Eric C. M. van Gorp

Long pentraxin PTX3 is associated with mortality and disease severity in severe Leptospirosis

OBJECTIVE To evaluate the long pentraxin PTX3 in patients with severe leptospirosis and to compare the results with the widely used short pentraxin C-reactive protein and the pro-inflammatory cytokines IL-6 and IL-8. METHODS This observational cohort study was carried out in Semarang, Indonesia, where leptospirosis is endemic and mortality is high. Consecutive patients with severe leptospirosis were sampled on admission and during follow-up. RESULTS A total number of 52 patients entered the study, the mortality was 27%. Severe leptospirosis patient yielded elevated plasma PTX3 levels. PTX3 correlated with IL-8 and to a lesser extent with CRP and IL-6 levels. High levels of PTX3, IL-6 and IL-8 were associated with mortality (OR 5.6, 95%CI: 1.2-26; OR 3.2, 95%CI: 1.2-8.1; OR 6.5, 95%CI: 1.5-28). Moreover, PTX3 levels were associated with disease severity (OR 9.5; 95%CI: 2.9-45). This association was unique, since none of the other markers showed this relation. C-reactive protein was not able to differentiate the severe from the severest cases. CONCLUSIONS The long pentraxin PTX3 is elevated in patients with severe leptospirosis and is associated with fatal disease and disease severity. PTX3 may be used as a marker to monitor disease severity in severe leptospirosis or predict outcome.

Soluble ST2 Levels Are Associated with Bleeding in Patients with Severe Leptospirosis

Background Severe leptospirosis features bleeding and multi-organ failure, leading to shock and death. Currently it is assumed that both exaggerated inflammation and immune suppression contribute to mortality in sepsis. Indeed, several proinflammatory cytokines are reported to be induced during leptospirosis. Toll-like receptors, which play an important role in the initiation of an innate immune response, are inhibited by negative regulators including the membrane-bound ST2 (mST2) receptor. Soluble ST2 (sST2) has been implicated to inhibit signaling through mST2. The aim of this study was to determine the extent of sST2 and (pro-) inflammatory cytokine release in patients with severe leptospirosis. Methodology and Principal Findings In an observational study, 68 consecutive cases of severe leptospirosis were included. Soluble ST2 and cytokines (TNF-α, IL-1β, IL-6, IL-8, and IL-10) were repeatedly measured. To determine whether blood cells are a source of sST2 during infection, we undertook an in vitro experiment: human whole blood and peripheral blood mononuclear cells (PBMC) were stimulated with viable pathogenic Leptospira. All patients showed elevated sST2, IL-6, IL-8, and IL-10 levels on admission. Admission sST2 levels correlated with IL-6, IL-8, and IL-10. Thirty-four patients (50%) showed clinical bleeding. Soluble ST2 levels were significantly associated with bleeding overall (OR 2.0; 95%CI: 1.2–3.6) and severe bleeding (OR 5.1; 95%CI: 1.1–23.8). This association was unique, since none of the cytokines showed this correlation. Moreover, sST2 was associated with mortality (OR 2.4; 95%CI: 1.0–5.8). When either whole blood or isolated PBMCs were stimulated with Leptospira in vitro, no sST2 production could be detected. Conclusions Patients with severe leptospirosis demonstrated elevated plasma sST2 levels. Soluble ST2 levels were associated with bleeding and mortality. In vitro experiments showed that (white) blood cells are probably not the source. In this regard, sST2 could be an indicative marker for tissue damage in patients suffering from severe leptospirosis.

Effects on Coagulation and Fibrinolysis Induced by Influenza in Mice With a Reduced Capacity to Generate Activated Protein C and a Deficiency in Plasminogen Activator Inhibitor Type 1

Influenza infections increase the risk of diseases associated with a prothrombotic state, such as venous thrombosis and atherothrombotic diseases. However, it is unclear whether influenza leads to a prothrombotic state in vivo. To determine whether influenza activates coagulation, we measured coagulation and fibrinolysis in influenza-infected C57BL/6 mice. We found that influenza increased thrombin generation, fibrin deposition, and fibrinolysis. In addition, we used various anti- and prothrombotic models to study pathways involved in the influenza-induced prothrombotic state. A reduced capacity to generate activated protein C in TMpro/pro mice increased thrombin generation and fibrinolysis, whereas treatment with heparin decreased thrombin generation in influenza-infected C57Bl/6 mice. Thrombin generation was not changed in hyperfibrinolytic mice, deficient in plasminogen activator inhibitor type-1 (PAI-1−/−); however, increased fibrin degradation was seen. Treatment with tranexamic acid reduced fibrinolysis, but thrombin generation was unchanged. We conclude that influenza infection generates thrombin, increased by reduced levels of protein C and decreased by heparin. The fibrinolytic system appears not to be important for thrombin generation. These findings suggest that influenza leads to a prothrombotic state by coagulation activation. Heparin treatment reduces the influenza induced prothrombotic state.

Predictors and treatment strategies of HIV-related fatigue in the combined antiretroviral therapy era

Objective To assess predictors and reported treatment strategies of HIV-related fatigue in the combined antiretroviral (cART) era. Method Five databases were searched and reference lists of pertinent articles were checked. Studies published since 1996 on predictors or therapy of HIV-related fatigue measured by a validated instrument were selected. Results A total of 42 studies met the inclusion criteria. The reported HIV-related fatigue prevalence in the selected studies varied from 33 to 88%. The strongest predictors for sociodemographic variables were unemployment and inadequate income. Concerning HIV-associated factors, the use of cART was the strongest predictor. Comorbidity and sleeping difficulties were important factors when assessing physiological influences. Laboratory parameters were not predictive of fatigue. The strongest and most uniform associations were observed between fatigue and psychological factors such as depression and anxiety. Reported therapeutic interventions for HIV-related fatigue include testosterone, psycho-stimulants (dextroamphetamine, methylphenidate hydrochloride, pemoline, modafinil), dehydroepiandrosterone, fluoxetine and cognitive behavioural or relaxation therapy. Conclusion HIV-related fatigue has a high prevalence and is strongly associated with psychological factors such as depression and anxiety. A validated instrument should be used to measure intensity and consequences of fatigue in HIV-infected individuals. In the case of fatigue, clinicians should not only search for physical mechanisms, but should question depression and anxiety in detail. There is a need for intervention studies comparing the effect of medication (antidepressants, anxiolytics) and behavioural interventions (cognitive-behavioural therapy, relaxation therapy, graded exercise therapy) to direct the best treatment strategy. Treatment of HIV-related fatigue is important in the care for HIV-infected patients and requires a multidisciplinary approach.

Safety and immunogenicity of a modified-vaccinia-virus-Ankara-based influenza A H5N1 vaccine: a randomised, double-blind phase 1/2a clinical trial.

BACKGROUND Modified vaccinia virus Ankara (MVA) is a promising viral vector platform for the development of an H5N1 influenza vaccine. Preclinical assessment of MVA-based H5N1 vaccines showed their immunogenicity and safety in different animal models. We aimed to assess the safety and immunogenicity of the MVA-haemagglutinin-based H5N1 vaccine MVA-H5-sfMR in healthy individuals. METHODS In a single-centre, double-blind phase 1/2a study, young volunteers (aged 18-28 years) were randomly assigned with a computer-generated list in equal numbers to one of eight groups and were given one injection or two injections intramuscularly at an interval of 4 weeks of a standard dose (10(8) plaque forming units [pfu]) or a ten times lower dose (10(7) pfu) of the MVA-H5-sfMR (vector encoding the haemagglutinin gene of influenza A/Vietnam/1194/2004 virus [H5N1 subtype]) or MVA-F6-sfMR (empty vector) vaccine. Volunteers and physicians who examined and administered the vaccine were masked to vaccine assignment. Individuals who received the MVA-H5-sfMR vaccine were eligible for a booster immunisation 1 year after the first immunisation. Primary endpoint was safety. Secondary outcome was immunogenicity. The trial is registered with the Dutch Trial Register, number NTR3401. FINDINGS 79 of 80 individuals who were enrolled completed the study. No serious adverse events were identified. 11 individuals reported severe headache and lightheadedness, erythema nodosum, respiratory illness (accompanied by influenza-like symptoms), sore throat, or injection-site reaction. Most of the volunteers had one or more local (itch, pain, redness, and swelling) and systemic reactions (rise in body temperature, headache, myalgia, arthralgia, chills, malaise, and fatigue) after the first, second, and booster immunisations. Individuals who received the 10(7) dose had fewer systemic reactions. The MVA-H5-sfMR vaccine at 10(8) pfu induced significantly higher antibody responses after one and two immunisations than did 10(7) pfu when assessed with haemagglutination inhibition geometric mean titre at 8 weeks against H5N1 A/Vietnam/1194/2004 (30·2 [SD 3·8] vs 9·2 [2·3] and 108·1 [2·4] vs 15·8 [3·2]). 27 of 39 eligible individuals were enrolled in the booster immunisation study. A single shot of MVA-H5-sfMR 10(8) pfu prime immunisation resulted in higher antibody responses after the booster immunisation than did two shots of MVA-H5-sfMR at the ten times lower dose. INTERPRETATION The MVA-based H5N1 vaccine was well tolerated and immunogenic and therefore the vaccine candidates arising from the MVA platform hold great promise for rapid development in response to a future influenza pandemic threat. However, the immunogenicity of this vaccine needs to be compared with conventional H5N1 inactivated non-adjuvanted vaccine candidates in head-to-head clinical trials. FUNDING European Research Council.

Additional value of procalcitonin for diagnosis of infection in patients with fever at the emergency department

Objective: First, to determine whether procalcitonin (PCT) significantly adds diagnostic value in terms of sensitivity and specificity to a common set of markers of infection, including C-reactive protein (CRP), at the Emergency Department. Second, to create a simple scoring rule implementing PCT values. Third, to determine and compare associations of CRP and PCT with clinical outcomes. Design: The additional diagnostic value of PCT was determined using multiple logistic regression analysis. A score was developed to help distinguish patients with a culture-proven bacterial infection from patients not needing antibiotic treatment using 16 potential clinical and laboratory variables. The prognostic value of CRP and PCT was determined using Spearmans correlation and logistic regression. Setting: Emergency Department of a 310-bed teaching hospital. Patients: Patients between 18 and 85 years old presenting with fever to the Emergency Department. Interventions: None. Measurements and Main Results: A total of 211 patients were studied (infection confirmed, n = 73; infection likely, n = 58; infection not excluded, n = 46; no infection, n = 34). CRP and chills were the strongest predictors for the diagnosis of bacterial infection. After addition of PCT to these parameters, model fit significantly improved (p = .003). The resulting scoring rule (score = 0.01 * CRP + 2 * chills + 1 * PCT) was characterized by an AUC value of 0.83 (sensitivity 79%; specificity of 71%), which was more accurate than physician judgment or SIRS (systemic inflammatory response syndrome). PCT levels were significantly associated with admission to a special care unit, duration of intravenous antibiotic use, total duration of antibiotic treatment, and length of hospital stay, whereas CRP was related only to the latter two variables. Conclusions: These data suggest that PCT may be a valuable addition to currently used markers of infection for diagnosis of infection and prognosis in patients with fever at the Emergency Department.

Changes in the Plasma Lipid Profile as a Potential Predictor of Clinical Outcome in Dengue Hemorrhagic Fever

In 50 consecutive children admitted to the intensive care unit with the clinical diagnosis of dengue hemorrhagic fever (DHF)/dengue shock syndrome (grade III or IV), 20 patients with mild DHF (grade I or II), and 20 healthy control patients, the plasma lipid profile was measured. Levels of total plasma cholesterol, high-density lipoprotein, and low-density lipoprotein were significantly decreased in patients with the severest cases, compared with patients with mild DHF and healthy controls. Changes in the plasma lipid profile differentiate between patients with different stages of DHF disease severity and could be used as a potential predictor for clinical outcome.

Hepatotoxicity following nevirapine-containing regimens in HIV-1-infected individuals

To determine the incidence of hepatotoxicity and to investigate whether plasma concentrations of nevirapine, in addition to other risk factors, could predict hepatotoxicity during treatment with nevirapine-containing regimens, we conducted a retrospective analysis with data from 174 individuals infected with human immunodeficiency virus-1 (HIV-1). During regular visits to the clinic, blood samples were collected for the determination of nevirapine plasma concentrations and clinical chemistry parameters including liver enzymes (LEs) and total bilirubin (TBR). Severe hepatotoxicity was defined as a grade > or =3 elevation in at least one of the tested LEs or TBR levels while on therapy. Analysis of predictive factors was focused on increases in aspartate aminotransferase (ASAT) and/or alanine aminotransferase (ALAT) to grade > or =2. Grade > or =3 elevation developed with an incidence of 0.15 per patient year (PY); only 3.4% of the patients developed grade > or =3 values for ASAT and/or ALAT (incidence 0.03 per PY). We found that patients who use a protease inhibitor (PI) in a nevirapine-containing regimen and patients who have chronic hepatitis B (HBV) infection are at a higher risk for the development of increases in ASAT and/or ALAT to grade > or =2. In contrast, the plasma concentration of nevirapine does not appear to be a predictive factor in this study population.

Dengue disease severity in Indonesian children: an evaluation of the World Health Organization classification system

BackgroundDengue disease severity is usually classified using criteria set up by the World Health Organization (WHO). We aimed to assess the diagnostic accuracy of the WHO classification system and modifications to this system, and evaluated their potential practical usefulness.MethodsPatients, admitted consecutively to the hospital with severe dengue, were classified using the WHO classification system and modifications to this system. Treating physicians were asked to classify patients immediately after discharge. We calculated the sensitivity of the various classification systems for the detection of shock and the agreement between the various classification systems and the treating physicians classification.ResultsOf 152 patients with confirmed dengue, sixty-six (43%) had evidence of circulatory failure. The WHO classification system had a sensitivity of 86% (95%CI 76–94) for the detection of patients with shock. All modifications to the WHO classification system had a higher sensitivity than the WHO classification system (sensitivity ranging from 88% to 99%). The WHO classification system was in only modest agreement with the intuitive classification by treating physicians whereas several modified classification systems were in good agreement.ConclusionThe use of the WHO classification system to classify dengue disease severity is to be questioned, because it is not accurate in correctly classifying dengue disease severity and it lacks sufficient agreement with clinical practice.

Intracellular and plasma steady‐state pharmacokinetics of raltegravir, darunavir, etravirine and ritonavir in heavily pre‐treated HIV‐infected patients

AIM To study the steady-state plasma and intracellular pharmacokinetics of raltegravir, etravirine, darunavir and ritonavir in heavily pre-treated patients. METHODS Patients on a salvage regimen containing raltegravir, etravirine, darunavir and ritonavir were eligible for inclusion. During a 12 h dosing interval plasma and peripheral blood mononuclear cells were collected. Drug concentrations were measured using a validated LC-MS/MS assay and pharmacokinetic analysis was performed using non-linear mixed effect modelling. RESULTS Irregular absorption was observed with raltegravir and darunavir, which may be caused by enterohepatic cycling. Relative bioavailability of ritonavir was low, when compared with other ritonavir regimens. Raltegravir plasma pharmacokinetics showed wide interpatient variability, while intracellular raltegravir concentrations could not be detected (<0.001 mg l(-1) in cell lysate). The intracellular to plasma ratios for etravirine, darunavir and ritonavir were 12.9, 1.32 and 7.72, respectively, and the relative standard error of these estimates were 16.3%, 12.3% and 13.0%. CONCLUSIONS The observed distinct intracellular accumulation indicated that these drugs have different affinity for the cellular compartment. The relatively high intracellular accumulation of etravirine may explain its efficacy and its previously described absence of PK-PD relationships in the therapeutic concentration range, when compared with other non-nucleoside reverse transcriptase inhibitors. Lastly, the intracellular concentrations of ritonavir seem sufficient for inhibition of viral replication in the cellular compartment in PI-naive patients, but not in patients with HIV harbouring PI resistance.