scholarly journals Staphylococcus epidermidis Has Growth Phase Dependent Affinity for Fibrinogen and Resulting Fibrin Clot Elasticity

2021 ◽  
Vol 12 ◽  
Author(s):  
Carolyn Vitale ◽  
Tianhui Maria Ma ◽  
Janice Sim ◽  
Christopher Altheim ◽  
Erika Martinez-Nieves ◽  
...  
Keyword(s):  
Bacterial Infection ◽  
Growth Phase ◽  
Staphylococcus Epidermidis ◽  
Fibrin Clot ◽  
Exponential Phase ◽  
Clot Formation ◽  
Coagulase Negative Staphylococci ◽  
Phase Dependence ◽  
The Impact

Bacterial infection and thrombosis are highly correlated, especially in patients with indwelling medical devices. Coagulase-negative staphylococci, typified by Staphylococcus epidermidis, are a common cause of medical device infections owing to their biofilm forming capacity which provides protection from antibiotics and host immune response. Attention has been drawn to the interaction between S. epidermidis and host proteins, specifically fibrinogen. However, little is known regarding the impact of the transition from planktonic to biofilm forming phenotype on this interaction. Here we investigate the growth phase dependence of bacteria-fibrinogen interaction and the resulting effect on fibrin clot formation, structure, and mechanics. Flow cytometry demonstrated growth phase dependent affinity for fibrinogen. To mimic intravascular device seeding, we quantified the adhesion of S. epidermidis to a fibrinogen coated surface under continuous flow conditions in vitro. The bacterial deposition rate onto fibrinogen was significantly greater for stationary (5,360 ± 1,776 cells/cm2s) versus exponential phase (2,212 ± 264, cells/cm2 s). Furthermore, the expression of sdrG–a cell surface adhesion protein with specificity for fibrinogen–was upregulated ∼twofold in the stationary versus the exponential phase. Rheometry and confocal microscopy demonstrated that stationary phase S. epidermidis slows clot formation and generates a more heterogeneous fibrin network structure with greater elasticity (G′ = 5.7 ± 1.0 Pa) compared to sterile fibrinogen (G′ = l.5 ± 0.2 Pa), while exponential phase cells had little effect. This work contributes to the current understanding of the growth phase dependent regulation of bacterial virulence factors and the correlation between bacterial infection and thrombosis.

Abstract 356: Adhesion of Staphylococcus Epidermidis to Fibrinogen Alters Clot Formation Kinetics and Ultimate Stiffness

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Carolyn Vitale ◽  
Tianhui Ma ◽  
Michael J Solomon ◽  
J. Scott VanEpps
Keyword(s):  
Stationary Phase ◽  
Deposition Rate ◽  
Staphylococcus Epidermidis ◽  
Biofilm Formation ◽  
Fibrin Clot ◽  
Exponential Phase ◽  
Automated Image Analysis ◽  
Clot Formation ◽  
Coagulation Proteins

Bacterial infection is known to increase the risk for thromboembolism. The mechanism underlying this correlation remains largely unknown. We recently showed that the common pathogen Staphylococcus epidermidis retards clot formation, increases clot elasticity and generates a heterogeneous clot structure that remodels over time. Here, we elucidate the mechanism of this process by evaluating the capacity for S. epidermidis to bind to fibrinogen as a function of its growth phase. We hypothesized that the effect of S. epidermidis on a fibrin clot is related to its propensity toward biofilm formation. Therefore, stationary phase (biofilm-like) S. epidermidis will have a more robust effect on clot kinetics and elasticity than exponential phase (planktonic). Furthermore, this difference is mediated by increased adhesion to fibrinogen. Rheometry was used to evaluate the formation and resultant elasticity of fibrin clots with exponential or stationary phase S. epidermidis . A functional in vitro model was developed to evaluate adhesion of S. epidermidis to a fibrinogen coated surface in a continuously flowing environment. Fluorescent labeled exponential and stationary phase S. epidermidis were visualized flowing through a parallel plate microfluidic chamber past immobilized fibrinogen. Images were obtained every 3 seconds for 30 min. Bacterial deposition rate and mean adhesion time were quantified by automated image analysis. A paired Student’s t-test was used for statistical analysis. Stationary phase S. epidermidis retards clot formation and increases resultant elasticity while exponential phase only slightly reduces elasticity. The bacterial deposition rate onto fibrinogen was significantly (p=0.03) greater for stationary phase (1741 ± 1513 cells/cm 2 · sec -1 ) vs exponential phase (676 ± 270 cells/cm 2 · sec -1 ). The average adhesion time however was similar for exponential and stationary phase cells. Coagulation proteins can provide a framework for bacterial adhesion, biofilm formation and infection. In turn infected thrombi with (biofilm-like) bacteria are stiffer which correlates to more frequent bacterial binding to fibrinogen. This provides a potential molecular mechanism for infection mediated thromboembolic events.

scholarly journals Impact of fibrinogen carbamylation on fibrin clot formation and stability

2017 ◽  
Vol 117 (05) ◽  
pp. 899-910 ◽  
Author(s):  
Stéphane Jaisson ◽  
Philippe Gillery ◽  
Carsten Scavenius ◽  
Endy Spriet ◽  
Anne Nyhaug ◽  
...  
Keyword(s):  
Kidney Disease ◽  
Mechanical Stability ◽  
Inflammatory Diseases ◽  
Plasma Concentrations ◽  
Fibrin Clot ◽  
Clot Formation ◽  
Cross Linking ◽  
Post Translational Modification ◽  
The Impact

SummaryCarbamylation is a non-enzymatic post-translational modification induced upon exposure of free amino groups to urea-derived cyanate leading to irreversible changes of protein charge, structure and function. Levels of carbamylated proteins increase significantly in chronic kidney disease and carbamylated albumin is considered as an important biomarker indicating mortality risk. High plasma concentrations and long half-life make fibrinogen a prime target for carbamylation. As aggregation and cross-linking of fibrin monomers rely on lysine residues, it is likely that carbamylation impacts fibrinogen processing. In this study we investigated carbamylation levels of fibrinogen from kidney disease patients as well as the impact of carbamylation on fibrinogen cleavage by thrombin, fibrin polymerisation and cross-linking in vitro. In conjunction, all these factors determine clot structure and stability and thus control biochemical and mechanical properties. LC-MS/MS analyses revealed significantly higher homocitrulline levels in patient fibrinogen than in fibrinogen isolated from control plasma. In our in vitro studies we found that although carbamylation does not affect thrombin cleavage per se, it alters fibrin polymerisation kinetics and impairs cross-linking and clot degradation. In addition, carbamylated fibrin clots had reduced fiber size and porosity associated with decreased mechanical stability. Using mass spectroscopy, we discovered that N-terminally carbamylated fibrinopeptide A was generated in this process and acted as a strong neutrophil chemoattractant potentially mediating recruitment of inflammatory cells to sites of fibrin(ogen) turnover. Taken together, carbamylation of fibrinogen seems to play a role in aberrant fibrin clot formation and might be involved in haemostatic disorders associated with chronic inflammatory diseases.

scholarly journals Neonatal Coinfection Model of Coagulase-Negative Staphylococcus (Staphylococcus epidermidis) and Candida albicans: Fluconazole Prophylaxis Enhances Survival and Growth

2007 ◽  
Vol 51 (4) ◽  
pp. 1240-1245 ◽  
Author(s):  
Mohan Pammi Venkatesh ◽  
Don Pham ◽  
Mindy Fein ◽  
Lingkun Kong ◽  
Leonard E. Weisman
Keyword(s):  
Candida Albicans ◽  
Weight Gain ◽  
Staphylococcus Epidermidis ◽  
Single Species ◽  
Coagulase Negative Staphylococcus ◽  
Coagulase Negative Staphylococci ◽  
Mortality And Morbidity ◽  
Fluconazole Prophylaxis ◽  
The Impact

ABSTRACT Coagulase-negative staphylococci (CoNS) and Candida are among the most common causes of single infections and coinfections in neonates after 72 h of age. In neonates, coinfection increases the rate of mortality threefold and results in significantly greater morbidity compared to those that result from single infections. In an effort to better understand this phenomenon, we developed the first neonatal animal model of coinfection (with CoNS and Candida) and evaluated its effects on mortality and morbidity and the impact of antifungal prophylaxis with fluconazole. Neonatal Wistar rats were infected with Candida albicans and/or Staphylococcus epidermidis with doses of 2 × 108 and 2 × 106 CFU subcutaneously in different combinations and were monitored for mortality, weight gain, and bacteremia. The in vitro sensitivity of C. albicans to fluconazole was evaluated and the MIC was determined. A subset of rats in these experiments received fluconazole at 10 mg/kg of body weight/dose intraperitoneally starting 24 h before infection for 4 days, and the serum trough levels of fluconazole were measured. Coinfection in the suckling rat significantly increased the rate of mortality compared to that after infection with a single species (P < 0.001) and resulted in deaths even at sublethal doses. Coinfection also impaired weight gain significantly in severely infected pups compared to that achieved after infection with a single species (P < 0.001). Fluconazole prophylaxis significantly reduced mortality by 30% in the Candida group and 36% in the coinfection group and improved weight gain in this neonatal model of coinfection (P < 0.001). We developed a neonatal model of coinfection with Candida and CoNS, observed significantly greater mortality and morbidity with coinfection, and found that fluconazole prophylaxis significantly reduced the rates of both mortality and morbidity. Further research on neonatal coinfection is urgently needed to improve clinical outcomes.

scholarly journals Repeated social defeat exaggerates fibrin-rich clot formation in FeCl3-induced arterial thrombosis mouse model by enhancing NETs formation via modulation of neutrophil functional properties

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
T Sugimoto ◽  
H Yamada ◽  
H Kubota ◽  
D Miyawaki ◽  
M Saburi ◽  
...  
Keyword(s):  
Functional Properties ◽  
Arterial Thrombosis ◽  
Social Defeat ◽  
Arterial Injury ◽  
Physical Contact ◽  
Clot Formation ◽  
Positive Staining ◽  
Double Positive ◽  
The Impact

Abstract Background and objective Depression is an independent risk factor of cardiovascular disease (CVD). We have recently shown that repeated social defeat (RSD) precipitates depressive-like behaviors in apoE−/− mice and exaggerates atherosclerosis development by enhancing neutrophil extracellular traps (NETs) formation. Here, we investigated the impact of RSD on arterial thrombosis. Methods and results Eight-week-old male WT mice were exposed to RSD by housing with a larger CD-1 mouse in a shared home cage. They were subjected to vigorous physical contact daily for 10 consecutive days. Control mice were housed in the same gage without physical contact. After social interaction test to confirm depressive-like behaviors, defeated mice (19 of 31) and control mice (12 of 14) were underwent arterial injury at 10 wks of age. A filter paper saturated with 10% FeCl3 was applied on the adventitial surface of left carotid artery for 3 min and analyzed 3 hrs later. The volume of thrombi was comparable between the two groups. However, fibrinogen/fibrin-positive areas in immunofluorescent images significantly increased in defeated mice (27.8% vs. 48.8%, p&lt;0.01). The number of Ly-6G-positive cells in thrombi was markedly higher in defeated mice (144/mm2 vs. 878/mm2, p&lt;0.05). Further, Ly-6G-positive cells were almost accumulated at the inner surface of injured artery, which were co-localized with neutrophil elastase, Cit-H3, and CD41-positive staining. Treatment with DNase I completely diminished the exaggerated fibrin-rich clot formation in defeated mice to an extent similar to that in control mice (25.7% vs. 22.3%, p = ns), without affecting the volume of thrombi and accumulation of Ly-6G-positive cells. Given that platelet aggregations induced by ADP or collagen were comparable between the two groups, neutrophil functional properties primarily contribute to the exaggerated fibrin-rich clot formation in defeated mice. We then examined neutrophil subset and vulnerability to NETs formation. At 3 hrs after FeCl3 application, the numbers of immature neutrophils (Ly6Glo/+CXCR2-) were comparable between the two groups in both bone marrow (BM) and peripheral blood (PB). In contrast, the number of PB mature neutrophils (Ly6G+CXCR2+) was markedly higher in defeated mice than control mice (580±68 /μl vs. 1265±114, p&lt;0.01). We next examined in vitro NETs formation upon PMA in BM mature neutrophils by FACS and nucleic acid staining. The percentage of double-positive cells (Cit-H3, MPO) was significantly higher in defeated mice (7.5% vs. 10.2%, p&lt;0.05), as well as SYTOX green-positive cells expelling DNA fibers (8.1% vs. 11.8%, p&lt;0.05). Conclusions Our findings demonstrate for the first time that repeated social defeat enhances fibrin-rich clot formation after arterial injury by enhancing NETs formation via modulation of neutrophil functional properties, suggesting that NETosis could be a new therapeutic target in depression-related CVD development. Funding Acknowledgement Type of funding source: None

scholarly journals Impact of Bacterial Biofilm Formation on In Vitro and In Vivo Activities of Antibiotics

1998 ◽  
Vol 42 (4) ◽  
pp. 895-898 ◽  
Author(s):  
Silvia Schwank ◽  
Zarko Rajacic ◽  
Werner Zimmerli ◽  
Jürg Blaser
Keyword(s):  
Animal Model ◽  
Staphylococcus Epidermidis ◽  
Biofilm Formation ◽  
Bacterial Biofilm ◽  
Phenotypic Resistance ◽  
Vitro Model ◽  
The Impact ◽  
Isogenic Strains

ABSTRACT The impact of bacterial adherence on antibiotic activity was analyzed with two isogenic strains of Staphylococcus epidermidis that differ in the features of their in vitro biofilm formation. The eradication of bacteria adhering to glass beads by amikacin, levofloxacin, rifampin, or teicoplanin was studied in an animal model and in a pharmacokinetically matched in vitro model. The features of S. epidermidis RP62A that allowed it to grow on surfaces in multiple layers promoted phenotypic resistance to antibiotic treatment, whereas strain M7 failed to accumulate, despite initial adherence on surfaces and growth in suspension similar to those for RP62A. Biofilms of S. epidermidis M7 were better eradicated than those of strain RP62A in vitro (46 versus 31%;P < 0.05) as well as in the animal model (39 versus 9%; P < 0.01).

scholarly journals Molecular Epidemiology of a Vancomycin-Intermediate Heteroresistant Staphylococcus epidermidis Outbreak in a Neonatal Intensive Care Unit

2016 ◽  
Vol 60 (10) ◽  
pp. 5673-5681 ◽  
Author(s):  
Jasmine Chong ◽  
Caroline Quach ◽  
Ana C. Blanchard ◽  
Philippe Guillaume Poliquin ◽  
George R. Golding ◽  
...  
Keyword(s):  
Intensive Care ◽  
Molecular Epidemiology ◽  
Staphylococcus Epidermidis ◽  
Population Analysis ◽  
Tertiary Care ◽  
Hospital Environment ◽  
Coagulase Negative Staphylococci ◽  
Time Curve ◽  
Content Type ◽  
The Impact

ABSTRACTCoagulase-negative staphylococci (CoNS) have become the leading cause of bloodstream infections (BSIs) in intensive care units (ICUs), particularly in premature neonates. Vancomycin-intermediate heteroresistant CoNS (hVICoNS) have been identified as sources of BSIs worldwide, and their potential to emerge as significant pathogens in the neonatal ICU (NICU) remains uncertain. This study describes the molecular epidemiology of an outbreak of vancomycin-heteroresistant (hV)Staphylococcus epidermidiscentral-line-associated BSI (CLABSI) in a single tertiary care NICU and compares it to a second tertiary care NICU that had not been associated with an outbreak. Between November 2009 and April 2014, 119S. epidermidisCLABSIs were identified in two tertiary care NICUs in Quebec, Canada. Decreased vancomycin susceptibility was identified in about 88% of all collected strains using Etest methods. However, discrepancies were found according to the Etest and population analysis profiling–area under the concentration-time curve (PAP-AUC) methods used. All strains were susceptible to linezolid, and a few isolates were nonsusceptible to daptomycin. Great genetic diversity was observed within the collection, with 31 pulsed-field gel electrophoresis (PFGE) patterns identified. The outbreak strains were all determined to be heteroresistant to vancomycin and were polyclonal. The study identified two major clones, PFGE patterns E and G, which were found in both NICUs across the 5-year study period. This suggests the persistence of highly successful clones that are well adapted to the hospital environment. hVS. epidermidisseems more common than currently realized in the NICU, and certain hVS. epidermidisclones can become endemic to the NICU. The reservoirs for these clones remain unknown at this time, and identification of the reservoirs is needed to better understand the impact of hVS. epidermidisin the NICU and to inform infection prevention strategies. In addition, there is a need to investigate and validate hV determination protocols for different species of CoNS.

scholarly journals Lipopolysaccharide administration alters extracellular vesicles in human lung-cancer cells and mice

2020 ◽  
Author(s):  
Leandra B. Jones ◽  
Sanjay Kumar ◽  
Courtnee’ R. Bell ◽  
Brennetta J. Crenshaw ◽  
Mamie T. Coats ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Bacterial Infection ◽  
Cell Line ◽  
Extracellular Vesicles ◽  
A549 Cells ◽  
Human Lung Cancer ◽  
Diagnostic Tools ◽  
The Impact

AbstractExtracellular vesicles (EVs) play a fundamental role in cell and infection biology and have the potential to act as biomarkers for novel diagnostic tools. In this study, we explored the in vitro impact of bacterial lipopolysaccharide administration on a cell line that represents a target for bacterial infection in the host. Administration of lipopolysaccharide at varying concentrations to this A549 cell line caused only modest changes in cell death, but EV numbers were significantly changed. After treatment with the highest concentration of lipopolysaccharide, EVs derived from A549 cells packaged significantly less interleukin-6 and lysosomal-associated membrane protein 1. We also examined the impact of lipopolysaccharide administration on exosome biogenesis and cargo composition in BALB/c mice. Serum-isolated EVs from lipopolysaccharide-treated mice showed significantly increased lysosomal-associated membrane protein 1 and toll-like receptor 4 levels compared with EVs from control mice. In summary, this study demonstrated that EV numbers and cargo were altered using these in vitro and in vivo models of bacterial infection.

Development of MDL 28,050, a Small Stable Antithrombin Agent Based on a Functional Domain of the Leech Protein, Hirudin

1990 ◽  
Vol 63 (02) ◽  
pp. 208-214 ◽  
Author(s):  
John L Krstenansky ◽  
Robert J Broersma ◽  
Thomas J Owen ◽  
Marguerite H Payne ◽  
Mark T Yates ◽  
...  
Keyword(s):  
Structural Optimization ◽  
Systematic Study ◽  
Fibrin Clot ◽  
Clot Formation ◽  
Functional Domain ◽  
In Vivo Models ◽  
Agent Based ◽  
Structure Activity

SummaryMDL 28,050 is a decapeptide antithrombin agent that inhibits a-thrombin-induced fibrin clot formation by binding to a non-catalytic site on α-thromhin. It is the result of chemical and structural optimization of a functional domain of the leech anticoagulant, hirudin. In contrast to the contention that the polyanionic nature of this C-terminal functional domain governs its interaction with α-thrombin, systematic study of this region has shown the importance of the lipophilic residues for providing the functionality necessary foi potent binding to a-thrombin. The development of MDL 28,050 and other effective antithrombin agents are outlined through the description of the structure-activity relationships (SAR) for these peptides. These peptides are effective in a variety of in vitro and in vivo models of thrombosis.

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