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Biomedical subjects

R A Proctor

Publications and source records attributed to R A Proctor.

At least 91 records · Page 5Linked to original sources
PubMed1985

Genetic differences in oxygen toxicity are correlated with cytochrome P-450 inducibility.

Susceptibility to oxygen toxicity was studied in three inbred and two hybrid strains of mice. Because in vitro studies have shown that the cytochrome P-450 enzymes can produce oxygen radicals and H2O2, we tested the hypothesis that inducibility of these enzymes might play a role in oxygen toxicity. Mice responsive to hepatic microsomal enzyme induction by aromatic hydrocarbons [C3H/HeJ, C3H/HeN, C3H/HeJ X DBA/2J (designated C3D2F1/J), C3H/HeN X DBA/2J (designated C3D2F1/N)] were more sensitive to the toxic effects of 100% oxygen exposure than were genetically unresponsive mice (DBA/2J). DBA/2J mice survived significantly longer exposure periods with less lung damage. Lung and liver cytochrome P-450 levels increased 2-to 3-fold in C3H and F1 mice during 100% oxygen exposure (maximum levels at 72-96 hr) and subsequently fell prior to death. No increases were seen in cytochrome P-450 levels in DBA/2J mice. Metabolic pathways involving cytochrome P-450 enzymes may initiate or modulate oxidative damage due to oxygen radicals. The difference in responsiveness of mice to microsomal enzyme induction may imply genetic differences in susceptibility to oxidative stress, may help to explain species differences in susceptibility, and may have long-term implications in therapeutics and patient care if similar inherited differences exist in humans.

Animals↗
PubMed1985

Cilastatin does not alter superoxide dismutase activity.

Cilastatin inhibits dehydropeptidase-I, a zinc metaloenzyme that metabolizes imipenem. Because zinc stabilizes the mammalian superoxide dismutase, we postulated that cilastatin would also inhibit the dismutase. Cilastatin concentrations at levels threefold higher than those expected in urine, however, did not inhibit the superoxide dismutase activity.

Cilastatin↗
PubMed1985

Role of fibronectin in human monocyte and macrophage bactericidal activity.

Fibronectin is a high-molecular-weight glycoprotein found as a soluble dimer in plasma and as an insoluble multimer in tissues. It has been proposed that plasma fibronectin facilitates phagocytic removal of lysed cells and damaged tissues. Fibronectin binds avidly to several species of gram-positive bacteria and enhances staphylococcal and streptococcal attachment to cultured cells. Determination of whether fibronectin will enhance the bactericidal activity of monocytes and macrophages has not been reported. The bactericidal activity of freshly isolated monocytes, cultured monocytes, or lymphokine-activated macrophages was tested in the presence of either dimeric or multimeric fibronectin. Freshly isolated monocytes and lymphokine-activated macrophages killed Staphylococcus aureus effectively in the absence of fibronectin or whole serum. In contrast, monocytes cultured for 7 to 10 days had diminished staphylocidal capacity. When the monocytes were cultured with either dimeric or multimeric fibronectin, however, bactericidal capacity was maintained. Thus, although fibronectin did not enhance the bactericidal activity of mononuclear phagocytes, both multimeric and dimeric fibronectin were effective at maintaining the bactericidal capacity.

Blood Bactericidal Activity↗
PubMed1985

Activation and inhibition of Limulus amebocyte lysate coagulation by chemically defined substructures of lipid A.

Recent work with lipid mutants of Escherichia coli and Salmonella typhimurium has helped to elucidate the correct structure of lipid A and has suggested a biosynthetic pathway. Precursor molecules include diacylglucosamine 1-phosphates and tetraacyl disaccharide bis-phosphates. The activities of several of these compounds and of their derivatives were measured by Limulus amebocyte lysate (LAL) assay. We report that (i) both mono- and disaccharide precursors of lipid A activate LAL, (ii) two acyl chains on the monosaccharide subunit of lipid A are necessary for activation of LAL, and (iii) the monosaccharide, 2-monoacylglucosamine 1-phosphate can competitively inhibit LAL activation by diacyl monosaccharide lipid A precursors. However, 2-monoacylglucosamine 1-phosphate did not inhibit endotoxin activation of LAL. One unanticipated finding was that the activities of the monosaccharides were reduced upon storage even though their covalent structures were unchanged. Perhaps this is due to alterations in physical state. Thus, these lipid A precursors and derivatives offer some insight into the structural features required for activation of the LAL assay and may in the future provide derivatives which are competitive inhibitors of endotoxin.

Drug Stability↗
PubMed1985

Pulmonary pathophysiological changes in sheep caused by endotoxin precursor, lipid X.

The chemical structure of the biologically active lipid A portion of Gram-negative endotoxin [lipopolysaccharide (LPS)] has recently been elucidated. This was greatly facilitated by the isolation of an Escherichia coli mutant that accumulates large quantities of lipid X, a novel monosaccharide precursor of lipid A (C. R. H. Raetz, Rev. Infect. Dis. 6: 463-471, 1984). We now report on the activity of lipid X in the lung-lymph model in sheep. We have measured the response to cumulative bolus injections of lipid X (2,3-diacylglucosamine 1-phosphate) in six chronically instrumented unanesthetized sheep. Lipid X at a total dose of 40 micrograms/kg produced a biphasic pattern of changes. The early phase was characterized by a rapid transient pulmonary arterial constrictive response that was dose dependent, accompanied by a delayed transient increase in lung-lymph flow (P less than 0.05), a significant (P less than 0.01) decrease in arterial blood O2 tension and an increase (P less than 0.05) in lung-lymph protein clearance. Protein permeability changes in the first phase are not usually seen following endotoxin injection. However, like endotoxin, lipid X also produced a late phase (3-6 h later) of increased lung vascular permeability to fluid and protein as reflected by significant (P less than 0.05) increases in both lung-lymph flow and lung-lymph protein clearance in the presence of stable pulmonary vascular pressures at or below base-line levels. We conclude that some of the pulmonary pressor activity of the endotoxin molecule can be attributed to the lipid X substructure. Furthermore, changes in vascular permeability may also be initiated by this substance.

Animals↗
PubMed1984

Effect of chelating agents and superoxide on human neutrophil NAD(P)H oxidation.

NAD(P)H oxidation is frequently measured to assay the activity of the neutrophil O-2-generating oxidase. It was found that 10(-4) M ethylene glycol bis (beta-aminoethyl ether)-N-N'-tetraacetic acid (EGTA) increased NAD(P)H oxidation by the 27,000 g granule fraction of resting and stimulated human neutrophils without altering net O-2 production. The commonly used chelating agents EDTA and diethylene triamine pentaacetic acid had similar effects. The addition of superoxide dismutase eliminated the effect of the chelating agents and thus demonstrated that the stimulated reaction was dependent upon O-2. KCN and bathophenanthroline disulfonate, an iron-chelating agent, prevented O-2-dependent NADPH oxidation by neutrophil granule fractions in the presence of EGTA. In contrast, bathocuproine disulfonate, a copper-chelating agent, mimicked the EGTA effect. The effects of both bathophenanthroline disulfonate and bathocuproine disulfonate were completely abolished when the agents were saturated with iron and copper, respectively. All the chelating agents studied, except bathophenonthroline disulfonate, also promoted O-2-dependent NADPH oxidation in a system wherein O-2 was generated by xanthine oxidase. Thus, commonly used chelating agents, by interacting with available iron and copper, may alter the apparent stoichiometry of the neutrophil O-2-generating oxidase and artifactually increase NADPH oxidation in other systems where O-2 is present.

Cations, Divalent↗
PubMed1984

Inefficient in vitro killing of virulent or nonvirulent Salmonella typhimurium by murine polymorphonuclear neutrophils.

The bactericidal capability of murine peritoneal polymorphonuclear neutrophils against virulent and nonvirulent Salmonella typhimurium was examined in an in vitro system. Although preincubation of the bacteria in specific murine antiserum elicited greater chemiluminescence from phagocytizing neutrophils than did incubation in normal murine serum, antiserum did not enhance ingestion, as less than 5% of the challenge was taken up by neutrophils under any of the conditions studied. Nonvirulent salmonellae showed a transient decrease in viable numbers early during in vitro incubation with or without intact neutrophils. Virulent salmonellae, however, were able to multiply without a lag period except when these bacteria were pretreated with antiserum and incubated in association with intact murine neutrophils. Results of these in vitro studies suggest that the murine polymorphonuclear neutrophil and antisalmonella antibody must act together to effect neutrophil-associated bactericidal activity against virulent salmonellae, and thus, that the neutrophil alone does not play a major role in the protection of unvaccinated, sensitive mice from disease caused by S. typhimurium.

Animals↗
PubMed1984

Host defences and the vaginal mucosa. A re-evaluation.

The portal of entry for most pathogens is at the mucosal surface, and mucosal defences afford the host the earliest opportunity to ward off infection. The vagina is exposed to considerable trauma, and harbours a complex microflora, but only a small number of microorganisms are associated with disease. It seems likely that mucosal defenses are important in this process. Mucosal secretions contain mucus, lysozyme, lactoferrin, zinc, fibronectin, and complement, all of which afford non-specific protection. In addition, secretory immunoglobulin A (IgA) concentration in these secretions increases in response to specific pathogens. Blood leukocytes migrate onto mucosal surfaces during infection with some microorganisms, and the interaction between phagocytic cells and genital pathogens has been the focus of intense investigation. Improved understanding of mucosal defences is crucial for prevention of infection.

Aging↗
PubMed1984

Fibronectin-induced agglutination of Staphylococcus aureus correlates with invasiveness.

Studies on the interactions of staphylococci with fibrinogen, fibrin split products, and prothrombin have formed the basis for the clumping tests for coagulase and fibrin degradation products. We investigated the role of another circulating protein, fibronectin, in clumping Staphylococcus aureus. Fibronectin is a dimeric glycoprotein with high molecular weight that is present in both blood and tissue and is involved in opsonization, clotting, healing of wounds, cell-to-cell attachment, and differentiation. Each fibronectin molecule has two S. aureus binding sites, thus allowing lattice formation. We defined conditions under which fibronectin will cause agglutination of S. aureus. Strains of S. aureus that were most easily clumped had the largest number of fibronectin receptors. Trypsinization or gentle sonication removed the fibronectin binding and agglutinating receptors from S. aureus. These treatments did not alter viability, which suggests that binding is a superficial component of the organisms. Invasive fibronectin-binding strains were from a wide variety of phage types. Twenty two S. aureus isolates from patients with invasive disease were more readily agglutinated and had a greater number of fibronectin binding sites than 19 noninvasive strains (p less than 2.5 X 10(-4)). This suggests that the pathogenicity of S. aureus invasion may be enhanced by binding of bacteria to tissue fibronectin or by agglutination of bacteria by plasma fibronectin. Thus, the fibronectin receptors on S. aureus that mediate agglutination might also permit invasion of host tissues.

Fibrinogen↗
PubMed1983

Effect of Staphylococcus aureus enterotoxin F on human neutrophil oxidative metabolism.

Epidemiologic and animal challenge studies have suggested that SEF may play a critical role in toxic-shock syndrome [1]. However, the pathogenic mechanism of SEF activity is not known. One means by which this toxin could elicit the widespread organ dysfunction observed in toxic-shock syndrome would be by directly promoting PMN production of toxic oxygen species and the resultant secondary endothelial cell damage [2]. To evaluate this possibility, we assayed the effect of SEF on PMN oxidative metabolism. SEF at 0.01-100 ng/ml did not stimulate O2- release or O2 consumption by inactive PMNs. Similarly, incubation of PMNs with 10 ng of SEF/ml for 1 hr neither potentiated nor inhibited cellular O2 consumption stimulated by optimal (10 mg/ml) or suboptimal (0.1 mg/ml) concentrations of opsonized zymosan. Finally, SEF had no effect on O2- release by PMNs stimulated by PMA. PMN viability, as assessed by trypan blue exclusion, was unaffected by SEF. This study did not address the possibility that SEF might indirectly activate PMN oxidative metabolism by promoting leukocytic pyrogen production by monocytes and macrophages [3]. SEF neither directly activated PMN oxidative activity nor potentiated the cellular oxidative response to particulate or soluble stimuli. Consequently, direct stimulation of PMN-derived, O2- mediated damage to endothelial cells is not a tenable hypothesis to explain the mechanism of SEF toxicity.

Bacterial Toxins↗
PubMed1983

Effects of subinhibitory concentrations of antibiotics on Staphylococcus aureus interactions with fibronectin.

Bacterial adherence to host tissues relies on interactions between tissue macromolecules and bacterial surface molecules. One of the major predisposing factors to infection with Staphylococcus aureus is trauma to tissues. A common element in traumatized tissues is fibronectin. In previous studies, we have shown that fibronectin binds to Staph. aureus. In this paper, we have investigated the effects of subinhibitory concentrations of antibiotics on fibronectin interactions with Staph. aureus. Exposure of Staph. aureus to 1/4 MIC of penicillin increases the number of binding sites and enhances adherence of Staph. aureus to a collagen-fibronectin matrix. Chloramphenicol, erythromycin, clindamycin, and U57,930E all decreased the number of binding sites. Also, U57,930E reduced Staph. aureus adherence to a collagen-fibronectin matrix. Taken together, these data suggest that penicillin may enhance Staph. aureus adherence to tissue fibronectin whereas U57,930E might reduce such binding.

Anti-Bacterial Agents↗
PubMed1983

Subinhibitory concentrations of antibiotics alter fibronectin binding to Staphylococcus aureus.

Fibronectin, a high-molecular-weight glycoprotein, is found in plasma and on mammalian cell surfaces. Recent reports have suggested that bacterial-fibronectin interactions play a role in bacterial attachment to host cells. Subinhibitory concentrations of lincosamines, erythromycin, and chloramphenicol decreased fibronectin binding to Staphylococcus aureus, whereas beta-lactam antibiotics enhanced this interaction.

Anti-Bacterial Agents↗
PubMed1982

Fibronectin binding to Staphylococcus aureus.

Bacteria are able to interact with a number of macromolecules which act as opsonins or tissue-adherence factors. Because soluble fibronectin may be important factors. Because soluble fibronectin may be important in the phagocytic removal of bacteria and insoluble fibronectin may serve as a bridge between bacteria and host tissues, we have characterized the binding of soluble plasma fibronectin to Staphylococcus aureus as a first step to understanding these interactions. The binding of 125I-fibronectin to clinical and laboratory strains of S. aureus was studied. Bound fibronectin was separated from free fibronectin by centrifugation. Specific binding was determined by subtracting the amount bound in the presence of excess fibronectin from the total amount bound. We found that (i) fibronectin bound saturably, irreversibly, and noncovalently to S. aureus when the binding reaction was carried out at pH 7.4 or greater; (ii) S. aureus harvested in logarithmic phase of growth from media buffered to pH 8.4, and from brain-heart infusion media which demonstrated the greatest number of fibronectin-binding sites; (iii) high molecular weight dextrans, fibrinogen, cyanogen bromide fragment 7 of collagen, cationic proteins, dibromide fragment 7 of collagen, cationic proteins, dithiothreitol, and protein A did not alt er fibronectin binding to S. aureus; (iv) nonsaturable binding occurred below pH 7.0 with peak binding occurring at pH 5.8; and (v) there were marked differences in the amounts of fibronectin that bind to different strains of S. aureus. S. aureus ATCC 25923, when harvested in logarithmic phase of growth from tryptic soy broth and tested for fibronectin binding in (2-hydroxyethyl)-1-piperazineethanesulfonic acid-buffered saline, pH 7.4, had 7500 binding sites/organism with an apparent association constant of 5.6 X 10(9) M-1.

Fibronectins↗
PubMed1982

Elicitation of peritoneal polymorphonuclear neutrophils from mice.

Although the mouse has been used extensively as a model for the study of host-parasite relationships, murine neutrophils have not been used nearly as often as PMNs from other species for in vitro functional assays due to lack of a commonly used procedure for murine neutrophil collection. These studies compared two eliciting agents and characterized the phagocytic and bactericidal activity of murine polymorphonuclear neutrophils elicited from the peritoneal cavity. We examined the effects of mouse strain (BALB/c, C57BL/6 and DBA/2) and sex, eliciting agent (0.2% glycogen vs. 3% fluid thioglycolate medium) and donor sacrifice method (ether vs. cervical dislocation) on the number of neutrophils recovered in peritoneal exudate. The greatest number of neutrophils was harvested when mice were sacrificed 5 h after intraperitoneal injection of 2.5 ml of 3% thioglycolate medium. This method as described allows reproducible collection of adequate numbers of neutrophils for use in in vitro assays of neutrophils phagocytic and bactericidal function.

Animals↗
PubMed1980

Binding and factor XIIIa-mediated cross-linking of a 27-kilodalton fragment of fibronectin to Staphylococcus aureus.

A 27-kilodalton tryptic fragment, derived from the amino terminus of the 200-kilodalton fibronectin subunit, inhibited binding of intact fibronectin to Staphylococcus aureus and could be cross-linked to Staphylococcus aureus by blood coagulation Factor XIIIa. Interactions of fibronectin with Staphylococcus aureus via this fragment may be important for bacterial opsonization and attachment.

Amino Acid Sequence↗
PubMed1979

Endotoxin in vitro interactions with human neutrophils: depression of chemiluminescence, oxygen consumption, superoxide production, and killing.

Endotoxin was shown to depress neutrophil bactericidal activity while enhancing Nitro Blue Tetrazolium reduction and hexose monophosphate shunt activity. Separation of bactericidal action from oxidative metabolism suggests that the effect of endotoxin might involve the formation of reactive oxygen radicals such as superoxide. Chemiluminescence often accompanies metabolic activation of polymorphonuclear neutrophils (PMNs). However, human PMNs did not show chemiluminescence when challenged with endotoxin (lipopolysaccharide; LPS) or lipid A. Superoxide formation was also unaffected by endotoxin. In contrast, preincubation of PMNs with LPS for 30 min produced significant depression of chemiluminescence, oxygen consumption, and superoxide formation. Decreased chemiluminescence was not the result of complement consumption. In a cell-free system, superoxide was not scavenged by LPS, nor did LPS stimulate superoxide dismutase. Oxidase enzymes for reduced nicotinamide adenine dinucleotide or reduced nicotinamide adenine dinucleotide phosphate harvested from broken cells were not affected by LPS. The toxicity of LPS may reside in its ability to activate the PMNs while simultaneously blocking bactericidal capacity.

Blood Bactericidal Activity↗