Integrative Multi-omics Analysis of Childhood Aggressive Behavior.

This study introduces and illustrates the potential of an integrated multi-omics approach in investigating the underlying biology of complex traits such as childhood aggressive behavior. In 645 twins (cases = 42%), we trained single- and integrative multi-omics models to identify biomarkers for subclinical aggression and investigated the connections among these biomarkers. Our data comprised transmitted and two non-transmitted polygenic scores (PGSs) for 15 traits, 78,772 CpGs, and 90 metabolites. The single-omics models selected 31 PGSs, 1614 CpGs, and 90 metabolites, and the multi-omics model comprised 44 PGSs, 746 CpGs, and 90 metabolites. The predictive accuracy for these models in the test (N = 277, cases = 42%) and independent clinical data (N = 142, cases = 45%) ranged from 43 to 57%. We observed strong connections between DNA methylation, amino acids, and parental non-transmitted PGSs for ADHD, Autism Spectrum Disorder, intelligence, smoking initiation, and self-reported health. Aggression-related omics traits link to known and novel risk factors, including inflammation, carcinogens, and smoking.

Heritability of Urinary Amines, Organic Acids, and Steroid Hormones in Children.

Variation in metabolite levels reflects individual differences in genetic and environmental factors. Here, we investigated the role of these factors in urinary metabolomics data in children. We examined the effects of sex and age on 86 metabolites, as measured on three metabolomics platforms that target amines, organic acids, and steroid hormones. Next, we estimated their heritability in a twin cohort of 1300 twins (age range: 5.7-12.9 years). We observed associations between age and 50 metabolites and between sex and 21 metabolites. The monozygotic (MZ) and dizygotic (DZ) correlations for the urinary metabolites indicated a role for non-additive genetic factors for 50 amines, 13 organic acids, and 6 steroids. The average broad-sense heritability for these amines, organic acids, and steroids was 0.49 (range: 0.25-0.64), 0.50 (range: 0.33-0.62), and 0.64 (range: 0.43-0.81), respectively. For 6 amines, 7 organic acids, and 4 steroids the twin correlations indicated a role for shared environmental factors and the average narrow-sense heritability was 0.50 (range: 0.37-0.68), 0.50 (range; 0.23-0.61), and 0.47 (range: 0.32-0.70) for these amines, organic acids, and steroids. We conclude that urinary metabolites in children have substantial heritability, with similar estimates for amines and organic acids, and higher estimates for steroid hormones.

The mode of action of sugammadex on coagulation.

OBJECTIVE: To explore the mode of action (MoA) by which sugammadex interferes with coagulation. MATERIALS AND METHODS: The effect of sugammadex on various steps in the coagulation cascade including thrombin generation, factor Xa activity, and factor Xa generation was explored in human plasma. RESULTS: Sugammadex did not affect a conventional thrombin generation test (TGT), while it prolongs activated partial thromboplastin time (APTT) and prothrombin time (PT). However, a customized TGT with PT reagent revealed sugammadex effects. In addition, sugammadex prolonged a one-step prothrombinase-induced clotting time (PiCT) using human factor Xa. Furthermore, sugammadex interfered with factor Xa generation induced by an intrinsic and not by an extrinsic activator, nor by Russell's viper venom factor X (RVV-X). CONCLUSION: Adapted, rather than standard, experiments show that sugammadex is likely to decrease factor Xa activity in the common pathway and activation of factor X specifically in the intrinsic pathway.

Interactions of sugammadex with various anticoagulants
.

OBJECTIVE: To investigate in vitro the effect of sugammadex on activated partial thromboplastin time (APTT) and prothrombin time (PT) prolongations with various anticoagulants as well as the neutralizing effect of rocuronium and vecuronium on sugammadex effects on APTT and PT. MATERIALS AND METHODS: We investigated in vitro the effect of sugammadex on APTT and/or PT in plasma of patients on a vitamin K antagonist and with elevated international normalized ratios (INRs), in plasma of healthy subjects spiked with either a low or high concentration of enoxaparin, fondaparinux, rivaroxaban, and dabigatran, and in perioperatively collected patient plasma. In addition, we explored whether the effects of sugammadex persisted in the presence of rocuronium or vecuronium, or whether they were counteracted by these compounds. RESULTS: Sugammadex concentration-dependently increased APTT and PT(INR) in all anticoagulant conditions, mainly in a proportional manner, with no differences between perioperatively collected patient and control plasma. Rocuronium and vecuronium both neutralized the effects of sugammadex on APTT and PT. CONCLUSION: Sugammadex has a transient effect on coagulation and is unlikely to increase bleeding risk, this possibility cannot be excluded for scenarios not clinically studied.

Urinary Amine and Organic Acid Metabolites Evaluated as Markers for Childhood Aggression: The ACTION Biomarker Study.

Biomarkers are of interest as potential diagnostic and predictive instruments in personalized medicine. We present the first urinary metabolomics biomarker study of childhood aggression. We aim to examine the association of urinary metabolites and neurotransmitter ratios involved in key metabolic and neurotransmitter pathways in a large cohort of twins (N = 1,347) and clinic-referred children (N = 183) with an average age of 9.7 years. This study is part of ACTION (Aggression in Children: Unraveling gene-environment interplay to inform Treatment and InterventiON strategies), in which we developed a standardized protocol for large-scale collection of urine samples in children. Our analytical design consisted of three phases: a discovery phase in twins scoring low or high on aggression (N = 783); a replication phase in twin pairs discordant for aggression (N = 378); and a validation phase in clinical cases and matched twin controls (N = 367). In the discovery phase, 6 biomarkers were significantly associated with childhood aggression, of which the association of O-phosphoserine (ß = 0.36; SE = 0.09; p = 0.004), and gamma-L-glutamyl-L-alanine (ß = 0.32; SE = 0.09; p = 0.01) remained significant after multiple testing. Although non-significant, the directions of effect were congruent between the discovery and replication analyses for six biomarkers and two neurotransmitter ratios and the concentrations of 6 amines differed between low and high aggressive twins. In the validation analyses, the top biomarkers and neurotransmitter ratios, with congruent directions of effect, showed no significant associations with childhood aggression. We find suggestive evidence for associations of childhood aggression with metabolic dysregulation of neurotransmission, oxidative stress, and energy metabolism. Although replication is required, our findings provide starting points to investigate causal and pleiotropic effects of these dysregulations on childhood aggression.

Whole blood assay as a model for in vitro evaluation of inflammasome activation and subsequent caspase-mediated interleukin-1 beta release.

Processing of pro-interleukin (IL)-1ß and IL-18 is regulated by multiprotein complexes, known as inflammasomes. Inflammasome activation results in generation of bioactive IL-1ß and IL-18, which can exert potent pro-inflammatory effects. Our aim was to develop a whole blood-based assay to study the inflammasome in vitro and that also can be used as an assay in clinical studies. We show whole blood is a suitable milieu to study inflammasome activation in primary human monocytes. We demonstrated that unprocessed human blood cells can be stimulated to activate the inflammasome by the addition of adenosine 5'-triphosphate (ATP) within a narrow timeframe following lipopolysaccharide (LPS) priming. Stimulation with LPS resulted in IL-1ß release; however, addition of ATP is necessary for "full-blown" inflammasome stimulation resulting in high IL-1ß and IL-18 release. Intracellular cytokine staining demonstrated monocytes are the major producers of IL-1ß in human whole blood cultures, and this was associated with activation of caspase-1/4/5, as detected by a fluorescently labelled caspase-1/4/5 probe. By applying caspase inhibitors, we show that both the canonical inflammasome pathway (via caspase-1) as well as the non-canonical inflammasome pathway (via caspases-4 and 5) can be studied using this whole blood-based model.

Genome-wide analysis of DNA methylation in buccal cells: a study of monozygotic twins and mQTLs.

BACKGROUND: DNA methylation arrays are widely used in epigenome-wide association studies and methylation quantitative trait locus (mQTL) studies. Here, we performed the first genome-wide analysis of monozygotic (MZ) twin correlations and mQTLs on data obtained with the Illumina MethylationEPIC BeadChip (EPIC array) and compared the performance of the EPIC array to the Illumina HumanMethylation450 BeadChip (HM450 array) for buccal-derived DNA. RESULTS: Good-quality EPIC data were obtained for 102 buccal-derived DNA samples from 49 MZ twin pairs (mean age = 7.5 years, range = 1-10). Differences between MZ twins in the cellular content of buccal swabs were a major driver for differences in their DNA methylation profiles, highlighting the importance to adjust for cellular composition in DNA methylation studies of buccal-derived DNA. After adjusting for cellular composition, the genome-wide mean correlation (r) between MZ twins was 0.21 for the EPIC array, and cis mQTL analysis in 84 twins identified 1,296,323 significant associations (FDR 5%), encompassing 33,749 methylation sites and 616,029 genetic variants. MZ twin correlations were slightly larger (p < 2.2 × 10-16) for novel EPIC probes (N = 383,066, mean r = 0.22) compared to probes that are also present on HM450 (N = 406,822, mean r = 0.20). In line with this observation, a larger percentage of novel EPIC probes was associated with genetic variants (novel EPIC probes with significant mQTL 4.7%, HM450 probes with mQTL 3.9%, p < 2.2 × 10-16). Methylation sites with a large MZ correlation and sites associated with mQTLs were most strongly enriched in epithelial cell DNase I hypersensitive sites (DHSs), enhancers, and histone mark H3K4me3. CONCLUSIONS: We conclude that the contribution of familial factors to individual differences in DNA methylation and the effect of mQTLs are larger for novel EPIC probes, especially those within regulatory elements connected to active regions specific to the investigated tissue.

A Dose-Escalating Study With the Fetal Estrogen Estetrol in Healthy Men.

Context: Luteinizing hormone-releasing hormone (LHRH) agonists have replaced estrogens for endocrine treatment of advanced prostate cancer (PC) because of cardiovascular side effects. The fetal estrogen estetrol (E4) may be safer for PC treatment and is expected to decrease testosterone (T) and prevent estrogen deficiency. Objective: To investigate the safety and T-suppressive effect of E4 in healthy men. Design: Double-blind, randomized, placebo-controlled, dose-escalating study. Setting: The study was conducted at a phase I clinical unit (QPS, Netherlands). Participants: Healthy male volunteers aged 40 to 70 years. Intervention(s): Three treatment cohorts of 15 volunteers with placebo (n = 5) and E4 (n = 10). Estetrol doses tested were 20, 40, and 60 mg/d. Subjects were treated for 4 weeks. Main Outcome Measures: Subjective side effects, pharmacodynamic effects on hemostatic variables, lipids, glucose, bone parameters, and endocrine parameters related to T metabolism. Results: Total and free T decreased dose-dependently and significantly. Nipple tenderness occurred in 40% and decrease of libido occurred in 30% of E4-treated men. The unwanted estrogenic effects on hemostasis were small, dose dependent, and in some cases significant. Lipid and bone parameters showed a favorable trend. Conclusion: The effect of E4 on testosterone levels is insufficient for standalone PC treatment. Taking all clinical and pharmacodynamic variables into consideration, a daily dose of 40 mg E4 seems safe for further evaluation of endocrine PC treatment in combination with LHRH analogs.

The BCR-ABL1 Inhibitors Imatinib and Ponatinib Decrease Plasma Cholesterol and Atherosclerosis, and Nilotinib and Ponatinib Activate Coagulation in a Translational Mouse Model.

Treatment with the second and third generation BCR-ABL1 tyrosine kinase inhibitors (TKIs) increases cardiovascular risk in chronic myeloid leukemia (CML) patients. We investigated the vascular adverse effects of three generations of TKIs in a translational model for atherosclerosis, the APOE*3Leiden.CETP mouse. Mice were treated for sixteen weeks with imatinib (150 mg/kg BID), nilotinib (10 and 30 mg/kg QD) or ponatinib (3 and 10 mg/kg QD), giving similar drug exposures as in CML-patients. Cardiovascular risk factors were analyzed longitudinally, and histopathological analysis of atherosclerosis and transcriptome analysis of the liver was performed. Imatinib and ponatinib decreased plasma cholesterol (imatinib, -69%, p < 0.001; ponatinib 3 mg/kg, -37%, p < 0.001; ponatinib 10 mg/kg-44%, p < 0.001) and atherosclerotic lesion area (imatinib, -78%, p < 0.001; ponatinib 3 mg/kg, -52%, p = 0.002; ponatinib 10 mg/kg, -48%, p = 0.006), which were not affected by nilotinib. In addition, imatinib increased plaque stability. Gene expression and pathway analysis demonstrated that ponatinib enhanced the mRNA expression of coagulation factors of both the contact activation (intrinsic) and tissue factor (extrinsic) pathways. In line with this, ponatinib enhanced plasma levels of FVII, whereas nilotinib increased plasma FVIIa activity. While imatinib showed a beneficial cardiovascular risk profile, nilotinib and ponatinib increased the cardiovascular risk through induction of a pro-thrombotic state.

Thrombin Generation in Vitreous and Subretinal Fluid of Patients with Retinal Detachment.

PURPOSE: To measure prothrombin fragments (F1+2) and thrombin-antithrombin complex (TAT) in vitreous and subretinal fluid (SRF) of rhegmatogenous retinal detachment (RRD) patients and to validate and further specify our earlier finding of increased thrombin activity in patients with proliferative vitreoretinopathy (PVR). METHODS: F1+2 and TAT were measured in 31 vitreous and 16 SRF samples using the Enzygnost® immunoassays. RESULTS: We found significant levels of F1+2 and TAT in the vitreous of all patients with RRD compared to patients with macular hole or macular pucker. However, there was no significant difference between patients who would develop PVR in the future, had established PVR, and patients with uncomplicated RRD both in vitreous concentrations of F1+2 (Kruskal-Wallis p = 0.963) and TAT (p = 0.516). CONCLUSION: The analysis of F1+2 and TAT confirmed significant thrombin generation in both vitreous and SRF of patients with RRD. An imbalance between the thrombin regulation mechanisms TAT and α2-macroglobulin possibly explains the difference from our previous findings.

Uncovering a Predictive Molecular Signature for the Onset of NASH-Related Fibrosis in a Translational NASH Mouse Model.

BACKGROUND & AIMS: The incidence of nonalcoholic steatohepatitis (NASH) is increasing. The pathophysiological mechanisms of NASH and the sequence of events leading to hepatic fibrosis are incompletely understood. The aim of this study was to gain insight into the dynamics of key molecular processes involved in NASH and to rank early markers for hepatic fibrosis. METHODS: A time-course study in low-density lipoprotein-receptor knockout. Leiden mice on a high-fat diet was performed to identify the temporal dynamics of key processes contributing to NASH and fibrosis. An integrative systems biology approach was used to elucidate candidate markers linked to the active fibrosis process by combining transcriptomics, dynamic proteomics, and histopathology. The translational value of these findings were confirmed using human NASH data sets. RESULTS: High-fat-diet feeding resulted in obesity, hyperlipidemia, insulin resistance, and NASH with fibrosis in a time-dependent manner. Temporal dynamics of key molecular processes involved in the development of NASH were identified, including lipid metabolism, inflammation, oxidative stress, and fibrosis. A data-integrative approach enabled identification of the active fibrotic process preceding histopathologic detection using a novel molecular fibrosis signature. Human studies were used to identify overlap of genes and processes and to perform a network biology-based prioritization to rank top candidate markers representing the early manifestation of fibrosis. CONCLUSIONS: An early predictive molecular signature was identified that marked the active profibrotic process before histopathologic fibrosis becomes manifest. Early detection of the onset of NASH and fibrosis enables identification of novel blood-based biomarkers to stratify patients at risk, development of new therapeutics, and help shorten (pre)clinical experimental time frames.

2SNP heritability and effects of genetic variants for neutrophil-to-lymphocyte and platelet-to-lymphocyte ratio.

Neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) are important biomarkers for disease development and progression. To gain insight into the genetic causes of variance in NLR and PLR in the general population, we conducted genome-wide association (GWA) analyses and estimated SNP heritability in a sample of 5901 related healthy Dutch individuals. GWA analyses identified a new genome-wide significant locus on the HBS1L-MYB intergenic region for PLR, which replicated in a sample of 2538 British twins. For platelet count, we replicated three known genome-wide significant loci in our cohort (at CCDC71L-PIK3CG, BAK1 and ARHGEF3). For neutrophil count, we replicated the PSMD3 locus. For the identified top SNPs, we found significant cis and trans expression quantitative trait loci effects for several loci involved in hematological and immunological pathways. Linkage Disequilibrium score (LD) regression analyses for PLR and NLR confirmed that both traits are heritable, with a polygenetic SNP heritability for PLR of 14.1%, and for NLR of 2.4%. Genetic correlations were present between ratios and the constituent counts, with the genetic correlation (r=0.45) of PLR with platelet count reaching statistical significance. In conclusion, we established that two important biomarkers have a significant heritable SNP component, and identified the first genome-wide locus for PLR.

Fibrin clot structure - pro-fibrinolytic effect of oral contraceptives in apparently healthy women.

Fibrin metabolism is influenced by many factors. The velocity of fibrin formation, genetic polymorphisms, fibrinolytic features and the structure of the fibrin clot are determinants of fibrin turnover. Oral contraceptives (OCs) have significant impact on the haemostatic system, by increasing the concentration of coagulation factors, plasminogen and tissue plasminogen activator activity, and decreasing the concentration of haemostatic inhibitors. The present study addresses the influence of OCs on fibrin structure and fibrin metabolism. The study included 70 women treated with seven different OC-formulations. Blood was collected at baseline and after six months of OCs. The plasma concentration of fibrinogen, thrombin-antithrombin complex (TAT), plasminogen, plasmin-antiplasmin complex (PAP), D-Dimer and thrombin generation measures were determined. Fibrin structure measures and fibrin clot lysis not affected by the plasma concentration of plasminogen activators and inhibitors were determined. OCs increased the concentration of fibrinogen, TAT, plasminogen, PAP and D-dimer significantly and affected measures of thrombin generation (p<0.001). The maximal optical density of fibrin (p<0.001), the fibrin fibre density (p=0.03), fibrin fibre diameter (p=0.003), fibrin mass-length ratio (p<0.001) and lysis per hour (p<0.001) increased significantly upon OC-treatment. Lysis per hour was not correlated to the concentration of plasminogen. We conclude that the effect of OCs on the coagulation system is balanced by alterations in fibrin structure, facilitating clot lysis and contributing to the fibrinolytic susceptibility already present in women treated with OC. These alterations may counterbalance the OC-induced increased thrombin generation and reduced coagulation inhibitory potential, contributing to maintenance of the haemostatic balance in women receiving OCs.

Assessing Safety of Thrombolytic Therapy.

Thrombolytic therapy involves thrombolytic agents administered to patients suffering from venous or arterial thrombosis. The therapy induces systemic effects interrelated with the thrombolytic agent used. Bleeding is a prominent complication of thrombolytic therapy. Exhaustion of coagulation factors, generation of excessive amounts of fibrin degradation products (FDPs), therapy-induced activation of coagulation, therapy-induced anticoagulation, and formation of new fibrin all illustrate the complexity of effects of the treatment and challenges the hemostatic balance in the patients. The therapy-induced effects can be modulated by parallel administration of anticoagulants. Risk assessment is mandatory prior to thrombolytic therapy. Anticoagulated and unconscious patients represent particular safety concerns, and should be fully evaluated. Several guidelines describe the choice of tests and their safety limits in relation to pretreatment evaluation of anticoagulated patients. Fibrinogen depletion and FDPs during treatment may be promising markers for the evaluation of bleeding risk posttreatment. Future risk assessment measures should focus on the dynamics of the hemostatic balance. Here, thromboelastography may be considered a tool addressing clot formation, fibrin structure, and fibrinolytic resistance in parallel. Suitable laboratory analysis performed shortly after treatment may help to recognize severe treatment-induced systemic effects that can be counteracted by rational treatment, thereby reducing bleeding risk.

Reduced hemostatic effects with drospirenone-based oral contraceptives containing estetrol vs. ethinyl estradiol.

OBJECTIVE: The effects of estetrol (E4), a natural fetal estrogen, combined with drospirenone (DRSP) were evaluated on plasma levels of sex hormone-binding globulin (SHBG), angiotensinogen and 12 hemostasis markers. STUDY DESIGN: Combinations of 3 mg DRSP with 5 or 10 mg E4 were compared with YAZ® (20 mcg ethinyl estradiol and 3 mg DRSP; EE/DRSP) in parallel groups of 15-18 healthy young women. Main outcome was the relative change from pretreatment to the end (day 24±1) of the third treatment cycle. RESULTS: All E4 combinations showed low estrogen impact compared to EE/DRSP. Effects on SHBG and angiotensinogen of 10 mg E4 combined with DRSP were 15%-20% that of EE/DRSP. Both E4/DRSP combinations reduced D-dimer level and the 5 mg E4/DRSP combination also decreased fragment 1+2. CONCLUSIONS: The reduction in coagulation markers suggests an anticoagulant effect from DRSP. The indications of a low thrombosis risk for E4 preparations should be validated in larger studies. IMPLICATION STATEMENT.

Causes of variation in the neutrophil-lymphocyte and platelet-lymphocyte ratios: a twin-family study.

AIM: Neutrophil-lymphocyte ratio (NLR) and platelet-lymphocyte ratio (PLR) are biomarkers for disease development, for whom little is known about causes of variation in the general population. MATERIALS & METHODS: We estimated the heritability of PLR and NLR and examined their association with gender, demographic, lifestyle and environmental factors in a Dutch nonpatient twin family population (n = 8108). RESULTS: Heritability was estimated at 64% for PLR and 36% for NLR. Men had on average higher NLR, but lower PLR levels than women. PLR and NLR increased significantly with age, decreased in colder months and showed small but significant sex- and age-specific associations with body composition and smoking. CONCLUSION: NLR and PLR levels are heritable and influenced by age, sex and environmental factors, such as seasonal conditions and lifestyle.

Biological Variation of Hemostasis Variables in Thrombosis and Bleeding: Consequences for Performance Specifications.

BACKGROUND: Levels of hemostasis factors vary between and within individuals as a result of genetic and environmental factors and analytical variation of the assays. The current state of the art for defining analytical precision requirements for analytical testing is based on this between- and within-individual (biological) variation. However, information on biological variation in hemostasis variables is still limited.The aim of this study was to determine the biological variation of coagulation variables involved in thrombosis and bleeding to provide a recommendation for performance specifications and to assess whether hemostasis assays fulfill the recommendation. METHODS: We performed a longitudinal study by repeated blood sampling (in total 13 times over a 1-year period) in 40 healthy individuals and measured prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen, antithrombin, factor VIII, factor IX, von Willebrand factor (VWF), protein C, and protein S. We evaluated the effect of the biological variation on parameters of analytical variation and propose required performance specifications. RESULTS: Biological variation was highly different for various hemostasis variables: the within-subject variation ranged from 2.6% (PT) to 25.6% [VWF collagen binding (CB) activity], the between-subject variation varied from 4.1% (PT) to 31.2% (VWF:ristocetin cofactor acitivity), and the assay variation from 1.3% (PT) to 12.9% (VWF:CB). CONCLUSIONS: With the reagents and analyzers used in this study, most of the hemostasis tests variables fulfill the current quality criteria for diagnosis and monitoring of routine hemostasis assays.

Vitreous and subretinal fluid concentrations of orally administered dabigatran in patients with rhegmatogenous retinal detachment.

PURPOSE: One of the factors that was shown to contribute to the development of proliferative vitreoretinopathy (PVR) is the coagulation factor thrombin. Therefore, a specific oral thrombin inhibitor such as dabigatran might be a possible therapeutic option. An oral drug has the advantage of patient-friendly prolonged administration in contrast to drugs that can only be applied during vitrectomy, on condition that the drug reaches the target site. We tested whether dabigatran reaches the vitreous and subretinal fluid (SRF) after a single oral dose of dabigatran. METHODS: Twenty-eight patients with a retinal detachment received a single dose of 220 mg dabigatran etexilate 2-8 hr prior to surgery. During surgery, we took a blood sample and a vitreous or subretinal fluid sample. The concentration of dabigatran was measured using liquid chromatography-tandem mass spectrometry (LC-MS/MS). RESULTS: The dabigatran concentration between 2 and 9 hr after administration was higher in SRF than in vitreous (max 8.5 and 3.8 ng/ml). Corresponding plasma concentrations ranged from 15 to 225 ng/ml. There was a significant relationship between SRF levels and plasma levels (rs  = 0.68, p = 0.014); the levels in vitreous fluid showed no such relationship (rs  = 0.20, p = 0.48). In addition, we measured the vitreous concentration of a non-study patient using 150 mg dabigatran twice daily. The concentration was approximately 10 times higher than after a single dosage (25.8 ng/ml). CONCLUSION: We demonstrate that oral intake of dabigatran, a candidate drug to modulate PVR, results in potentially relevant intraocular concentrations. We suggest that repeated dosing may lead to higher concentrations, but this should be further explored.