Comparison of diverse platelet activation markers as indicators for left atrial thrombus in atrial fibrillation.

Atrial fibrillation (AF) is well known for being a major risk factor of thromboembolic stroke. We could recently demonstrate an association of monocyte-platelet aggregates (MPAs) with the degree of thrombogenicity in patients with AF. This study investigated platelet activation markers, as potential biomarkers for the presence of left atrial (LA) thrombus in patients with AF. One hundred and eight patients with symptomatic AF underwent transesophageal echocardiography (TEE) before scheduled cardioversion or pulmonary vein isolation. In order to determine the content of MPAs by flow-cytometric quantification analyses, blood was drawn on the day of TEE. The soluble CD40 Ligand (sCD40L) and soluble P-selectin (sP-selectin) were obtained by Cytometric Bead Arrays (CBA). D-dimer levels were detected by quantitative immunological determination of fibrin degradation products. Clinical, laboratory, and echocardiographic standard parameters were obtained from all patients, including the determination of the flow in the left atrial appendage (LAA). Patients with detected LA thrombus (n = 28) compared with patients without thrombus (n = 80) showed an increased number of common risk factors, such as age, diabetes, heart failure, and coronary artery disease (CAD). The presence of LA thrombus was associated with significantly increased levels of MPAs (147 ± 12 vs. 304 ± 29 per µl; p < 0.00), sCD40L (106.3 ± 31.0 vs. 33.5 ± 2.1 pg/ml, p = 0.027), and D-dimer (0.13 ± 0.02 vs. 0.69 ± 0.21 mg FEU/l, p = 0.015). In contrast, sP-selectin showed no association with LA thrombus. A multivariate regression analysis showed that MPAs, sCD40L as well as D-dimers were independent indicators for the existence of LA thrombus. MPAs above 170 cells/µl indicated LA thrombus with a high sensitivity of 93% and a specificity of 73% (OR 62, 95% CI. 6.9-557.2, p < 0.001) in patients with AF, whereas the D-dimer lost their quality as independent indicator by using the conventional cut-off of 0.5 mg/l within the regression analysis. MPAs, as well as the D-dimer, correlated significantly negatively with the flow in the LAA measured during TEE. The content of MPAs, sCD40L, and D-dimer, but not sP-selectin showed an increased dependence on LA thrombus in patients with AF. In our study group, MPAs showed the best diagnostic test accuracy of the compared platelet markers. The different results of the examined platelet activation markers could be an indication of diverse mechanisms of LA thrombus in AF. Further studies should evaluate whether determination of MPAs in clinical routine may suffice to indicate the presence of LA thrombus in patients with AF.

TAK1 and IKK2, novel mediators of SCF-induced signaling and potential targets for c-Kit-driven diseases.

NF-κB activation depends on the IKK complex consisting of the catalytically active IKK1 and 2 subunits and the scaffold protein NEMO. Hitherto, IKK2 activation has always been associated with IκBα degradation, NF-κB activation, and cytokine production. In contrast, we found that in SCF-stimulated primary bone marrow-derived mast cells (BMMCs), IKK2 is alternatively activated. Mechanistically, activated TAK1 mediates the association between c-Kit and IKK2 and therefore facilitates the Lyn-dependent IKK2 activation which suffices to mediate mitogenic signaling but, surprisingly, does not result in NF-κB activation. Moreover, the c-Kit-mediated and Lyn-dependent IKK2 activation is targeted by MyD88-dependent pathways leading to enhanced IKK2 activation and therefore to potentiated effector functions. In neoplastic cells, expressing constitutively active c-Kit mutants, activated TAK1 and IKKs do also not induce NF-κB activation but mediate uncontrolled proliferation, resistance to apoptosis and enables IL-33 to mediate c-Kit-dependent signaling. Together, we identified the formation of the c-Kit-Lyn-TAK1 signalosome which mediates IKK2 activation. Unexpectedly, this IKK activation is uncoupled from the NF-κB-machinery but is critical to modulate functional cell responses in primary-, and mediates uncontrolled proliferation and survival of tumor-mast cells. Therefore, targeting TAK1 and IKKs might be a novel approach to treat c-Kit-driven diseases.

The receptor tyrosine kinase c-Kit controls IL-33 receptor signaling in mast cells.

Members of the Toll/interleukin-1 receptor (TIR) family are of importance for host defense and inflammation. Here we report that the TIR-family member interleukin-33R (IL-33R) cross-activates the receptor tyrosine kinase c-Kit in human and murine mast cells. The IL-33R-induced activation of signal transducer and activator of transcription 3 (STAT3), extracellular signal-regulated kinase 1/2 (Erk1/2), protein kinase B (PKB), and Jun NH(2)-terminal kinase 1 (JNK1) depends on c-Kit and is required to elicit optimal effector functions. Costimulation with the c-Kit ligand stem cell factor (SCF) is necessary for IL-33-induced cytokine production in primary mast cells. The structural basis for this cross-activation is the complex formation between c-Kit, IL-33R, and IL-1R accessory protein (IL-1RAcP). We found that c-Kit and IL-1RAcP interact constitutively and that IL-33R joins this complex upon ligand binding. Our findings support a model in which signals from seemingly disparate receptors are integrated for full cellular responses.