, M-negative strain of group A streptococci to PMN was inhibited by LTA, but not by other bacterial somatic antigens tested. Occupation of 30% of the LTA binding sites resulted in greater than 70% inhibition of streptococcal attachment to PMN. In contrast, LTA failed to block attachment of Escherichia coli or antibody-coated streptococci, indicating that binding sites for E. coli and the Fc portion of immunoglobulin G are distinct from those for LTA. Immunofluorescent studies demonstrated that LTA remained uniformly bound to PMN membranes for as long as 2 h at 37 degrees C. Cross-linking of PMN-bound LTA with anti-LTA resulted in rapid capping of LTA receptor sites. The results suggest that LTA is a monovalent ligand interacting with mobile receptors in the plasma membrane of PMN.

Binding Sites↗
PubMed1980

Binding of streptococcal lipoteichoic acid to the fatty acid binding sites on serum albumin.

The ability of the fatty acid binding sites of serum albumin to bind lipoteichoic acid of Streptococcus pyogenes was investigated. Initial studies indicated that lipoteichoic acid, but not its deacylated deprivative, protected albumin from being denatured by heat (80C for 1 h) and changed its mobility in an electrical field. Albumin covalently linked to agarose beads bound radiolabeled lipoteichoic acid, and the bound [3H]lipoteichoic acid could be specifically eluted with unlabeled lipoteichoic acid or albumin but not with other proteins tested. After binding to albumins, the lipoteichoic acid also could be quantitatively eluted with 50% ethanol and various detergents but not with up to 1.0 M sodium chloride. Binding of lipoteichoic acid to albumin followed first order kinetics, reaching saturation at 12 h. Analysis of the binding data by a Scatchard plot indicated heterogeneity of the binding sites on the albumin molecule similar to that previously reported for fatty acids. The affinity of binding of lipoteichoic acid to albumin was found to be intermediate between that previously reported for octanoic and palmitic acids, respectively. Based on these findings, we prepared affinity columns of immobilized albumin and were able to separate biologically active lipoteichoic acid from heterogeneous extracts of S. pyogenes.

Binding Sites↗
PubMed1980

Characteristics of the binding of streptococcal lipoteichoic acid to human oral epithelial cells.

Membrane receptors for lipoteichoic acid (LTA) may be involved in the adherence of streptococci to human pharyngeal epithelial cells. Since streptococcal binding to oral epithelial cells has been shown to be low at birth with a gradual increase to adult levels within three days, the characteristics of the binding of radiolabeled LTA to epithelial cells were investigated. LTA binding to epithelial cells from both infants and adults was time- and cell concentration-dependent. Binding was reversible in the presence of a 50-fold excess of unlabeled LTA. The number of LTA-binding sites in infant cells was only one-half that of adult cells; the amount of LTA bound increased to adult levels within three days of birth and paralleled an increase in the ability of these cells to bind streptococci. Thus, oral epithelial cells may possess specific binding sites for LTA, and a critical number or arrangement of LTA-binding sites may be required for the optimal binding of streptococci.

Adult↗