Search

Eine Langzeittherapie mit Methotrexat (MTX) ist als Auslöser einer MTX-assoziierten lymphoproliferativen Erkrankung (MTX-LPD) bekannt. Unter einer Kombinationstherapie mit MTX und Ustekinumab kam es bei einer 58-jährigen Patientin mit Psoriasis vulgaris und Pityriasis rubra pilaris innerhalb von 4 Monaten zu einer generalisierten Lymphadenopathie. Die Histologie deutete zunächst auf ein Hodgkin-Lymphom hin. Nur die Zusammenschau mit dem klinischen Hintergrund konnte die Diagnose einer MTX-LPD aufzeigen. Unseres Wissens ist dies der erste Fall einer MTX-LPD nach nur 4 Monaten Therapie.

Hintergrund: Der Pikosekundenlaser gehört zu den jüngsten in der Dermatologie genutzten Lasersystemen. Ursprünglich wurde er zur Optimierung von Tätowierungsentfernungen entwickelt, Fortschritte in dieser Technologie erweiterten das Indikationsspektrum des Pikosekundenlasers jedoch erheblich. Ziel der Arbeit: Dieser Artikel gibt eine Übersicht über den technischen Hintergrund sowie die Indikationen des Pikosekundenlasers in der dermatologischen Lasermedizin und erläutert die Möglichkeiten und Grenzen dieses Lasersystems. Material und Methoden: Grundlage dieses Beitrages sind systematische Literaturanalyse sowie Erfahrungen aus der klinischen Praxis in der universitären Laserambulanz. Ergebnisse: Der Pikosekundenlaser ermöglicht durch Impulse im Pikosekundenbereich und den Wirkmechanismus des „laser-induced optical breakdown“ eine besonders schonende und effektive Behandlung. Im Vergleich zu den gütegeschalteten Lasern weist der Pikosekundenlaser weniger hitzeinduzierte Nebenwirkungen auf und geht mit einer geringeren Schmerzintensität sowie einer kürzeren Ausfallzeit einher. Insbesondere durch die fraktionierte nichtablative Anwendung haben sich zusätzliche Anwendungsgebiete ergeben, darunter die Hautverjüngung und Narbentherapie. Schlussfolgerung: Der Pikosekundenlaser findet ein breites Anwendungsspektrum in der dermatologischen Lasermedizin. Die aktuelle Datenlage deutet darauf hin, dass der Laser eine effektive Methode mit einem geringen Nebenwirkungsprofil ist. Um die Wirksamkeit, Verträglichkeit und Patientenzufriedenheit evidenzbasiert beurteilen zu können, sind weitere prospektive Studien notwendig.

Clinically relevant brain metastases (BMs) frequently form in cancer patients, with limited options for effective treatment. Circulating cancer cells must first permanently arrest in brain microvessels to colonize the brain, but the critical factors in this process are not well understood. Here, in vivo multiphoton laser-scanning microscopy of the entire brain metastatic cascade allowed unprecedented insights into how blood clot formation and von Willebrand factor (VWF) deposition determine the arrest of circulating cancer cells and subsequent brain colonization in mice. Clot formation in brain microvessels occurred frequently (>95%) and specifically at intravascularly arrested cancer cells, allowing their long-term arrest. An extensive clot embedded ∼20% of brain-arrested cancer cells, and those were more likely to successfully extravasate and form a macrometastasis. Mechanistically, the generation of tissue factor-mediated thrombin by cancer cells accounted for local activation of plasmatic coagulation in the brain. Thrombin inhibition by treatment with low molecular weight heparin or dabigatran and an anti-VWF antibody prevented clot formation, cancer cell arrest, extravasation, and the formation of brain macrometastases. In contrast, tumor cells were not able to directly activate platelets, and antiplatelet treatments did reduce platelet dispositions at intravascular cancer cells but did not reduce overall formation of BMs. In conclusion, our data show that plasmatic coagulation is activated early by intravascular tumor cells in the brain with subsequent clot formation, which led us to discover a novel and specific mechanism that is crucial for brain colonization. Direct or indirect thrombin and VWF inhibitors emerge as promising drug candidates for trials on prevention of BMs.

Clinically relevant brain metastases (BMs) frequently form in cancer patients, with limited options for effective treatment. Circulating cancer cells must first permanently arrest in brain microvessels to colonize the brain, but the critical factors in this process are not well understood. Here, in vivo multiphoton laser-scanning microscopy of the entire brain metastatic cascade allowed unprecedented insights into how blood clot formation and von Willebrand factor (VWF) deposition determine the arrest of circulating cancer cells and subsequent brain colonization in mice. Clot formation in brain microvessels occurred frequently (>95%) and specifically at intravascularly arrested cancer cells, allowing their long-term arrest. An extensive clot embedded ∼20% of brain-arrested cancer cells, and those were more likely to successfully extravasate and form a macrometastasis. Mechanistically, the generation of tissue factor-mediated thrombin by cancer cells accounted for local activation of plasmatic coagulation in the brain. Thrombin inhibition by treatment with low molecular weight heparin or dabigatran and an anti-VWF antibody prevented clot formation, cancer cell arrest, extravasation, and the formation of brain macrometastases. In contrast, tumor cells were not able to directly activate platelets, and antiplatelet treatments did reduce platelet dispositions at intravascular cancer cells but did not reduce overall formation of BMs. In conclusion, our data show that plasmatic coagulation is activated early by intravascular tumor cells in the brain with subsequent clot formation, which led us to discover a novel and specific mechanism that is crucial for brain colonization. Direct or indirect thrombin and VWF inhibitors emerge as promising drug candidates for trials on prevention of BMs.

The frequent von Willebrand factor (VWF) variant p.Phe2561Tyr is located within the C4 domain, which also harbors the platelet GPIIb/IIIa-binding RGD sequence. To investigate its potential effect on hemostasis, we genotyped 865 patients with coronary artery disease (CAD), 915 with myocardial infarction (MI), and 417 control patients (Ludwigshafen Risk and Cardiovascular Health Study) and performed functional studies of this variant. A univariate analysis of male and female carriers of the Tyr2561 allele aged 55 years or younger revealed an elevated risk for repeated MI (odds ratio, 2.53; 95% confidence interval [CI], 1.07-5.98). The odds ratio was even higher in females aged 55 years or younger, at a value of 5.93 (95% CI, 1.12-31.24). Cone and plate aggregometry showed that compared with Phe2561, Tyr2561 was associated with increased platelet aggregate size both in probands’ blood and with the recombinant variants. Microfluidic assays revealed that the critical shear rate for inducing aggregate formation was decreased to 50% by Tyr2561 compared with Phe2561. Differences in C-domain circular dichroism spectra resulting from Tyr2561 suggest an increased shear sensitivity of VWF as a result of altered association of the C domains that disrupts the normal dimer interface. In summary, our data emphasize the functional effect of the VWF C4 domain for VWF-mediated platelet aggregation in a shear-dependent manner and provide the first evidence that a functional variant of VWF plays a role in arterial thromboembolism.

The frequent von Willebrand factor (VWF) variant p.Phe2561Tyr is located within the C4 domain, which also harbors the platelet GPIIb/IIIa-binding RGD sequence. To investigate its potential effect on hemostasis, we genotyped 865 patients with coronary artery disease (CAD), 915 with myocardial infarction (MI), and 417 control patients (Ludwigshafen Risk and Cardiovascular Health Study) and performed functional studies of this variant. A univariate analysis of male and female carriers of the Tyr2561 allele aged 55 years or younger revealed an elevated risk for repeated MI (odds ratio, 2.53; 95% confidence interval [CI], 1.07-5.98). The odds ratio was even higher in females aged 55 years or younger, at a value of 5.93 (95% CI, 1.12-31.24). Cone and plate aggregometry showed that compared with Phe2561, Tyr2561 was associated with increased platelet aggregate size both in probands' blood and with the recombinant variants. Microfluidic assays revealed that the critical shear rate for inducing aggregate formation was decreased to 50% by Tyr2561 compared with Phe2561. Differences in C-domain circular dichroism spectra resulting from Tyr2561 suggest an increased shear sensitivity of VWF as a result of altered association of the C domains that disrupts the normal dimer interface. In summary, our data emphasize the functional effect of the VWF C4 domain for VWF-mediated platelet aggregation in a shear-dependent manner and provide the first evidence that a functional variant of VWF plays a role in arterial thromboembolism.

To measure and to modulate the invasive potential of urothelial carcinoma of the bladder (UCB) cells in a standardized preclinical setting using broad-spectrum matrix-metalloproteinase (MMPs) inhibitors and specific targeting of MMP7. MMP expression levels in UCB cells were determined by quantitative real-time PCR (qRT-PCR) and gel zymographies of cell supernatants (MMP9, MMP2 and MMP1) and cell lysates (MMP7). The invasiveness of human UCB cells (HT1197 and T24/83) and human benign urothelial cells (UROtsa) was modulated by a broad-spectrum MMP inhibitor (4-Aminobenzoyl-Gly-Pro-D-Leu-D-Ala hydroxamic acid; AHA) and by MMP7 specific siRNAs. MMP7 knockdown efficiency was assessed by qRT-PCR and western blot. Invasive potential of UCB cells was measured by a standardized trans-epithelial electrical resistance (TEER) assay. Different MMP secretion profiles were measured in UCB cells. The active form of MMP7 was exclusively detected in HT1197 cells. Characteristic TEER breakdown patterns were observed in UCB cells when compared to benign cells. Invasive potentials were significantly higher in HT1197 cells than in T24/83 and in UROtsa cells [14.8±5.75 vs. 1.5±0.56 and 1.2±0.15, respectively; p < 0.01]. AHA treatment reduced the invasive potential of HT1197 cells. Also the specific downregulation of MMP7 by siRNA lowered the HT1197 cell invasiveness [20±1.0 vs. 16±2.8; p < 0.05]. Neither AHA nor MMP-7 siRNA transfection altered the invasive potential of T24/83 cells. Invasion of UCB is partially dependent on MMPs. Specific targeting of MMP7 by siRNA reduces the invasive potential in a subgroup of UCB cells. Therefore, MMP7 represents a potential therapeutic target which warrants further investigation.

The dynamic change from a globular conformation to an elongated fiber determines the ability of von Willebrand factor (VWF) to trap platelets. Fiber formation is favored by the anchorage of VWF to the endothelial cell surface, and VWF-platelet aggregates on the endothelium contribute to inflammation, infection, and tumor progression. Although P-selectin and a?ß3-integrins may bind VWF, their precise role is unclear, and additional binding partners have been proposed. In the present study, we evaluated whether the endothelial glycocalyx anchors VWF fibers to the endothelium. Using microfluidic experiments, we showed that stabilization of the endothelial glycocalyx by chitosan oligosaccharides or overexpression of syndecan-1 (SDC-1) significantly supports the binding of VWF fibers to endothelial cells. Heparinase-mediated degradation or impaired synthesis of heparan sulfate (HS), a major component of the endothelial glycocalyx, reduces VWF fiber–dependent platelet recruitment. Molecular interaction studies using flow cytometry and live-cell fluorescence microscopy provided further evidence that VWF binds to HS linked to SDC-1. In a murine melanoma model, we found that protection of the endothelial glycocalyx through the silencing of heparanase increases the number of VWF fibers attached to the wall of tumor blood vessels. In conclusion, we identified HS chains as a relevant binding factor for VWF fibers at the endothelial cell surface in vitro and in vivo.

Tumor progression is associated with aberrant hemostasis, and patients with malignant diseases have an elevated risk of developing thrombosis. A crosstalk among the vascular endothelium, components of the coagulation cascade, and cancer cells transforms the intravascular milieu to a prothrombotic, proinflammatory, and cell-adhesive state. We review the existing evidence on activation of the coagulation system and its implication in genitourinary malignancies and discuss the potential therapeutic benefit of antithrombotic agents. A literature review was performed searching the Medline database and the Cochrane Library for original articles and reviews. A second search identified studies reporting on oncological benefit of anticoagulants in genitourinary cancer. An elevated expression of procoagulatory tissue factor on tumor cells and tumor-derived microparticles seems to stimulate cancer development and progression. Several components of the hemostatic system, including D-dimers, von Willebrand Factor, thrombin, fibrin-/ogen, soluble P-selectin, and prothrombin fragments 1 + 2 were either overexpressed or overactive in genitourinary cancers. Hypercoagulation was in general associated with a poorer prognosis. Experimental models and small trials in humans showed reduced cancer progression after treatment with anticoagulants. Main limitations of these studies were heterogeneous experimental methodology, small patient numbers, and a lack of prospective validation. In conclusion, experimental and clinical evidence suggests procoagulatory activity of genitourinary neoplasms, particularly in prostate, bladder and kidney cancer. This may promote the risk of vascular thrombosis but also metastatic progression. Clinical studies linked elevated biomarkers of hemostasis with poor prognosis in patients with genitourinary cancers. Thus, anticoagulation may have a therapeutic role beyond prevention of thromboembolism.

Livedoid vasculopathy is a thrombotic skin disease characterised by recurrent occlusion of the cutaneous microcirculation in lower extremities, which results in skin infarctions with painful ulcerations and irreversible scar formation. Rivaroxaban is a direct factor Xa inhibitor that prevents thrombus formation. We investigated whether rivaroxaban is effective for the treatment of livedoid vasculopathy.

Tumor-mediated procoagulatory activity leads to venous thromboembolism and supports metastasis in cancer patients. A prerequisite for metastasis formation is the interaction of cancer cells with endothelial cells (ECs) followed by their extravasation. Although it is known that activation of ECs and the release of the procoagulatory protein von Willebrand factor (VWF) is essential for malignancy, the underlying mechanisms remain poorly understood. We hypothesized that VWF fibers in tumor vessels promote tumor-associated thromboembolism and metastasis. Using in vitro settings, mouse models, and human tumor samples, we showed that melanoma cells activate ECs followed by the luminal release of VWF fibers and platelet aggregation in tumor microvessels. Analysis of human blood samples and tumor tissue revealed that a promoted VWF release combined with a local inhibition of proteolytic activity and protein expression of ADAMTS13 (a disintegrin-like and metalloproteinase with thrombospondin type I repeats 13) accounts for this procoagulatory milieu. Blocking endothelial cell activation by the low-molecular-weight heparin tinzaparin was accompanied by a lack of VWF networks and inhibited tumor progression in a transgenic mouse model. Our findings implicate a mechanism wherein tumor-derived vascular endothelial growth factor-A (VEGF-A) promotes tumor progression and angiogenesis. Thus, targeting EC activation envisions new therapeutic strategies attenuating tumor-related angiogenesis and coagulation.

Tumor-mediated procoagulatory activity leads to venous thromboembolism and supports metastasis in cancer patients. A prerequisite for metastasis formation is the interaction of cancer cells with endothelial cells (ECs) followed by their extravasation. Although it is known that activation of ECs and the release of the procoagulatory protein von Willebrand factor (VWF) is essential for malignancy, the underlying mechanisms remain poorly understood. We hypothesized that VWF fibers in tumor vessels promote tumor-associated thromboembolism and metastasis. Using in vitro settings, mouse models, and human tumor samples, we showed that melanoma cells activate ECs followed by the luminal release of VWF fibers and platelet aggregation in tumor microvessels. Analysis of human blood samples and tumor tissue revealed that a promoted VWF release combined with a local inhibition of proteolytic activity and protein expression of ADAMTS13 (a disintegrin-like and metalloproteinase with thrombospondin type I repeats 13) accounts for this procoagulatory milieu. Blocking endothelial cell activation by the low-molecular-weight heparin tinzaparin was accompanied by a lack of VWF networks and inhibited tumor progression in a transgenic mouse model. Our findings implicate a mechanism wherein tumor-derived vascular endothelial growth factor-A (VEGF-A) promotes tumor progression and angiogenesis. Thus, targeting EC activation envisions new therapeutic strategies attenuating tumor-related angiogenesis and coagulation.

Von Willebrand factor (VWF) plays a central role in hemostasis. Triggered by shear-stress, it adheres to platelets at sites of vascular injury. Inactivation of VWF has been associated to the shielding of its adhesion sites and proteolytic cleavage. However, the molecular nature of this shielding and its coupling to cleavage under shear-forces in flowing blood remain unknown. In this study, we describe, to our knowledge, a new force-sensory mechanism for VWF-platelet binding, which addresses these questions, based on a combination of molecular dynamics (MD) simulations, atomic force microscopy (AFM), and microfluidic experiments. Our MD simulations demonstrate that the VWF A2 domain targets a specific region at the VWF A1 domain, corresponding to the binding site of the platelet glycoprotein Ibα(GPIbα) receptor, thereby causing its blockage. This implies autoinhibition of the VWF for the binding of platelets mediated by the A1-A2 protein-protein interaction. During force-probe MD simulations, a stretching force dissociated the A1A2 complex, thereby unblocking the GPIbαbinding site. Dissociation was found to be coupled to the unfolding of the A2 domain, with dissociation predominantly occurring before exposure of the cleavage site in A2, an observation that is supported by our AFM experiments. This suggests that the A2 domain prevents platelet binding in a force-dependent manner, ensuring that VWF initiates hemostasis before inactivation by proteolytic cleavage. Microfluidic experiments with an A2-deletion VWF mutant resulted in increased platelet binding, corroborating the key autoinhibitory role of the A2 domain within VWF multimers. Overall, autoinhibition of VWF mediated by force-dependent interdomain interactions offers the molecular basis for the shear-sensitive growth of VWF-platelet aggregates, and might be similarly involved in shear-induced VWF self-aggregation and other force-sensing functions in hemostasis.

Tumor cell extravasation is a critical step in the metastatic cascade and requires interaction between the tumor cell and the endothelium. Although cancer progression depends on a complex network of mechanisms, including inflammation and coagulation, the involvement of tumor-induced endothelium activation and the subsequent release of procoagulatory factors in this process are not well understood. Using tissue sections from patients with malignant melanoma, immunofluorescence studies for the presence of von Willebrand factor (VWF) clearly demonstrated endothelium activation and the formation of ultra-large VWF fibers in these patients. In vitroanalyses revealed that supernatants from highly invasive melanoma cells induced an acute endothelium activation measured by VWF, P-selectin, and angiopoietin-2 release. Proteome profiling identified vascular endothelial growth factor A (VEGF-A) as the main mediator of endothelium activation. Inhibition and knock-down of VEGF-A in melanoma cells led to a rigorous decrease in VWF exocytosis. Selective small-interfering RNA to matrix metalloproteinase-2 (MMP-2) inhibited endothelium activation, and this effect correlated with reduced VEGF-A content in the supernatants of melanoma cells. Further experiments showed that active MMP-2 regulates VEGF-A in melanoma cells on a transcriptional level via an integrin αvβ5/phosphoinositide-3-kinase–dependent pathway. In conclusion, these results indicate an important role of VEGF-A in acute endothelium activation and provide clear evidence that MMP-2 plays a pivotal role in the autocrine regulation of VEGF-A expression in melanoma cells.

Increasing incidence of inflammatory skin diseases such as Atopic Dermatitis (AD) has been noted in the past years. According to recent estimations around 15% of newborn subjects are affected with a disease severity that requires medical treatment. Although its pathogenesis is multifactorial, recent reports indicate that an impaired physical skin barrier predispose for the development of AD. The major part of this barrier is formed by the stratum corneum (SC) wherein corneocytes are embedded in a complex matrix of proteins and lipids. Its components were synthesized in the stratum granulosum (SG) and secreted via lamellar bodies at the SC/SG interface. Within a clinical in vivo study we focused on the skin metabolism at the SC/SG interface in AD affected patients in comparison to healthy subjects. Measurement of fluorescence life-time of NADH provides access to the metabolic state of skin. Due to the application of a 5D intravital tomographic skin analysis we facilitate the non-invasive investigation of human epidermis in the longitudinal course of AD therapy. We could ascertain by blinded analysis of 40 skin areas of 20 patients in a three month follow-up that the metabolic status at the SC/SG interface was altered in AD compromised skin even in non-lesional, apparent healthy skin regions. This illustrates an impaired skin barrier formation even at non-affected skin of AD subjects appearing promotive for the development of acute skin inflammation. Therefore, our findings allow a deeper understanding of the individual disease development and the improved management of the therapeutic intervention in clinical application.

von Willebrand factor (VWF) is an important player in hemostasis but has also been suggested to promote inflammatory processes. Gene ablation of VWF causes a simultaneous defect in P-selectin expression making it difficult to identify VWF-specific functions. Therefore, we analyzed whether blocking antibodies against VWF would be able to interfere with neutrophil extravasation. We found that these antibodies inhibited neutrophil recruitment into thioglycollate-inflamed peritoneum and KC-stimulated cremaster by approximately 50%. Whereas platelet-VWF was not involved, the contribution of VWF to granulocyte recruitment was strictly dependent on the presence of platelets and the accessibility of their VWF-receptor glycoprotein Ib. Surprisingly, platelet P-selectin was largely dispensable for leukocyte extravasation, in agreement with our observation that anti-VWF antibodies did not affect leukocyte rolling and adhesion. Searching for possible effects downstream of leukocyte capture, we found that anti-VWF antibodies significantly inhibited thioglycollate-induced vascular permeability. The increase of permeability was independent of circulating granulocytes, showing that it was not a side effect of neutrophil diapedesis. Collectively, our results demonstrate that VWF-associated platelets strongly support neutrophil extravasation at a step downstream of leukocyte docking to the vessel wall. This step could be related to leukocyte diapedesis facilitated by destabilization of the endothelial barrier.

von Willebrand factor (VWF) is an important player in hemostasis but has also been suggested to promote inflammatory processes. Gene ablation of VWF causes a simultaneous defect in P-selectin expression making it difficult to identify VWF-specific functions. Therefore, we analyzed whether blocking antibodies against VWF would be able to interfere with neutrophil extravasation. We found that these antibodies inhibited neutrophil recruitment into thioglycollate-inflamed peritoneum and KC-stimulated cremaster by approximately 50%. Whereas platelet-VWF was not involved, the contribution of VWF to granulocyte recruitment was strictly dependent on the presence of platelets and the accessibility of their VWF-receptor glycoprotein Ib. Surprisingly, platelet P-selectin was largely dispensable for leukocyte extravasation, in agreement with our observation that anti-VWF antibodies did not affect leukocyte rolling and adhesion. Searching for possible effects downstream of leukocyte capture, we found that anti-VWF antibodies significantly inhibited thioglycollate-induced vascular permeability. The increase of permeability was independent of circulating granulocytes, showing that it was not a side effect of neutrophil diapedesis. Collectively, our results demonstrate that VWF-associated platelets strongly support neutrophil extravasation at a step downstream of leukocyte docking to the vessel wall. This step could be related to leukocyte diapedesis facilitated by destabilization of the endothelial barrier.

The electrical resistance breakdown of the Madin-Darby canine kidney (MDCK) cell monolayer provides a continuous assay system for cancer invasion that detects functional changes before morphological alterations. In this study, we address the question of whether physical contact between tumor cell and epithelial monolayer is a prerequisite for tumor cell invasion. When human melanoma cells were seeded directly (i.e., physical contact) on top of an electrically tight epithelial cell layer (5,800 ± 106 Ω · cm2), electrical monolayer leakage led to an 18 ± 3% reduction of transepithelial electrical resistance within 24 h. However, when melanoma cells were seeded close to the basolateral surface of the epithelial cell monolayer but separated by a filter membrane (i.e., no physical contact), electrical leakage occurred even more quickly (42 ± 3% reduction in 24 h). Atomic force microscopy detected discrete structural changes between cells. Electrical leakage was effectively blocked by α2-macroglobulin or ilomastat, inhibitors of matrix metalloproteinases. We conclude that exocytosis of soluble proteases causes electrical breakdown of the MDCK monolayer, independently of physical contact between tumor cells and the monolayer.

Atomic force microscopy (AFM) is a useful technique for imaging the surface of living cells in three dimensions. The authors applied AFM to obtain morphological information of individual cultured endothelial cells of bovine aorta under stationary and strain conditions and to simultaneously measure changes in cell volume in response to aldosterone. This mineralocorticoid hormone is known to have acute, non-genomic effects on intracellular pH, intracellular electrolytes and inositol-1,4,5-triphosphate production. In this study whether endothelial cells under tension change their volume in response to aldosterone was tested. Such changes were already shown in human leukocytes measured by Coulter counter. In contrast to leukocytes that are more or less spherical and live in suspension, endothelial cells exhibit a complex morphology and adhere to a substrate. Thus, measurements of discrete cell volume changes in endothelial cells under physiological condition is only feasible with more sophisticated techniques. By using AFM we could precisely measure the absolute cell volume of individual living endothelial cells. Before the addition of aldosterone the cell volume of mechanically stressed endothelial cells mimicking arterial blood pressure was 1827±172fl. Cell volume was found to increase by 28% 5min after hormone exposure. Twenty-five minutes later cell volume was back to normal despite the continuous presence of aldosterone in the medium. Amiloride, a blocker of the plasma membrane Na+/H+exchanger prevented the initial aldosterone-induced volume increase. Taken together, AFM disclosed a transient swelling of endothelial cells induced by the activation of an aldosterone sensitive plasma membrane Na+/H+exchanger.

Pathomechanisms in IgA pemphigus are assumed to rely on Fc-dependent cellular activation by antigen-specific IgA autoantibodies, however, models for disease and more detailed pathophysiologic data are lacking. We here aimed to establish in vitromodels of disease for IgA pemphigus, allowing to study effects of the interaction of anti-keratinocyte IgA with cell-surface Fc alpha receptors. Employing multiple in vitroassays, such as a skin cryosection assay and a human skin organ culture model, we here present mechanistic data for the pathogenesis of IgA pemphigus, mediated by anti-desmoglein 3 IgA autoantibodies. Our results reveal that this disease is dependent on Fc alpha receptor-mediated activation of leukocytes in the epidermis. Importantly, this cell-dependent pathology can be dose-dependently abrogated by peptide-mediated inhibition of Fc alpha receptor:IgA interaction, as confirmed in an additional model for IgA-dependent disease, i.e., IgA vasculitis. These data suggest that IgA pemphigus can be modeled in vitro, and IgA pemphigus and IgA vasculitis are Fc alpha receptor-dependent disease entities that can be specifically targeted in these experimental systems.

Schneppenheim: SHIRE: Consultancy; CSL Behring: Consultancy.

The frequent von Willebrand factor (VWF) variant p.Phe2561Tyr is located within the C4 domain, which also harbors the platelet GPIIb/IIIa-binding RGD sequence. To investigate its potential effect on hemostasis, we genotyped 865 patients with coronary artery disease (CAD), 915 with myocardial infarction (MI) and 417 controls (Ludwigshafen Risk and Cardiovascular Health Study (LURIC)) and performed functional studies of this variant. A univariate analysis of male and female carriers of the Tyr2561 allele ≤55 years revealed an elevated risk for repeated MI (Odds ratio: 2.53, 95% CI:1.07-5.98). The Odds ratio was even higher in females ≤55 years at a value of 5.93 (95% CI:1.12-31.24). Cone and plate aggregometry showed, that compared to Phe2561, Tyr2561 was associated with increased platelet aggregate size, both in probands' blood and with the recombinant variants. Inhibition of the VWF-GPIIb/IIIa interaction abrogated the effect. Microfluidic assays revealed that the critical shear rate for inducing aggregate formation was decreased to 50% by Tyr2561 compared to Phe2561. Differences in C-domain circular dichroism spectra resulting from Tyr2561 suggest an increased shear sensitivity of VWF, as a result of altered association of the C-domains that disrupts the normal dimer interface. In summary, our data emphasize the functional impact of the VWF C4-domain for VWF-mediated platelet aggregation in a GPIIb/IIIa-dependent manner and provide the first evidence that a functional variant of VWF plays a role in arterial thromboembolism.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.