Angiotensin II directly triggers endothelial exocytosis via protein kinase C-dependent protein kinase D2 activation
ABSTRACT Angiotensin II (AII) has been reported to induce leukocyte adhesion to endothelium through up-regulation of P-selectin surface expression. However, the underlying molecular and cellular mechanisms remain unknown. P-selectin is stored in Weibel-Palade bodies (WPBs), large secretory granules, in endothelial cells. In this study, we examined the role of protein kinase D (PKD), a newly identified regulator of protein transport, in AII-induced WPB exocytosis and the resultant P-selectin surface expression. We demonstrated that PKD2 was rapidly activated by AII in endothelial cells through phosphorylation of the activation loop at Ser744/748. AII-induced PKD2 activation correlated with increased P-selectin surface expression. Furthermore, AII-regulated PKD2 activation is protein kinase C (PKC) alpha-dependent. Importantly, knock-down of either PKD2 or PKCalpha expression inhibited AII-mediated P-selectin surface expression and monocyte adhesion. Our findings provide the first evidence that stimulation of P-selectin surface expression via PKCalpha-dependent PKD2 activation could be an important mechanism in the early onset of AII-initiated endothelial adhesiveness.
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ABSTRACT: It has become increasingly clear that the accumulation of proteins in specific regions of the plasma membrane can facilitate cellular communication. These regions, termed signaling microdomains, are found throughout the blood vessel wall where cellular communication, both within and between cell types, must be tightly regulated to maintain proper vascular function. We will define a cellular signaling microdomain and apply this definition to the plethora of means by which cellular communication has been hypothesized to occur in the blood vessel wall. To that end, we make a case for three broad areas of cellular communication where signaling microdomains could play an important role: 1) paracrine release of free radicals and gaseous molecules such as nitric oxide and reactive oxygen species; 2) role of ion channels including gap junctions and potassium channels, especially those associated with the endothelium-derived hyperpolarization mediated signaling, and lastly, 3) mechanism of exocytosis that has considerable oversight by signaling microdomains, especially those associated with the release of von Willebrand factor. When summed, we believe that it is clear that the organization and regulation of signaling microdomains is an essential component to vessel wall function.Pharmacological reviews 02/2014; 66(2):513-69. DOI:10.1124/pr.112.007351 · 18.55 Impact Factor
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ABSTRACT: Acute lung injury (ALI) and its most severe extreme the acute respiratory distress syndrome (ARDS) refer to increased-permeability pulmonary edema caused by a variety of pulmonary or systemic insults. ALI and in particular ARDS, are usually accompanied by refractory hypoxemia and the need for mechanical ventilation. In most cases, an exaggerated inflammatory and pro-thrombotic reaction to an initial stimulus, such as systemic infection, elicits disruption of the alveolo-capillary membrane and vascular fluid leak. The pulmonary endothelium is a major metabolic organ promoting adequate pulmonary and systemic vascular homeostasis, and a main target of circulating cells and humoral mediators under injury; pulmonary endothelium is therefore critically involved in the pathogenesis of ALI. In this review we will discuss mechanisms of pulmonary endothelial dysfunction and edema generation in the lung with special emphasis on the interplay between the endothelium, the immune and hemostatic systems, and highlight how these principles apply in the context of defined disorders and specific insults implicated in ALI pathogenesis.Vascular Pharmacology 08/2008; 49(4-6):119-33. DOI:10.1016/j.vph.2008.06.009 · 4.62 Impact Factor
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ABSTRACT: Exocytosis of endothelial Weibel-Palade bodies, which contain von Willebrand factor (VWF), P-selectin and other modulators, plays an important role in both inflammation and thrombosis. The present study investigates whether genipin, an aglycon of geniposide, inhibits endothelial exocytosis. Human umbilical vein endothelial cells (HUVECs) were isolated from umbilical cords and cultured. The concentration of VWF in cell supernatants was measured using an ELISA Kit. P-selectin translocation on the cell surface was analyzed by cell surface ELISA. Cell viability was measured using a Cell Counting Kit-8. Mouse bleeding times were measured by amputating the tail tip. Western blot analysis was used to determine the amount of endothelial nitric oxide synthase (eNOS) and phospho-eNOS present. Nitric oxide (NO) was measured in the cell supernatants as nitrite using an NO Colorimetric Assay. Genipin inhibited thrombin-induced VWF release and P-selectin translocation in HUVECs in a dose- and time-dependent manner. The drug had no cytotoxic effect on the cells at the same doses that were able to inhibit exocytosis. The functional study that demonstrated that genipin inhibited exocytosis in vivo also showed that genipin prolonged the mouse bleeding time. Furthermore, genipin activated eNOS phosphorylation, promoted enzyme activation and increased NO production. L-NAME, an inhibitor of NOS, reversed the inhibitory effects of genipin on endothelial exocytosis. Genipin inhibits endothelial exocytosis in HUVECs. The mechanism by which this compound inhibits exocytosis may be related to its ability to stimulate eNOS activation and NO production. Our findings suggest a novel anti-inflammatory mechanism for genipin. This compound may represent a new treatment for inflammation and/or thrombosis in which excess endothelial exocytosis plays a pathophysiological role.Acta Pharmacologica Sinica 05/2009; 30(5):589-96. DOI:10.1038/aps.2009.31 · 2.50 Impact Factor