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Objective: Leukocyte recruitment to the site of inflammation is a key event in a variety of cardiovascular pathologies. Infiltrating neutrophils constitute the first line of defense that precedes a second wave of emigrating monocytes reinforcing the inflammatory reaction. The mechanisms initiating this sequential process remained largely obscure., Approach and Results: Using advanced in vivo microscopy and in vitro/ex vivo techniques, we identified individual spatiotemporal expression patterns of selectins and their principal interaction partners on neutrophils, resident/inflammatory monocytes, and endothelial cells. Coordinating the intraluminal trafficking of neutrophils and inflammatory monocytes to common sites of extravasation, selectins assign different sites to these immune cells for their initial interactions with the microvascular endothelium. Whereas constitutively expressed leukocyte L-selectin/CD62L and endothelial P-selectin/CD62P together with CD44 and P-selectin glycoprotein ligand-1/CD162 initiate the emigration of neutrophils, de novo synthesis of endothelial E-selectin/CD62E launches the delayed secondary recruitment of inflammatory monocytes. In this context, P-selectin/CD62P and L-selectin/CD62L together with P-selectin glycoprotein ligand-1/CD162 and CD44 were found to regulate the flux of rolling neutrophils and inflammatory monocytes, whereas E-selectin/CD62E selectively adjusts the rolling velocity of inflammatory monocytes. Moreover, selectins and their interaction partners P-selectin glycoprotein ligand-1/CD162 and CD44 differentially control the intraluminal crawling behavior of neutrophils and inflammatory monocytes collectively enabling the sequential extravasation of these immune cells to inflamed tissue., Conclusions: Our findings provide novel insights into the mechanisms initiating the sequential infiltration of the perivascular tissue by neutrophils and monocytes in the acute inflammatory response and might thereby contribute to the development of targeted therapeutic strategies for prevention and treatment of cardiovascular diseases., (© 2015 American Heart Association, Inc.)

Obstructive sleep apnea (OSA) is a common sleep-related breathing disorder characterized by recurrent episodes of upper airway obstruction and subsequent hypoxia. In patients with OSA, severity and number of these hypoxic events positively correlate with the extent of associated cardiovascular pathology. The molecular mechanisms underlying intermittent hypoxia (IH)-driven cardiovascular disease in OSA, however, remain poorly understood—partly due to the lack of adequate experimental models. Here, we present a novel experimental approach that utilizes primary human endothelial cells cultivated under shear stress. Oxygen partial pressure dynamics were adopted in our in vitro model according to the desaturation-reoxygenation patterns identified in polysomnographic data of severe OSA patients (n = 10, with 892 severe desaturations, SpO2<80%). Using western blot analysis, we detected a robust activation of the two major inflammatory pathways ERK and NF-κB in endothelial cells, whereas no HIF1α and HIF2α protein stabilization was observed. In line with these findings, mRNA and protein expression of the pro-inflammatory adhesion and signaling molecule ICAM-1 and the chemokine CCL2 were significantly increased. Hence, we established a novel in vitro model for deciphering OSA-elicited effects on the vascular endothelium. First data obtained in this model point to the endothelial activation of pro-inflammatory rather than hypoxia-associated pathways in OSA. Future studies in this model might contribute to the development of targeted strategies against OSA-induced, secondary cardiovascular disease. [ABSTRACT FROM AUTHOR]

Aim: To assess the role of the endothelial glycocalyx (eGCX) for the uptake of nanoparticles by endothelial cells. Methods: The expression of the eGCX on cultured human umbilical vein endothelial cells was determined by immunostaining of heparan sulfate. Enzymatic degradation of the eGCX was achieved by incubating the cells with eGCX-shedding enzymes. The uptake of 50-nm polystyrene nanospheres was quantified by confocal microscopy. Results: Human umbilical vein endothelial cells expressed a robust eGCX when cultured for 10 days. The uptake of both carboxylated and aminated polystyrene nanospheres was significantly increased in cells in which the glycocalyx was enzymatically degraded, while it remained at a low level in cells with an intact glycocalyx. Conclusion: The eGCX constitutes a barrier against the internalization of blood-borne nanoparticles by endothelial cells. [ABSTRACT FROM AUTHOR]

Effective immune responses require the directed migration of leukocytes from the vasculature to the site of injury or infection. How immune cells “find” their site of extravasation remains largely obscure. Here, we identified a previously unrecognized role of platelets as pathfinders guiding leukocytes to their exit points in the microvasculature: upon onset of inflammation, circulating platelets were found to immediately adhere at distinct sites in venular microvessels enabling these cellular blood components to capture neutrophils and, in turn, inflammatory monocytes via CD40-CD40L-dependent interactions. In this cellular crosstalk, ligation of PSGL-1 by P-selectin leads to ERK1/2 MAPK-dependent conformational changes of leukocyte integrins, which promote the successive extravasation of neutrophils and monocytes to the perivascular tissue. Conversely, blockade of this cellular partnership resulted in misguided, inefficient leukocyte responses. Our experimental data uncover a platelet-directed, spatiotemporally organized, multicellular crosstalk that is essential for effective trafficking of leukocytes to the site of inflammation. [ABSTRACT FROM AUTHOR]

ABSTRACT Although carbon-based nanomaterials (CBNs) have been shown to exert prothrombotic effects in microvessels, it is poorly understood whether CBNs also have the potential to interfere with the process of leukocyte-endothelial cell interactions and whether the shape of CBNs plays a role in these processes. Thus, the aim of this study was to compare the acute effects of two differently shaped CBNs, fiber-shaped single-walled carbon nanotubes (SWCNT) and spherical ultrafine carbon black (CB), on thrombus formation as well as on leukocyte-endothelial cell interactions and leukocyte transmigration in the murine microcirculation upon systemic administration in vivo. Systemic administration of both SWCNT and CB accelerated arteriolar thrombus formation at a dose of 1 mg kg-1 body weight, whereas SWCNT exerted a prothrombotic effect also at a lower dose (0.1 mg kg-1 body weight). In vitro, both CBNs induced P-selectin expression on human platelets and formation of platelet-granulocyte complexes. In contrast, injection of fiber-shaped SWCNT or of spherical CB did not induce leukocyte-endothelial cell interactions or leukocyte transmigration. In vitro, both CBNs slightly increased the expression of activation markers on human monocytes and granulocytes. These findings suggest that systemic administration of CBNs accelerates arteriolar thrombus formation independently of the CBNs' shape, but does not induce leukocyte-endothelial cell interactions or leukocyte transmigration. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]