FDA-approved drug screening in patient-derived organoids demonstrates potential of drug repurposing for rare cystic fibrosis genotypes.

BACKGROUND: Preclinical cell-based assays that recapitulate human disease play an important role in drug repurposing. We previously developed a functional forskolin induced swelling (FIS) assay using patient-derived intestinal organoids (PDIOs), allowing functional characterization of CFTR, the gene mutated in people with cystic fibrosis (pwCF). CFTR function-increasing pharmacotherapies have revolutionized treatment for approximately 85% of people with CF who carry the most prevalent F508del-CFTR mutation, but a large unmet need remains to identify new treatments for all pwCF. METHODS: We used 76 PDIOs not homozygous for F508del-CFTR to test the efficacy of 1400 FDA-approved drugs on improving CFTR function, as measured in FIS assays. The most promising hits were verified in a secondary FIS screen. Based on the results of this secondary screen, we further investigated CFTR elevating function of PDE4 inhibitors and currently existing CFTR modulators. RESULTS: In the primary screen, 30 hits were characterized that elevated CFTR function. In the secondary validation screen, 19 hits were confirmed and categorized in three main drug families: CFTR modulators, PDE4 inhibitors and tyrosine kinase inhibitors. We show that PDE4 inhibitors are potent CFTR function inducers in PDIOs where residual CFTR function is either present, or created by additional compound exposure. Additionally, upon CFTR modulator treatment we show rescue of CF genotypes that are currently not eligible for this therapy. CONCLUSION: This study exemplifies the feasibility of high-throughput compound screening using PDIOs. We show the potential of repurposing drugs for pwCF carrying non-F508del genotypes that are currently not eligible for therapies. ONE-SENTENCE SUMMARY: We screened 1400 FDA-approved drugs in CF patient-derived intestinal organoids using the previously established functional FIS assay, and show the potential of repurposing PDE4 inhibitors and CFTR modulators for rare CF genotypes.

Man vs. machine: A meta-analysis on the added value of human support in text-based internet treatments ("e-therapy") for mental disorders.

Guided internet-based treatment is more efficacious than completely unguided or self-guided internet-based treatment, yet within the spectrum of guidance, little is known about the added value of human support compared to more basic forms of guidance. The primary aims of this meta-analysis were: (1) to examine whether human guidance was more efficacious than technological guidance in text-based internet treatments ("e-therapy") for mental disorders, and (2) whether more intensive human guidance outperformed basic forms of human guidance. PsycINFO, PubMed and Web of Science were systematically searched for randomized controlled trials that directly compared various types and degrees of online guidance. Thirty-one studies, totaling 6215 individuals, met inclusion criteria. Results showed that human guidance was slightly more efficacious than technological guidance, both in terms of symptom reduction (g = 0.11; p < .01) and adherence (0.26 < g < 0.29; p's < 0.01). On the spectrum of human support, results were slightly more favorable for regular guidance compared to optional guidance, but only in terms of adherence (OR = 1.89, g = 0.35; p < .05). Higher qualification of online counselors was not associated with efficacy. These findings extend and refine previous reports on guided and unguided online treatments.

The presence of cardiotropic viral genomes is not increased in atrial tissue of atrial fibrillation patients.

BACKGROUND: Infections with potentially cardiotropic viruses are associated with the development of atrial fibrillation (AF). However, whether direct viral infection of the atria is involved in the pathogenesis of AF is unclear. We have therefore analysed the presence of cardiotropic viral genomes in AF patients. METHODS: Samples of left atrial tissue were obtained from 50 AF patients (paroxysmal, n = 20; long-standing persistent/permanent, n = 30) during cardiac surgery and from autopsied control patients (n = 14). Herein, the presence of PVB19, EBV, CMV, HHV­6, adenovirus and enterovirus genomes was determined by polymerase chain reaction. The densities of CD45+ and CD3+ cells and fibrosis in the atria were quantified by (immuno)histochemistry. RESULTS: Of the tested viruses only the PVB19 genome was detected in the atria of 10% of patients, paroxysmal AF (2 of 20) and long-standing persistent/permanent AF (3 of 30). Conversely, in 50% of controls (7 of 14) PVB19 genome was found. No significant association was found between PVB19 and CD45+ and CD3+ cells, or between the presence of PVB19 and fibrosis, in either control or AF patients. CONCLUSION: The presence of viral genomes is not increased in the atria of AF patients. These results do not support an important role for viral infection of the atria in the pathogenesis of AF.

Atrial inflammation in different atrial fibrillation subtypes and its relation with clinical risk factors.

OBJECTIVE: Inflammation of the atria is an important factor in the pathogenesis of atrial fibrillation (AF). Whether the extent of atrial inflammation relates with clinical risk factors of AF, however, is largely unknown. This we have studied comparing patients with paroxysmal and long-standing persistent/permanent AF. METHODS: Left atrial tissue was obtained from 50 AF patients (paroxysmal = 20, long-standing persistent/permanent = 30) that underwent a left atrial ablation procedure either or not in combination with coronary artery bypass grafting and/or valve surgery. Herein, the numbers of CD45+ and CD3+ inflammatory cells were quantified and correlated with the AF risk factors age, gender, diabetes, and blood CRP levels. RESULTS: The numbers of CD45+ and CD3+ cells were significantly higher in the adipose tissue of the atria compared with the myocardium in all AF patients but did not differ between AF subtypes. The numbers of CD45+ and CD3+ cells did not relate significantly to gender or diabetes in any of the AF subtypes. However, the inflammatory infiltrates as well as CK-MB and CRP blood levels increased significantly with increasing age in long-standing persistent/permanent AF and a moderate positive correlation was found between the extent of atrial inflammation and the CRP blood levels in both AF subtypes. CONCLUSION: The extent of left atrial inflammation in AF patients was not related to the AF risk factors, diabetes and gender, but was associated with increasing age in patients with long-standing persistent/permanent AF. This may be indicative for a role of inflammation in the progression to long-standing persistent/permanent AF with increasing age.

Prophylactic veno-arterial extracorporeal membrane oxygenation in patients undergoing high-risk percutaneous coronary intervention.

PURPOSE: Complex high-risk percutaneous coronary intervention (PCI) is challenging and frequently accompanied by haemodynamic instability. Veno-arterial extracorporeal membrane oxygenation (VA-ECMO) can provide cardiopulmonary support in high-risk PCI. However, the outcome is unclear. METHODS: A two-centre, retrospective study was performed of all patients undergoing high-risk PCI and receiving VA-ECMO for cardiopulmonary support. RESULTS: A total of 14 patients (92% male, median age 69 (53-83) years), of whom 50% had previous coronary artery disease in the form of a coronary artery bypass graft (36%) and a PCI (14%) underwent high-risk PCI and received VA-ECMO support. The main target lesion was a left main coronary artery in 78%, a left anterior descending artery in 14%, a right coronary artery in 7%, and 71% underwent multi-vessel PCI in addition to main target vessel PCI. The median SYNTAX score was 27.2 (8-42.5) and in 64% (9/14) there was a chronic total occlusion. Left ventricular function was mildly impaired in 7% (1/14), moderately impaired in 14% (2/14) and severely impaired in 64% (9/14). Cannulation was femoral-femoral in all patients. Median ECMO run was 2.57 h (1-4). Survival was 93% (13/14). One patient died during hospitalisation due to refractory cardiac failure. All other patients survived to discharge. Complications occurred in 14% (2/14), with one patient developing a transient ischaemic attack post-ECMO and one patient developing a thrombus in the femoral vein used for ECMO cannulation. CONCLUSION: VA-ECMO in high-risk PCI is feasible with a good outcome. It can be successfully used for cardiopulmonary support in selected patients.

Hemangioma of the Atherosclerotic Changed Aortic Valve.

The incidence of heart valve hemangioma is very low and is mostly observed in the mitral and tricuspid valve. In 2006, two cases of aortic valve hemangioma were reported for the first time, including one with calcifying aortic valve stenosis. We now present a case of aortic valve hemangioma in a patient suffering from aortic valve insufficiency combined with atherosclerotic thickening.

Postoperative microcirculatory perfusion and endothelial glycocalyx shedding following cardiac surgery with cardiopulmonary bypass.

We investigated microcirculatory perfusion disturbances following cardiopulmonary bypass in the early postoperative period and whether the course of these disturbances mirrored restoration of endothelial glycocalyx integrity. We performed sublingual sidestream dark field imaging of the microcirculation during the first three postoperative days in patients who had undergone on-pump coronary artery bypass graft surgery. We calculated the perfused vessel density, proportion of perfused vessels and perfused boundary region. Plasma was obtained to measure heparan sulphate and syndecan-1 levels as glycocalyx shedding markers. We recruited 17 patients; the mean (SD) duration of non-pulsatile cardiopulmonary bypass was 103 (18) min, following which 491 (29) ml autologous blood was transfused through cell salvage. Cardiopulmonary bypass immediately decreased both microcirculatory perfused vessel density; 11 (3) vs. 16 (4) mm.mm-2 , p = 0.052 and the proportion of perfused vessels; 92 (5) vs. 69 (9) %, p < 0.0001. The proportion of perfused vessels did not increase after transfusion of autologous salvaged blood following cardiopulmonary bypass; 72 (7) %, p = 0.19 or during the first three postoperative days; 71 (5) %, p < 0.0001. The perfused boundary region increased after cardiopulmonary bypass; 2.2 (0.3) vs. 1.9 (0.3) µm, p = 0.037 and during the first three postoperative days; 2.4 (0.3) vs. 1.9 (0.3) µm, p = 0.003. Increased plasma heparan sulphate levels were inversely associated with the proportion of perfused vessels during cardiopulmonary bypass; R = -0.49, p = 0.02. Plasma syndecan-1 levels were inversely associated with the proportion of perfused vessels during the entire study period; R = -0.51, p < 0.0001. Our study shows that cardiopulmonary bypass-induced acute microcirculatory perfusion disturbances persist in the first three postoperative days, and are associated with prolonged endothelial glycocalyx shedding. This suggests prolonged impairment and delayed recovery of both microcirculatory perfusion and function after on-pump cardiac surgery.

Vasculotide, an angiopoietin-1 mimetic, reduces pulmonary vascular leakage and preserves microcirculatory perfusion during cardiopulmonary bypass in rats.

BACKGROUND: Cardiopulmonary bypass (CPB) during cardiac surgery impairs microcirculatory perfusion and is paralleled by vascular leakage. The endothelial angiopoietin/Tie2 system controls microvascular leakage. This study investigated whether targeting Tie2 with the angiopoietin-1 mimetic vasculotide reduces vascular leakage and preserves microcirculatory perfusion in a rat CPB model. METHODS: Rats were subjected to 75 min of CPB after treatment with vasculotide or phosphate buffered solution as control or underwent a sham procedure. Microcirculatory perfusion and leakage were assessed with intravital microscopy (n=10 per group) and Evans blue dye extravasation (n=13 per group), respectively. Angiopoietin-1, -2, and Tie2 protein and gene expression were determined in plasma, kidney, and lung. RESULTS: CPB immediately impaired microcirculatory perfusion [5 (4-8) vs 10 (7-12) vessels per recording, P=0.002] in untreated CPB rats compared with sham, which persisted after weaning from CPB. CPB increased circulating angiopoeietin-1, -2, and soluble Tie2 concentrations and reduced Tie2 messenger ribonucleic acid (mRNA) expression in kidney and lung. Moreover, CPB increased Evans blue dye leakage in kidney [12 (8-25) vs 7 (1-12) µg g-1, P=0.04] and lung [and 23 (13-60) vs 6 (4-16) µg g-1, P=0.001] compared with sham. Vasculotide treatment preserved microcirculatory perfusion during and after CPB. Moreover, vasculotide treatment reduced Evans blue dye extravasation in lung compared with CPB control [18 (6-28) µg g-1vs 23 (13-60) µg g-1, P=0.04], but not in kidney [10 (3-23) vs 12 (8-25) µg g-1, P=0.38]. Vasculotide did not affect circulating or mRNA expression of angiopoietin-1, -2, and Tie2 concentrations compared with untreated CPB controls. CONCLUSIONS: Treatment with the angiopoietin-1 mimetic vasculotide reduced pulmonary vascular leakage and preserved microcirculatory perfusion during CPB in a rat model.

Reduction of vascular leakage by imatinib is associated with preserved microcirculatory perfusion and reduced renal injury markers in a rat model of cardiopulmonary bypass.

BACKGROUND: Cardiopulmonary bypass during cardiac surgery leads to impaired microcirculatory perfusion. We hypothesized that vascular leakage is an important contributor to microcirculatory dysfunction. Imatinib, a tyrosine kinase inhibitor, has been shown to reduce vascular leakage in septic mice. We investigated whether prevention of vascular leakage using imatinib preserves microcirculatory perfusion and reduces organ injury markers in a rat model of cardiopulmonary bypass. METHODS: Male Wistar rats underwent cardiopulmonary bypass after treatment with imatinib or vehicle (n=8 per group). Cremaster muscle microcirculatory perfusion and quadriceps microvascular oxygen saturation were measured using intravital microscopy and reflectance spectroscopy. Evans Blue extravasation was determined in separate experiments. Organ injury markers were determined in plasma, intestine, kidney, and lungs. RESULTS: The onset of cardiopulmonary bypass decreased the number of perfused microvessels by 40% in the control group [9.4 (8.6-10.6) to 5.7 (4.8-6.2) per microscope field; P<0.001 vs baseline], whereas this reduction was not seen in the imatinib group. In the control group, the number of perfused capillaries remained low throughout the experiment, whilst perfusion remained normal after imatinib administration. Microvascular oxygen saturation was less impaired after imatinib treatment compared with controls. Imatinib reduced vascular leakage and decreased fluid resuscitation compared with control [3 (3-6) vs 12 ml (7-16); P=0.024]. Plasma neutrophil-gelatinase-associated-lipocalin concentrations were reduced by imatinib. CONCLUSIONS: Prevention of endothelial barrier dysfunction using imatinib preserved microcirculatory perfusion and oxygenation during and after cardiopulmonary bypass. Moreover, imatinib-induced protection of endothelial barrier integrity reduced fluid-resuscitation requirements and attenuated renal and pulmonary injury markers.

Anticoagulant and side-effects of protamine in cardiac surgery: a narrative review.

Neutralisation of systemic anticoagulation with heparin in cardiac surgery with cardiopulmonary bypass requires protamine administration. If adequately dosed, protamine neutralises heparin and reduces the risk of postoperative bleeding. However, as its anticoagulant properties are particularly exerted in the absence of heparin, overdosing of protamine may contribute to bleeding and increased transfusion requirements. This narrative review describes the mechanisms underlying the anticoagulant properties and side-effects of protamine, and the impact of protamine dosing on the activated clotting time and point-of-care viscoelastic test results, and explains the distinct protamine dosing strategies in relation to haemostatic activation and postoperative bleeding. The available evidence suggests that protamine dosing should not exceed a protamine-to-heparin ratio of 1:1. In particular, protamine-to-heparin dosing ratios >1 are associated with more postoperative 12 h blood loss. The optimal protamine-to-heparin ratio in cardiac surgery has, however, not yet been elaborated, and may vary between 0.6 and 1.0 based on the initial heparin dose.

Lung ultrasound compared with chest X-ray in diagnosing postoperative pulmonary complications following cardiothoracic surgery: a prospective observational study.

Postoperative pulmonary complications are common after cardiothoracic surgery and are associated with adverse outcomes. The ability to detect postoperative pulmonary complications using chest X-rays is limited, and this technique requires radiation exposure. Little is known about the diagnostic accuracy of lung ultrasound for the detection of postoperative pulmonary complications after cardiothoracic surgery, and we therefore aimed to compare lung ultrasound with chest X-ray to detect postoperative pulmonary complications in this group of patients. We performed this prospective, observational, single-centre study in a tertiary intensive care unit treating adult patients who had undergone cardiothoracic surgery. We recorded chest X-ray findings upon admission and on postoperative days 2 and 3, as well as rates of postoperative pulmonary complications and clinically-relevant postoperative pulmonary complications that required therapy according to the treating physician as part of their standard clinical practice. Lung ultrasound was performed by an independent researcher at the time of chest X-ray. We compared lung ultrasound with chest X-ray for the detection of postoperative pulmonary complications and clinically-relevant postoperative pulmonary complications. We also assessed inter-observer agreement for lung ultrasound, and the time to perform both imaging techniques. Subgroup analyses were performed to compare the time to detection of clinically-relevant postoperative pulmonary complications by both modalities. We recruited a total of 177 patients in whom both lung ultrasound and chest X-ray imaging were performed. Lung ultrasound identified 159 (90%) postoperative pulmonary complications on the day of admission compared with 107 (61%) identified with chest X-ray (p < 0.001). Lung ultrasound identified 11 out of 17 patients (65%) and chest X-ray 7 out of 17 patients (41%) with clinically-relevant postoperative pulmonary complications (p < 0.001). The clinically-relevant postoperative pulmonary complications were detected earlier using lung ultrasound compared with chest X-ray (p = 0.024). Overall inter-observer agreement for lung ultrasound was excellent (κ = 0.907, p < 0.001). Following cardiothoracic surgery, lung ultrasound detected more postoperative pulmonary complications and clinically-relevant postoperative pulmonary complications than chest X-ray, and at an earlier time-point. Our results suggest lung ultrasound may be used as the primary imaging technique to search for postoperative pulmonary complications after cardiothoracic surgery, and will enhance bedside decision making.

Detection of azole-susceptible and azole-resistant Aspergillus coinfection by cyp51A PCR amplicon melting curve analysis.

BACKGROUND: The AsperGenius® assay is a multiplex real-time PCR test that allows the simultaneous detection of Aspergillus species and identification of the most common mutations in the Aspergillus fumigatus cyp51A gene conferring resistance (TR34/L98H and TR46/T289A/Y121F) by using melting curve analysis. Mixed infections with azole-resistant and susceptible A. fumigatus have rarely been described. METHODS: The AsperGenius® multiplex real-time PCR assay (PathoNostics, Maastricht, the Netherlands) was used on bronchoalveolar lavage (BAL) samples of 91 consecutive patients with a suspected invasive Aspergillus infection at the Erasmus MC University Medical Center, Rotterdam. RESULTS: In three cases the AsperGenius® assay indicated the simultaneous presence of WT and mutant genes (two patients with TR34/L98H mutation and one patient with TR46/T289A/Y121F mutation) and therefore mixed infections with azole-susceptible and -resistant isolates. In one of the three cases, the mixed infection was confirmed by phenotypic antifungal testing of multiple A. fumigatus colonies. CONCLUSIONS: The use of a dedicated A. fumigatus cyp51A resistance PCR allowed the detection of mixed infections with azole-resistant and -susceptible Aspergillus strains. These mixed infections may remain undiagnosed with conventional phenotypic susceptibility testing.

Individual Differences in Heparin Sensitivity and Their Effect on Heparin Anticoagulation During Arterial Vascular Surgery.

OBJECTIVES: To investigate whether a fixed heparin dose results in adequate heparinisation levels and consequent inhibition of haemostatic activation in all patients. METHODS: This prospective clinical pilot study included 24 patients undergoing arterial vascular surgery. Individual heparin responsiveness was assessed using the Heparin Dose Response (HDR) test, while the activated clotting time (ACT) and heparin concentration were measured to monitor the peri-procedural degree of anticoagulation. Finally, peri-operative haemostasis was evaluated with rotational thromboelastometry (ROTEM). RESULTS: Eight patients were identified with reduced heparin sensitivity (RS group) and 16 patients with normal heparin sensitivity (NS group). Compared with the NS group, the RS group showed less prolonged ACTs after heparinisation with heparin concentrations below the calculated target heparin concentration. ROTEM revealed shorter clot formation times in the intrinsically activated coagulation test (INTEM) 3 min (114 ± 48 s vs. 210 ± 128 s) and 30 min after the initial heparin bolus (103 ± 48 s vs. 173 ± 81 s) in the RS group compared with the NS group. In the RS group, one patient developed a major thromboembolic complication. CONCLUSIONS: This study shows that a third of the study population had reduced heparin sensitivity, which was associated with lower levels of heparinisation, and lower inhibition levels of clot initiation and clot formation. Identifying patients with reduced heparin sensitivity by monitoring the anticoagulant effect of heparin could decrease the risk of thrombotic complications after arterial vascular surgery.

Benchmarking the use of blood products in cardiac surgery to stimulate awareness of transfusion behaviour : Results from a four-year longitudinal study.

INTRODUCTION: Cardiac operations account for a large proportion of the blood transfusions given each year, leading to high costs and an increased risk to patient safety. Therefore, it is important to explore initiatives to reduce transfusion rates. This study aims to provide a benchmark for transfusion practice by inter-hospital comparison of transfusion rates, blood product use and costs related to patients undergoing coronary artery bypass grafting (CABG), valve surgery or combined CABG and valve surgery. METHODS: Between 2010 and 2013, patients from four Dutch hospitals undergoing CABG, valve surgery or combined CABG and valve surgery (n = 11,150) were included by means of a retrospective longitudinal study design. RESULTS: In CABG surgery the transfusion rate ranged between 43 and 54%, in valve surgery between 54 and 67%, and in combined CABG and valve surgery between 80 and 88%. With the exception of one hospital, the trend in transfusion rate showed a significant decrease over time for all procedures. Hospitals differed significantly in the units of blood products given to each patient, and in the use of specific transfused combinations of blood products, such as red blood cells (RBCs) and a combination of RBCs, fresh frozen plasma (FFP) and platelets. CONCLUSION: This study indicates that benchmarking blood product usage stimulates awareness of transfusion behaviour, which may lead to better patient safety and lower costs. Further studies are warranted to improve awareness of transfusion behaviour and increase the standardisation of transfusion practice in cardiac surgery.

Interspecies discrimination of A. fumigatus and siblings A. lentulus and A. felis of the Aspergillus section Fumigati using the AsperGenius® assay.

The AsperGenius® assay detects several Aspergillus species and the A. fumigatus Cyp51A mutations TR34/L98H/T289A/Y121F that are associated with azole resistance. We evaluated its contribution in identifying A. lentulus and A. felis, 2 rare but intrinsically azole-resistant sibling species within the Aspergillus section Fumigati. Identification of these species with conventional culture techniques is difficult and time-consuming. The assay was tested on (i) 2 A. lentulus and A. felis strains obtained from biopsy proven invasive aspergillosis and (ii) control A. fumigatus (n=3), A. lentulus (n=6) and A. felis species complex (n=12) strains. The AsperGenius® resistance PCR did not detect the TR34 target in A. lentulus and A. felis in contrast to A. fumigatus. Melting peaks for L98H and Y121F markers differed and those of the Y121F marker were particularly suitable to discriminate the 3 species. In conclusion, the assay can be used to rapidly discriminate A. fumigatus, A. lentulus and A. felis.

Validation of a point-of-care prothrombin time test after cardiopulmonary bypass in cardiac surgery.

Point-of-care coagulation monitoring can be used for the guidance of haemostasis management. However, the influence of time on point-of-care prothrombin time testing following protamine administration after cardiopulmonary bypass has not been investigated. Bland-Altman and error grid analysis were used to analyse the level of agreement between prothrombin time measurements from point-of-care and laboratory tests before cardiopulmonary bypass, and then 3 min, 6 min and 10 min after protamine administration. Prothrombin times were expressed as International Normalised Ratios. While the point-of-care and laboratory prothrombin time measurements showed a high level of agreement before bypass, this agreement deteriorated following protamine administration to a mean (SD) bias of -0.22 (0.13) [limits of agreement 0.48-0.04]. Error grid analysis revealed that 35 (70%) of the paired values showed a clinically relevant discrepancy in international normalised ratio. At 3 min, 6 min and 10 min after cardiopulmonary bypass there is a clinical unacceptable discrepancy between the point-of-care and laboratory measurement of prothrombin time.

PCR-based detection of Aspergillus fumigatus Cyp51A mutations on bronchoalveolar lavage: a multicentre validation of the AsperGenius assay® in 201 patients with haematological disease suspected for invasive aspergillosis.

OBJECTIVES: In patients with invasive aspergillosis (IA), fungal cultures are mostly negative. Consequently, azole resistance often remains undetected. The AsperGenius® multiplex real-time PCR assay identifies clinically relevant Aspergillus species and four resistance-associated mutations (RAMs; TR34/L98H/T289A/Y121F) in the Cyp51A gene. This multicentre study evaluated the diagnostic performance of this assay on bronchoalveolar lavage (BAL) fluid and correlated the presence of RAMs with azole treatment failure and mortality. METHODS: Stored BAL samples from patients with haematological diseases with suspected IA were used. BAL samples that were galactomannan/culture positive were considered positive controls for the presence of Aspergillus. Azole treatment failure and 6 week mortality were compared in patients with and without RAMs that had received ≥5 days of voriconazole monotherapy. RESULTS: Two hundred and one patients each contributed one BAL sample, of which 88 were positive controls and 113 were negative controls. The optimal cycle threshold cut-off value for the Aspergillus species PCR was <38. With this cut-off, the PCR was positive in 74/88 positive controls. The sensitivity, specificity, positive predictive value and negative predictive value were 84%, 80%, 76% and 87%, respectively. 32/74 BAL samples were culture negative. Azole treatment failure was observed in 6/8 patients with a RAM compared with 12/45 patients without RAMs (P = 0.01). Six week mortality was 2.7 times higher in patients with RAMs (50.0% versus 18.6%; P = 0.07). CONCLUSIONS: The AsperGenius® assay had a good diagnostic performance on BAL and differentiated WT from Aspergillus fumigatus with RAMs, including in culture-negative BAL samples. Most importantly, detection of RAMs was associated with azole treatment failure.

Endothelial hyperpermeability after cardiac surgery with cardiopulmonary bypass as assessed using an in vitro bioassay for endothelial barrier function.

BACKGROUND: The mechanisms causing increased endothelial permeability after cardiopulmonary bypass (CPB) have not been elucidated. Using a bioassay for endothelial barrier function, we investigated whether endothelial hyperpermeability is associated with alterations in plasma endothelial activation and adhesion markers and can be attenuated by the use of pulsatile flow during CPB. METHODS: Patients undergoing cardiac surgery were randomized to non-pulsatile (n=20) or pulsatile flow CPB (n=20). Plasma samples were obtained before (pre-CPB) and after CPB (post-CPB), and upon intensive care unit (ICU) arrival. Changes in plasma endothelial activation and adhesion markers were determined by enzyme-linked immunosorbent assay. Using electric cell-substrate impedance sensing of human umbilical vein endothelial monolayers, the effects of plasma exposure on endothelial barrier function were assessed and expressed as resistance. RESULTS: Cardiopulmonary bypass was associated with increased P-selectin, vascular cell adhesion molecule-1, and von Willebrand factor plasma concentrations and an increase in the angiopoietin-2 to angiopoietin-1 ratio, irrespective of the flow profile. Plasma samples obtained after CPB induced loss of endothelial resistance of 21 and 23% in non-pulsatile and pulsatile flow groups, respectively. The negative effect on endothelial cell barrier function was still present with exposure to plasma obtained upon ICU admission. The reduction in endothelial resistance after exposure to post-CPB plasma could not be explained by CPB-induced haemodilution. CONCLUSION: The change in the plasma fingerprint during CPB is associated with impairment of in vitro endothelial barrier function, which occurs irrespective of the application of a protective pulsatile flow profile during CPB. CLINICAL TRIAL REGISTRATION: NTR2940.

Limited premature termination codon suppression by read-through agents in cystic fibrosis intestinal organoids.

Premature termination codon read-through drugs offer opportunities for treatment of multiple rare genetic diseases including cystic fibrosis. We here analyzed the read-through efficacy of PTC124 and G418 using human cystic fibrosis intestinal organoids (E60X/4015delATTT, E60X/F508del, G542X/F508del, R1162X/F508del, W1282X/F508del and F508del/F508del). G418-mediated read-through induced only limited CFTR function, but functional restoration of CFTR by PTC124 could not be confirmed. These studies suggest that better read-through agents are needed for robust treatment of nonsense mutations in cystic fibrosis.

Trends in incidence of infective endocarditis at the Medical Center of Alkmaar.

INTRODUCTION: Infective endocarditis (IE) is a life-threatening illness with a high morbidity and mortality, and with a rise in incidence in patients with prosthetic valves and cardiac devices. Recently the Dutch guidelines of IE prophylaxis have been revised, limiting IE prophylaxis to the highest-risk population. The aim of the present study was to investigate the incidence of IE and its trend between 2008-2013 in a regional hospital in the Netherlands. METHODS: This is an observational descriptive study of all patients who were admitted with IE to the Medical Center of Alkmaar (MCA) from 1 January 2008 to 31 December 2013. RESULTS: A total of 89 patients with IE, including 7 patients (7.9 %) with a cardiac device IE (CDIE), were identified. In 2008 there were 8 patients with IE, this increased to 26 patients in 2013. Patients with a prosthetic valve IE increased from 25 % in 2008 to 34.6 % in 2013. This increase was not seen in patients with CDIE. CONCLUSION: In the MCA we have observed an increase in patients with IE since 2010. This increase was in part attributable to prosthetic valve IE. A larger observational study is needed to investigate the increase of IE in the Netherlands.