Osteoprotegerin independently predicts mortality in patients with stable coronary artery disease: the CLARICOR trial.

OBJECTIVES: To elucidate the prognostic power of serum osteoprotegerin (OPG) in patients with stable coronary artery disease (CAD). METHODS: Serum OPG levels were measured in the CLARICOR trial cohort of 4063 patients with stable CAD on blood samples drawn at randomization. The follow-up was 2.6 years for detailed cardiovascular events and 6 years for all-cause mortality. RESULTS: OPG levels were significantly increased in non-survivors (21%) compared to survivors (median [quartiles] 2092 ng/L [1636; 2800] compared to 1695 ng/L [1322; 2193, p < 0.0001]). The 2.6-year follow-up showed that OPG adds to the prediction of both cardiovascular and all-cause mortality in combination with clinical risk factors (HR [one log10 unit increase] 6.1 [95% CI 2.4-15.6, p = 0.0001]) and HR 6.5 [95% CI 3.4-12.5, p < 0.0001], respectively). Similar, in the 6-year follow-up, OPG was found to be a strong predictor for all-cause mortality. Importantly, OPG remained an independent predictor of mortality even after adjustment for both clinical and conventional cardiovascular risk markers (HR 2.5 [95% CI 1.6-3.9, p < 0.0001]). CONCLUSIONS: Serum OPG has a long-lasting independent predictive power as to all-cause mortality and cardiovascular death in patients with stable CAD.

Escherichia coli uropathogenesis in vitro: invasion, cellular escape, and secondary infection analyzed in a human bladder cell infection model.

Uropathogenic Escherichia coli (UPEC) strains are capable of invading bladder epithelial cells (BECs) on the bladder luminal surface. Based primarily on studies in mouse models, invasion is proposed to trigger an intracellular uropathogenic cascade involving intracellular bacterial proliferation followed by escape of elongated, filamentous bacteria from colonized BECs. UPEC filaments on the mouse bladder epithelium are able to revert to rod-shaped bacteria, which are believed to invade neighboring cells to initiate new rounds of intracellular colonization. So far, however, these late-stage infection events have not been replicated in vitro. We have established an in vitro model of human bladder cell infection by the use of a flow chamber (FC)-based culture system, which allows investigation of steps subsequent to initial invasion. Short-term bacterial colonization on the FC-BEC layer led to intracellular colonization. Exposing invaded BECs to a flow of urine, i.e., establishing conditions similar to those faced by UPEC reemerging on the bladder luminal surface, led to outgrowth of filamentous bacteria similar to what has been reported to occur in mice. These filaments were capable of reverting to rods that could invade other BECs. Hence, under growth conditions established to resemble those present in vivo, the elements of the proposed uropathogenic cascade were inducible in a human BEC model system. Here, we describe the model and show how these characteristics are reproduced in vitro.

High-sensitivity C-reactive protein and N-terminal pro-B-type natriuretic peptide in patients with stable coronary artery disease: a prognostic study within the CLARICOR trial.

BACKGROUND: Patients with stable coronary artery disease (CAD) have a poor prognosis. The aim of the study was to evaluate the extent to which serum high-sensitivity C-reactive protein (hs-CRP) and N-terminal pro-B-type natriuretic peptide (NT-proBNP) measurement alone or together could be prognostic biomarkers in patients with stable CAD. MATERIALS AND METHODS: During the 2.6-year follow-up period 270 patients among the 4264 patients with stable CAD in the CLARICOR trial suffered myocardial infarction (MI) and 377 died (187 cardiovascular deaths (CVD)). RESULTS: Serum NT-proBNP was significantly associated with MI (hazard ratio (HR), 1. 65 (refers to a 2.72 fold increase in serum level, p = 0.0005), CVD (HR, 2.42, p < 0.0005) and non-CVD (HR, 1.79, p < 0.0005). When correcting for hs-CRP, NT-proBNP was still significantly associated with MI (HR, 1.63, p = 0.0005), CVD (HR, 2.36, p < 0.0005) and non-CVD (HR, 1.66, p < 0.0005). Serum hs-CRP was compared to NT-proBNP less associated with MI (HR, 1.20, p = 0.001), CVD (HR, 1.39, p < 0.0005) and non-CVD (HR, 1.67, p < 0.0005). When corrected for NT-proBNP, hs-CRP was only associated with non-CVD (HR, 1.51, p < 0.0005). When adjusting for cardiovascular risk factors hs-CRP predicted non-CVD (HR, 1.46) and all-cause death (HR, 1.24) and NT-proBNP predicted MI (HR, 1.50), CVD (HR, 1.98), non-CVD (HR, 1.39), and all-cause death (HR, 1.62)(p < 0.0005 for all). CONCLUSION: Increased serum NT-proBNP was a stronger predictor of MI, cardiovascular death and non-cardiovascular death than hs-CRP in patients with stable CAD. Once NT-proBNP was taken into account, hs-CRP did not improve predictions.

Pig-associated methicillin-resistant Staphylococcus aureus: family transmission and severe pneumonia in a newborn.

Carriage of pig-associated methicillin-resistant Staphylococcus aureus (MRSA) is known to occur in pig farmers. Zoonotic lineages of MRSA have been considered of low virulence and with limited capacity for inter-human spread. We present a case of family transmission of pig-associated MRSA ST398, which resulted in a severe infection in a newborn.

A Method for Quantification of Epithelium Colonization Capacity by Pathogenic Bacteria.

Most bacterial infections initiate at the mucosal epithelium lining the gastrointestinal, respiratory, and urogenital tracts. At these sites, bacterial pathogens must adhere and increase in numbers to effectively breach the outer barrier and invade the host. If the bacterium succeeds in reaching the bloodstream, effective dissemination again requires that bacteria in the blood, reestablish contact to distant endothelium sites and form secondary site foci. The infectious potential of bacteria is therefore closely linked to their ability to adhere to, colonize, and invade epithelial and endothelial surfaces. Measurement of bacterial adhesion to epithelial cells is therefore standard procedure in studies of bacterial virulence. Traditionally, such measurements have been conducted with microtiter plate cell cultures to which bacteria are added, followed by washing procedures and final quantification of retained bacteria by agar plating. This approach is fast and straightforward, but yields only a rough estimate of the adhesive properties of the bacteria upon contact, and little information on the ability of the bacterium to colonize these surfaces under relevant physiological conditions. Here, we present a method in which epithelia/endothelia are simulated by flow chamber-grown human cell layers, and infection is induced by seeding of pathogenic bacteria on these surfaces under conditions that simulate the physiological microenvironment. Quantification of bacterial adhesion and colonization of the cell layers is then performed by in situ time-lapse fluorescence microscopy and automatic detection of bacterial surface coverage. The method is demonstrated in three different infection models, simulating Staphylococcus aureus endothelial infection and Escherichia coli intestinal- and uroepithelial infection. The approach yields valuable information on the fitness of the bacterium to successfully adhere to and colonize epithelial surfaces and can be used to evaluate the influence of specific virulence genes, growth conditions, and antimicrobial treatment on this process.

Co-release of dicloxacillin and thioridazine from catheter material containing an interpenetrating polymer network for inhibiting device-associated Staphylococcus aureus infection.

Approximately half of all nosocomial bloodstream infections are caused by bacterial colonization of vascular catheters. Attempts have been made to improve devices using anti-adhesive or antimicrobial coatings; however, it is often difficult to bind coatings stably to catheter materials, and the low amounts of drug in thin-film coatings limit effective long-term release. Interpenetrating polymer networks (IPNs) are polymer hybrid materials with unique drug release properties. While IPNs have been extensively investigated for use in tablet- or capsule-based drug delivery systems, the potential for use of IPNs in drug release medical devices remains largely unexplored. Here, we investigated the use of silicone-hydrogel IPNs as a catheter material to provide slow anti-bacterial drug-release functionality. IPN catheters were produced by the sequential method, using supercritical CO2 as a solvent to polymerize and crosslink poly(2-hydroxyethyl methacrylate) (PHEMA) in silicone elastomer. The design was tested against Staphylococcus aureus colonization after loading with dicloxacillin (DCX) alone or in combination with thioridazine (TDZ), the latter of which is known to synergistically potentiate the antibacterial effect of DCX against both methicillin-sensitive and methicillin-resistant S. aureus. The hydrophilic PHEMA component allowed for drug loading in the catheters by passive diffusion and provided controlled release properties. The drug-loaded IPN material inhibited bacterial growth on agar plates for up to two weeks and in blood cultures for up to five days, and it withstood 24h of seeding with resilient biofilm aggregates. The combined loading of DCX+TDZ enhanced the antibacterial efficiency in static in vitro experiments, although release analyses revealed that this effect was due to an enhanced loading capacity of DCX when co-loaded with TDZ. Lastly, the IPN catheters were tested in a novel porcine model of central venous catheter-related infection, in which drug-loaded IPN catheters were found to significantly decrease the frequency of infection.

Does C-reactive protein independently predict mortality in adult community-acquired bacteremia patients with known sepsis severity?

We evaluated whether sepsis severity and C-reactive protein (CRP) level on admission prognostically corroborated or annulled each other in adult patients with incident community-acquired bacteremia (Funen, Denmark, 2000-2008). We used logistic regression and area under the receiver operating characteristic curve (AUC) to evaluate 30-day mortality in four models: (i) age, gender, comorbidity, bacteria, and ward. (ii) Model 1 and sepsis severity. (iii) Model 1 and CRP. (iv) Model 1, sepsis severity, and CRP. Altogether, 416 of 1999 patients died within 30 days. CRP independently predicted 30-day mortality [Model 4, odds ratio (95% CIs) for 100 mg/L: 1.16 (1.06-1.27)], but it did not contribute to the AUC (Model 2 vs Model 4: p = 0.31). In the 963 non-severe sepsis patients, CRP independently predicted 30-day mortality [Model 4: 1.42 (1.20-1.69)] and it increased the AUC (Model 2 vs Model 4: p = 0.06), thus CRP contributed as much as sepsis severity to prognosis.

Serum YKL-40 predicts long-term mortality in patients with stable coronary disease: a prognostic study within the CLARICOR trial.

OBJECTIVE: We investigated whether the inflammatory biomarker YKL-40 could improve the long-term prediction of death made by common risk factors plus high-sensitivity C-reactive protein (hs-CRP) and N-terminal-pro-B natriuretic peptide (NT-proBNP) in patients with stable coronary artery disease (CAD). BACKGROUND: Non-hospitalized CAD patients are usually followed in general practice. There is a need for identify biomarkers which could help to foresee the prognoses of these patients. Elevated serum YKL-40 is a short-term predictor for myocardial infarction, cardiovascular mortality and all-cause mortality in patients with stable CAD. METHODS: Serum YKL-40, hs-CRP, and NT-proBNP were measured in 4265 (97.6%) of the 4372 patients with stable CAD included in the CLARICOR trial, and death was registered in a 6-years follow-up period. RESULTS: The median serum YKL-40 was 110 µg/L [IQR=93], hs-CRP 2.8 mg/L [IQR=4.74], and NT-proBNP 203 ng/L [IQR=407]. During 6 years follow-up period 923 (21.1%) patients died. After adjustment for type of intervention, risk factors (age, sex, hypertension, diabetes, smoking status, and previous myocardial infarction) and medical treatment (diuretics, digoxin, and statin) serum YKL-40 (transformed as ln(max(82, YKL-40/µg/L)) was significantly associated with all-cause mortality [hazard ratio (HR)=1.55, 95% CI=1.39-1.73, p<0.001]. After additional adjustment for ln(hs-CRP) and ln(NT-proBNP) this was still true [HR=1.38, 95% CI=1.21-1.53, p<0.001]. CONCLUSIONS: Serum YKL-40 is a predictor of long-term mortality in patients with stable CAD independent of common risk factors and ln(hs-CRP) and ln(NT-proBNP). Serum YKL-40 can be used for prognostication in these patients.

Good interobserver agreement was attainable on outcome adjudication in patients with stable coronary heart disease.

OBJECTIVE: In clinical trials, agreement on outcomes is of utmost importance for valid estimation of intervention effects. As there is limited knowledge about adjudicator agreement in cardiology, we examined the level of agreement among three cardiology specialists adjudicating all possible events in a randomized controlled clinical trial of patients with stable coronary heart disease. STUDY DESIGN AND SETTING: All information (hospital records, death certificates, etc.) was forwarded to two randomly selected blinded adjudicators. If they disagreed, the third arbiter had to choose the more likely of the two alternatives. Files of 5,475 nonfatal and 362 fatal events were evaluated. RESULTS: For nonfatal outcomes, pairwise kappa values ranged from 0.75 to 0.80. The three adjudicators had 4.3%, 9.5%, and 6.1% of their nonfatal outcome classifications overruled by their arbiter. If stable angina pectoris, unstable angina pectoris, and acute myocardial infarction were treated as one, agreement increased minimally. For fatal outcomes, the pairwise kappa values ranged from 0.65 to 0.90. The three adjudicators had 12%, 9%, and 10% of their death classifications overruled. CONCLUSION: Specialists in cardiology can attain a reasonably high agreement on outcomes in patients with stable coronary heart disease.

Decreased material-activation of the complement system using low-energy plasma polymerized poly(vinyl pyrrolidone) coatings.

In the current study we investigate the activation of blood complement on medical device silicone rubber and present a plasma polymerized vinyl pyrrolidone (ppVP) coating which strongly decreases surface-activation of the blood complement system. We show that uncoated silicone and polystyrene are both potent activators of the complement system, measured both as activated, deposited C3b and quantifying fluid-phase release of the cleavage fragment C3c. The ppVP coated silicone exhibits approximately 90% reduced complement activation compared to untreated silicone. Quartz crystal microbalance with dissipation (QCM-D) measurements show relatively strong adsorption of blood proteins including native C3 to the ppVP surface, indicating that reduction of complement activation on ppVP is neither a result of low protein adsorption nor lower direct C3-binding, and is therefore possibly a consequence of differences in the adsorbed protein layer composition. The alternative and classical complement pathways are barely detectable on ppVP while the lectin pathway through MBL/ficolin-2 deposition remains active on ppVP suggesting this pathway is responsible for the remaining subtle activation on the ppVP coated surface. The ppVP surface is furthermore characterized physically and chemically using scanning electron microscopy (SEM), x-ray photoelectron spectroscopy (XPS) and Fourier transform infrared (FTIR), which indicates preservation of chemical functionality by the applied plasma process. Overall, the ppVP coating shows a potential for increasing complement-compatibility of blood-contacting devices.

Antibacterial properties of EMLA and lidocaine in wound tissue biopsies for culturing.

If a tissue biopsy from a chronic wound is sampled for culture, the antibacterial properties of local anesthetics may pose a problem in producing false-negative results. The purpose of this study was to investigate the effects of EMLA (AstraZeneca) and lidocaine on common wound pathogenic bacteria. An in vitro study of a total of 25 clinical isolates and ATCC reference strains of Staphylococcus aureus (including methicillin-resistant S. aureus), Escherichia coli, Pseudomonas aeruginosa, and Streptococcus pyogenes was conducted. The isolates were exposed to the local anesthetic drugs (and some of their contents separately) at 35 degrees C over a 24-hour period and time-kill curves were recorded. No culture media were used and saline was used for controls. EMLA was found to have a rapid acting and powerful antibacterial effect and should not be used before culturing tissue samples. Lidocaine 1% was found not to inhibit the bacterial strains when exposure time was held below 2 hours. We conclude that culturing tissue from a wound biopsy is safe within 2 hours when a pure, preservative-free lidocaine 1% solution is used.