Purinergic Regulation of Endothelial Barrier Function.

Increased vascular permeability is a hallmark of several cardiovascular anomalies, including ischaemia/reperfusion injury and inflammation. During both ischaemia/reperfusion and inflammation, massive amounts of various nucleotides, particularly adenosine 5'-triphosphate (ATP) and adenosine, are released that can induce a plethora of signalling pathways via activation of several purinergic receptors and may affect endothelial barrier properties. The nature of the effects on endothelial barrier function may depend on the prevalence and type of purinergic receptors activated in a particular tissue. In this review, we discuss the influence of the activation of various purinergic receptors and downstream signalling pathways on vascular permeability during pathological conditions.

Coronary Artery Disease as a Relevant Risk Factor in Screening of Abdominal Aortic Ectasia and Aneurysm.

BACKGROUND: The aim of this study was to investigate the prevalence of abdominal aortic aneurysm (AAA) and abdominal aortic ectasia (AAE) in coronary artery disease (CAD) patients in a multicenter setting to obtain significant data to establish an AAA screening program in our departments. METHODS: Between January and September 2016, 500 patients with suspected or diagnosed CAD planned for coronary angiography or coronary artery bypass graft (CABG) underwent a sonographic examination of the infrarenal abdominal aorta to diagnose AAA or AAE. We calculated the prevalence of AAA and AAE in patients diagnosed of CAD and investigated factors potentially associated with the occurrence of AAA. RESULTS: The overall prevalence in all grades of CAD for AAE was 35.1% and for AAA 5.4%. In patients with three-vessel CAD, the prevalence of AAE was 34% and of AAA 6.8%. Significant correlation was found between the three-vessel CAD and AAA (p = 0.039). The logistic regression analysis showed significant correlation between AAA and age > 65 years (p = 0.05). The multivariate analysis of risk factors and CAD revealed significant correlations between one-vessel CAD and arterial hypertension (AH) (p = 0.004) and age > 65 years (p = 0.001) as well as between three-vessel CAD and AH (p = 0.01), peripheral artery disease (p = 0.01), and age > 65 years (p = 0.03). CONCLUSION: Our results confirm, that in comparison to other data, the prevalence of AAA in patients with CAD is high. Thus, it is recommended to include patients with CAD, especially elderly patients with three-vessel CAD, in future AAA screening programs.

Venous access closure using a purse-string suture without heparin antagonism or additional compression after MitraClip implantation.

OBJECTIVES: This observational study was designed to analyze the safety and feasibility of percutaneous skin closure using a purse-string suture (PSS) after MitraClip procedures. METHODS: Forty-one consecutive patients with severe mitral regurgitation who underwent MitraClip implantation from February 2018 to January 2019 at our institution received a PSS after percutaneous mitral valve repair before withdrawal of the 24-French (Fr) sheath. Protamine was not administered after venous closure at procedure end. No compression therapy (e.g., compression bandage or pneumatic compression device) was used. Patients were on bed rest for 6 hrs prior to suture removal, which was accomplished 18-24 hrs after MitraClip implantation. We analyzed the occurrence of any vascular or thromboembolic complications during the hospital stay and until the 3-month follow-up. RESULTS: The primary endpoint-any access-related major complication-did not occur in any patients. None of the patients revealed a pseudoaneurysm or an arteriovenous fistula, a thromboembolic complication, or local stenosis related to the PSS closure. The secondary endpoint- minor access-site vascular complications (hematoma)- was documented in six (14.6%) patients. CONCLUSIONS: Venous access-site closure with a PSS without the need for protamine administration or compression therapy appears to be safe and feasible in patients undergoing MitraClip implantation with access via a 24-Fr sheath.

Inhibition of Protein Prenylation of GTPases Alters Endothelial Barrier Function.

The members of Rho family of GTPases, RhoA and Rac1 regulate endothelial cytoskeleton dynamics and hence barrier integrity. The spatial activities of these GTPases are regulated by post-translational prenylation. In the present study, we investigated the effect of prenylation inhibition on the endothelial cytoskeleton and barrier properties. The study was carried out in human umbilical vein endothelial cells (HUVEC) and protein prenylation is manipulated with various pharmacological inhibitors. Inhibition of either complete prenylation using statins or specifically geranylgeranylation but not farnesylation has a biphasic effect on HUVEC cytoskeleton and permeability. Short-term treatment inhibits the spatial activity of RhoA/Rho kinase (Rock) to actin cytoskeleton resulting in adherens junctions (AJ) stabilization and ameliorates thrombin-induced barrier disruption whereas long-term inhibition results in collapse of endothelial cytoskeleton leading to increased basal permeability. These effects are reversed by supplementing the cells with geranylgeranyl but not farnesyl pyrophosphate. Moreover, long-term inhibition of protein prenylation results in basal hyper activation of RhoA/Rock signaling that is antagonized by a specific Rock inhibitor or an activation of cAMP signaling. In conclusion, inhibition of geranylgeranylation in endothelial cells (ECs) exerts biphasic effect on endothelial barrier properties. Short-term inhibition stabilizes AJs and hence barrier function whereas long-term treatment results in disruption of barrier properties.

Role of PI3K/Akt and MEK/ERK Signalling in cAMP/Epac-Mediated Endothelial Barrier Stabilisation.

BACKGROUND AND AIMS: Activation of the cAMP/Epac signalling stabilises endothelial barrier function. Moreover, its activation is accompanied by an activation of PI3K/Akt and MEK/ERK signalling in diverse cell types but their impact on endothelial barrier function is largely unknown. Here the role of PI3K/Akt and MEK/ERK signalling in cAMP/Epac-mediated endothelial barrier stabilisation was analysed. METHODS: Endothelial barrier function was analysed in cultured human umbilical vein endothelial cells (HUVECs) by measuring flux of albumin. A modified cAMP analogue 8-pCPT-2'-O-Me-cAMP (Epac agonist) was used to specifically activate cAMP/Epac signalling. RESULTS: Epac agonist reduces the basal and attenuates thrombin-induced endothelial hyperpermeability accompanied by an activation of PI3K/Akt and MEK/ERK signalling. The qPCR data demonstrate HUVECs express PI3Kα, PI3Kß, and PI3Kγ but not PI3Kδ isoforms. The western blot data demonstrate Epac agonist activates PI3Kα and PI3Kß isoforms. Inhibition of MEK/ERK but not PI3K/Akt pathway potentiates the endothelial barrier protective effects of cAMP/Epac signalling. Inhibition of MEK/ERK signalling in the presence of Epac agonist induces a reorganisation of actin cytoskeleton to the cell periphery, enhanced VE-cadherin localisation at cell-cell junctions, and dephosphorylation of myosin light chains (MLC) but not inhibition of RhoA/Rock signalling. Moreover, Epac agonist promotes endothelial cell (EC) survival via reduction in activities of pro-apoptotic caspases in a PI3K/Akt and MEK/ERK signalling-dependent manner. CONCLUSION: Our data demonstrate that the Epac agonist simultaneously activates diverse signalling pathways in ECs, which may have differential effects on endothelial barrier function. It activates PI3K/Akt and MEK/ERK signalling which mainly govern its pro-survival effects on ECs. Inhibition of MEK/ERK but not PI3K/Akt signalling enhances barrier stabilising and barrier protective effects of cAMP/Epac activation. CHEMICAL COMPOUNDS USED IN THIS STUDY: 8-pCPT-2'-O-Me-cAMP (PubChem CID: 9913268); Akt inhibitor VIII (PubChem CID: 10196499); AS-252424 (PubChem CID: 11630874); IC-87114 (PubChem CID: 9908783); PD 98059 (PubChem CID: 4713); PIK-75 (PubChem CID: 10275789); TGX-221 (PubChem CID: 9907093); Thrombin (PubChem CID: 90470996); U0126 (PubChem CID: 3006531); Wortmannin (PubChem CID: 312145).

Assessment of Cardiac Involvement in Fabry Disease (FD) with Native T1 Mapping.

PURPOSE: Fabry disease (FD) is an X-linked multi-organ disorder of lysosomal metabolism with cardiac disease being the leading cause of death. Identifying early FD-specific pathologies is important in the context of maximum therapeutic benefit in these stages. Therefore, the aim of this study was to investigate the value of quantitative cardiac T1 mapping as a potential disease-specific surrogate. METHODS: 16 consecutive FD patients (9 female, 7 male; median age: 54 years, IQR 17) and 16 control patients (9 female, 7 male; median age: 52 years, IQR 20) were investigated at 1.5 Tesla. Native T1 mapping was performed using a modified look locker inversion recovery sequence (MOLLI) and native T1 times were measured within the septal myocardium at the midventricular short-axis section. Also functional parameters, left ventricular morphology, presence of late-gadolinium enhancement, cTnI- and Lyso-Gb3-Levels were evaluated. RESULTS: The median native septal T1 time for FD was 889.0 ms and 950.6 ms for controls (p < 0.003). LGE and positive cTnI values (0.26 ±â€Š0.21) were present in 5 FD patients (31.25 %), and left ventricular hypertrophy (LVH) was present in 4 FD patients (25.00 %). The 4 cTnI and 8 Lyso-Gb3 positive FD patients had significantly lower native T1 values (p < 0.05, respectively p < 0.01). Assuming a T1 cut-off value of 900 ms for the identification of increased cardiac lipid deposit, 9 patients with FD (56.25 %) had pathologic values (4 patients cTnI and 8 patients Lyso-Gb3 positive). Moreover, native septal T1 showed a good negative correlation to Lyso-Gb3 (r = - 0.582; p = 0.018). CONCLUSION: A pathologic cardiac native T1 time obviously reflects cardiac involvement in the scope of FD at tissue level. In the future native T1 mapping as an imaging biomarker might allow identification of early stages of cardiac involvement in FD before morphological changes are obvious. KEY POINTS: · Native T1 values are significantly decreased in Fabry disease.. · Native T1 shows promising correlation to cardiac and Fabry-specific biomarkers.. · Native T1 mapping might have great potential for early disease detection and therapy monitoring.. CITATION FORMAT: · Roller FC, Fuest S, Meyer M et al. Assessment of Cardiac Involvement in Fabry Disease (FD) with Native T1 Mapping. Fortschr Röntgenstr 2019; 191: 932 - 939.

Inhibition of Cx43 attenuates ERK1/2 activation, enhances the expression of Cav­1 and suppresses cell proliferation.

In addition to being an important component of the gap junction, connexin 43 (Cx43) has been shown to regulate other cellular functions, including cell proliferation. This regulatory role of Cx43 may be important in therapeutic situations, including wound healing or ischemic injuries. Caveolin­1 (Cav­1) has been shown to regulate angiogenesis. The aim of the present study was to analyze whether Cx43 counter­regulates Cav­1 in controlling the proliferation and migration of endothelial cells. The inhibition of Cx43 with niflumic acid, flufenamic acid and 18­α­glycyrrhetinic acid in cultured human umbilical vein endothelial cells resulted in decreased phosphorylation of extracellular signal­regulated kinase (ERK)1/2 and increased expression of Cav­1, as shown by western blot analysis. Furthermore, the inhibition of Cx43 resulted in a 50±7% decrease in cell proliferation, determined using a crystal violet assay, a 48±5% decrease in migration, determined using a migration assay, and a 49±6% decrease in endothelial tube formation, determined using a Matrigel assay, compared with the control. Similar results were obtained following specific inhibition of Cx43 by mimetic peptides (Gap26 and Gap27). Inhibition of the mitogen­activated protein kinase kinase/ERK pathway with PD­98059 resulted in an increased expression of Cav­1 and a reduction in the expression of Cx43. Furthermore, cell proliferation, migration and tube formation in endothelial cells were impaired. By contrast, downregulation of the protein expression of Cav­1 by small interference RNA resulted in increased expression of Cx43 and phosphorylation of ERK1/2. Accordingly, the number of cells in the Cav­1 treated­group increased by 35±5% compared with the controls. The data of the present study showed that Cav­1 suppressed cell proliferation by inhibiting the activity of Cx43, which is upstream of ERK1/2. The downregulation of Cav­1 protein resulted in loss of the inhibitory activity of Cav­1 on cell proliferation and led to increased cell proliferation. This counter­regulatory effect of Cx43 may be of importance in therapeutic angiogenesis.

Simultaneous Isolation of High Quality Cardiomyocytes, Endothelial Cells, and Fibroblasts from an Adult Rat Heart.

The rat is an important animal model used in cardiovascular research, and rat cardiac cells are used routinely for in vitro analysis of the molecular mechanisms of cardiovascular disease progression such as cardiac hypertrophy, fibrosis, and atherosclerosis. Although several attempts with variable success have been made to develop immortalized cell lines from the cardiovascular system to understand these cellular mechanisms, primary cells offer a more natural and close to in vivo environment for such studies. Therefore, different laboratories working on a particular cell type have developed protocols to isolate individual types of rat cardiac cells of interest. A protocol that allows the isolation of more than one cell type, however, is missing. Here an optimized protocol is described that allows the isolation of high-quality major cardiac cell types (cardiomyocytes, endothelial cells, and fibroblasts) from a single preparation and enables their use for cellular analyses. This permits the most efficient use of available resources, which may save time and reduce research costs.

Targeting of Extracellular RNA Reduces Edema Formation and Infarct Size and Improves Survival After Myocardial Infarction in Mice.

BACKGROUND: Following myocardial infarction (MI), peri-infarct myocardial edema formation further impairs cardiac function. Extracellular RNA (eRNA) released from injured cells strongly increases vascular permeability. This study aimed to assess the role of eRNA in MI-induced cardiac edema formation, infarct size, cardiac function, and survival after acute MI and to evaluate the therapeutic potential of ribonuclease 1 (RNase-1) treatment as an eRNA-degrading intervention. METHODS AND RESULTS: C57BL/6J mice were subjected to MI by permanent ligation of the left anterior descending coronary artery. Plasma eRNA levels were significantly increased compared with those in controls starting from 30 minutes after ligation. Systemic application of RNase-1, but not DNase, significantly reduced myocardial edema formation 24 hours after ligation compared with controls. Consequently, eRNA degradation by RNase-1 significantly improved the perfusion of collateral arteries in the border zone of the infarcted myocardium 24 hours after ligation of the left anterior descending coronary artery, as detected by micro-computed tomography imaging. Although there was no significant difference in the area at risk, the area of vital myocardium was markedly larger in mice treated with RNase-1 compared with controls, as detected by Evans blue and 2,3,5-triphenyltetrazolium chloride staining. The increase in viable myocardium was associated with significantly preserved left ventricular function, as assessed by echocardiography. Moreover, RNase-1 significantly improved 8-week survival following MI. CONCLUSIONS: eRNA is an unrecognized permeability factor in vivo, associated with myocardial edema formation after acute MI. RNase-1 counteracts eRNA-induced edema formation and preserves perfusion of the infarction border zone, reducing infarct size and protecting cardiac function after MI.

Uridine Triphosphate Thio Analogues Inhibit Platelet P2Y12 Receptor and Aggregation.

Platelet P2Y12 is an important adenosine diphosphate (ADP) receptor that is involved in agonist-induced platelet aggregation and is a valuable target for the development of anti-platelet drugs. Here we characterise the effects of thio analogues of uridine triphosphate (UTP) on ADP-induced platelet aggregation. Using human platelet-rich plasma, we demonstrate that UTP inhibits P2Y12 but not P2Y1 receptors and antagonises 10 µM ADP-induced platelet aggregation in a concentration-dependent manner with an IC50 value of ~250 °µM. An eight-fold higher platelet inhibitory activity was observed with a 2-thio analogue of UTP (2S-UTP), with an IC50 of 30 µM. The 4-thio analogue (4S-UTP) with an IC50 of 7.5 µM was 33-fold more effective. A three-fold decrease in inhibitory activity, however, was observed by introducing an isobutyl group at the 4S- position. A complete loss of inhibition was observed with thio-modification of the γ phosphate of the sugar moiety, which yields an enzymatically stable analogue. The interaction of UTP analogues with P2Y12 receptor was verified by P2Y12 receptor binding and cyclic AMP (cAMP) assays. These novel data demonstrate for the first time that 2- and 4-thio analogues of UTP are potent P2Y12 receptor antagonists that may be useful for therapeutic intervention.

Effect of ticagrelor on endothelial calcium signalling and barrier function.

The P2Y12 receptor is a Gi-coupled receptor whose activation inhibits adenylyl cyclase and thereby reduces the concentration of intracellular cAMP. Here the hypothesis was tested whether AR-C 66096 or ticagrelor, two direct-acting and reversibly binding P2Y12 receptor antagonists, protect endothelial cell (EC) barrier function by raising intracellular cAMP in ECs. The study was carried out on primary human umbilical vein ECs (HUVECs) and human pulmonary microvascular ECs (hPMECs). AR-C66096 (10 µM) induced a 50 % increase in cAMP in ECs whereas ticagrelor (2-10 µM) had no effect. Likewise, AR-C666096 antagonised thrombin-induced hyperpermeability in both HUVECs and hPMECs, but ticagrelor had no effect on basal EC monolayer permeability. Ticagrelor, however, sensitised ECs for thrombin-induced hyperpermeability and potentiated the thrombin effect. Ticagrelor but not AR-C66096 caused an increase in cytosolic calcium ([Ca2+]i). This increase in [Ca2+]i was abrogated by LaCl3 (Ca2+ influx inhibitor) but not by xestospongin C (IP3 receptor antagonist) or by depletion of intracellular stores with thapsigargin, suggesting a Ca2+ influx from the extracellular space. Accordingly, ticagrelor caused an increase in myosin light chain (MLC) phosphorylation, an important regulator of EC contractile machinery and thus permeability, which was abrogated by LaCl3. The ability of ticagrelor to potentiate EC permeability was abrogated by a MLC kinase inhibitor (ML-7; 10 µM). Our data demonstrate that the P2Y12 receptor antagonist AR-C66096 exerts a protective effect on ECs in vitro, possibly by raising intracellular cAMP, whereas ticagrelor sensitises EC barrier function by inducing Ca2+ influx and activating downstream EC contractile machinery.

Diagnosing moyamoya syndrome using ultrasound - a case report.

BACKGROUND: Moyamoya syndrome is a vasculopathy characterised by progressive occlusion of the cerebral arteries resulting in the development of abnormal collateral circulation. To diagnose this syndrome, imaging of the cerebral arteries is required including CT- or MR-angiography and conventional angiography. We present a case of moyamoya disease with typical findings detected in the sonography. The diagnosis was suspected after reviewing the initial ultrasound images of the cerebral arteries with evidence for obliterated intracranial arteries and the detection of an existing collateral circulation network. CASE PRESENTATION: A 62 years old male patient presented in the hospital's emergency department with symptoms indicating a subacute cerebrovascular event. Immediate sonographic studies showed a right-sided pulsatile Doppler-signal in the common and internal carotid arteries, suggestive of distal stenoses. In addition, the transcranial examination indicated obliteration of both middle cerebral arteries. Numerous arterial vessels suggestive of leptomeningeal collateral arteries revealed a strong arterial leptomeningeal flow. At this stage of the diagnostic work-up, the collateral circulation network, characteristic of moyamoya disease, was indicated by sonography. Moyamoya syndrome was verified by conventional angiography. The aetiological work remained empty, so the diagnosis of moyamoya disease was established. CONCLUSION: Our case report indicates that sonography can be a useful tool for detecting the vaculopathy in moyamoya syndrome. In case routine procedures, such as the CT- or MR-angiography, with evidence for obliterated intracerebral arteries, ultrasound studies might provide important information regarding an existing collateral network in the scope of a moyamoya syndrome.

Compound C inhibits in vitro angiogenesis and ameliorates thrombin-induced endothelial barrier failure.

Compound C (comp. C) is a cell-permeable pyrrazolopyrimidine derivative and widely used as adenosine monophosphate-activated protein kinase (AMPK) inhibitor to characterise the role of AMPK in various physiological processes. However, its AMPK-independent effects have also been reported. In the present study we investigated the effects of moderate dose (1-10µM) comp. C on endothelial cell (EC) proliferation, in vitro angiogenesis, and endothelial barrier function. Comp. C was unable to inhibit AMPK phosphorylation (activation) induced by metformin and A-769662 in ECs even at concentration of 10µM. At lower concentration (1µM), comp. C inhibited and potentiated the inhibitory effects of metformin and A-769662 on EC proliferation, migration, tube formation, and sprouting without inducing apoptosis. However, at higher concentration (10µM), it strongly induced apoptosis as measured by enhanced caspase 3/7 activity. Moreover, comp. C antagonised thrombin-induced EC hyperpermeability accompanied by activation of Rac1 and strengthening of adherens junctions (AJs). This EC barrier protective effect was not affected by the presence of AMPK activators. The data of the present study demonstrate that long-term treatment of ECs with low concentration comp. C inhibits EC proliferation and angiogenesis without induction of apoptosis. While short-term incubation antagonises thrombin-induced EC hyperpermeability presumably via Rac1-dependent strengthening of AJs. Furthermore, higher concentration of comp. C (10µM or above) is toxic for ECs and warns that this agent should be used with caution to demonstrate the AMPK-mediated effects.

Cleaved high-molecular-weight kininogen inhibits neointima formation following vascular injury.

Cleaved high-molecular-weight kininogen (HKa) or its peptide domain 5 (D5) alone exert anti-adhesive properties in vitro related to impeding integrin-mediated cellular interactions. However, the anti-adhesive effects of HKa in vivo remain elusive. In this study, we investigated the effects of HKa on leukocyte recruitment and neointima formation following wire-induced injury of the femoral artery in C57BL/6 mice. Local application of HKa significantly reduced the accumulation of monocytes and also reduced neointimal lesion size 14 days after injury. Moreover, C57BL/6 mice transplanted with bone marrow from transgenic mice expressing enhanced green fluorescence protein (eGFP) showed a significantly reduced accumulation of eGFP+-cells at the arterial injury site and decreased neointimal lesion size after local application of HKa or the polypeptide D5 alone. A differentiation of accumulating eGFP+-cells into highly specific smooth muscle cells (SMC) was not detected in any group. In contrast, application of HKa significantly reduced the proliferation of locally derived neointimal cells. In vitro, HKa and D5 potently inhibited the adhesion of SMC to vitronectin, thus impairing their proliferation, migration, and survival rates. In conclusion, application of HKa or D5 decreases the inflammatory response to vascular injury and exerts direct effects on SMC by impeding the binding of integrins to extracellular matrix components. Therefore, HKa and D5 may hold promise as novel therapeutic substances to prevent neointima formation.

cAMP controls the restoration of endothelial barrier function after thrombin-induced hyperpermeability via Rac1 activation.

Inflammatory mediators like thrombin disrupt endothelial adherens junctions (AJs) and barrier integrity leading to oedema formation followed by resealing of AJs and a slow recovery of the barrier function. The molecular mechanisms of this process have not yet been fully delineated. The aim of the present study was to analyse the molecular mechanism of endothelial barrier recovery and thrombin was used as model inflammatory mediator. Thrombin caused a strong increase in endothelial permeability within 10 min accompanied by loss of Rac1 but not cdc42 activity, drop in cellular cAMP contents, and a strong activation of the endothelial contractile machinery mainly via RhoA/Rock signalling. Activation of RhoA/Rock signalling precedes and is dependent upon a rise in the cytosolic Ca(2+) concentration. Inhibition of cytosolic Ca(2+) rise but not MLCK or Rock enhances the recovery of endothelial barrier function. The cellular cAMP contents increased gradually during the barrier recovery phase (30-60 min after thrombin challenge) accompanied by an increase in Rac1 activity. Inhibition of Rac1 activity using a specific pharmacological inhibitor (NSC23766) abrogated the endothelial barrier recovery process, suggesting a Rac1-dependent phenomenon. Likewise, inhibition of either adenylyl cyclase or the cAMP-effectors PKA and Epac (with PKI and ESI-09, respectively) caused an abrogation of Rac1 activation, resealing of endothelial AJs and recovery of endothelial barrier function. The data demonstrate that endothelial barrier recovery after thrombin challenge is regulated by Rac1 GTPase activation. This Rac1 activation is due to increased levels of cellular cAMP and activation of downstream signalling during the barrier recovery phase.

Continuous cardiac troponin I release in Fabry disease.

BACKGROUND: Fabry disease (FD) is a rare lysosomal storage disorder also affecting the heart. The aims of this study were to determine the frequency of cardiac troponin I (cTNI) elevation, a sensitive parameter reflecting myocardial damage, in a smaller cohort of FD-patients, and to analyze whether persistent cTNI can be a suitable biomarker to assess cardiac dysfunction in FD. METHODS: cTNI values were determined at least twice per year in 14 FD-patients (6 males and 8 females) regularly followed-up in our centre. The data were related to other parameters of heart function including cardiac magnetic resonance imaging (cMRI). RESULTS: Three patients (21%) without specific vascular risk factors other than FD had persistent cTNI-elevations (range 0.05-0.71 ng/ml, normal: <0.01). cMRI disclosed late gadolinium enhancement (LGE) in all three individuals with cTNI values ≥0.01, while none of the 11 patients with cTNI <0.01 showed a pathological enhancement (p<0.01). Two subjects with increased cTNI-values underwent coronary angiography, excluding relevant stenoses. A myocardial biopsy performed in one during this procedure demonstrated substantial accumulation of globotriaosylceramide (Gb3) in cardiomyocytes. CONCLUSION: Continuous cTNI elevation seems to occur in a substantial proportion of patients with FD. The high accordance with LGE, reflecting cardiac dysfunction, suggests that cTNI-elevation can be a useful laboratory parameter for assessing myocardial damage in FD.