Rhinocetin, a Venom-derived Integrin-specific Antagonist Inhibits Collagen-induced Platelet and Endothelial Cell Functions

Vaiyapuri, Sakthivel; Hutchinson, E. Gail; Ali, Marfoua S.; Dannoura, Abeer; Stanley, Ronald G.; Harrison, Robert A.; Bicknell, Andrew B.; Gibbins, Jonathan M.

doi:10.1074/jbc.m112.381483

Cited by 24 publications

(18 citation statements)

References 54 publications

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“…Human platelet preparation, aggregation and activation assays were performed according to well-established methods [ 62 , 63 ]. Briefly, blood was obtained from aspirin-free, healthy human volunteers with written informed consent as approved by the University of Reading Research Ethics Committee.…”

Section: Methodsmentioning

confidence: 99%

Mycolactone-Dependent Depletion of Endothelial Cell Thrombomodulin Is Strongly Associated with Fibrin Deposition in Buruli Ulcer Lesions

et al. 2015

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A well-known histopathological feature of diseased skin in Buruli ulcer (BU) is coagulative necrosis caused by the Mycobacterium ulcerans macrolide exotoxin mycolactone. Since the underlying mechanism is not known, we have investigated the effect of mycolactone on endothelial cells, focussing on the expression of surface anticoagulant molecules involved in the protein C anticoagulant pathway. Congenital deficiencies in this natural anticoagulant pathway are known to induce thrombotic complications such as purpura fulimans and spontaneous necrosis. Mycolactone profoundly decreased thrombomodulin (TM) expression on the surface of human dermal microvascular endothelial cells (HDMVEC) at doses as low as 2ng/ml and as early as 8hrs after exposure. TM activates protein C by altering thrombin’s substrate specificity, and exposure of HDMVEC to mycolactone for 24 hours resulted in an almost complete loss of the cells’ ability to produce activated protein C. Loss of TM was shown to be due to a previously described mechanism involving mycolactone-dependent blockade of Sec61 translocation that results in proteasome-dependent degradation of newly synthesised ER-transiting proteins. Indeed, depletion from cells determined by live-cell imaging of cells stably expressing a recombinant TM-GFP fusion protein occurred at the known turnover rate. In order to determine the relevance of these findings to BU disease, immunohistochemistry of punch biopsies from 40 BU lesions (31 ulcers, nine plaques) was performed. TM abundance was profoundly reduced in the subcutis of 78% of biopsies. Furthermore, it was confirmed that fibrin deposition is a common feature of BU lesions, particularly in the necrotic areas. These findings indicate that there is decreased ability to control thrombin generation in BU skin. Mycolactone’s effects on normal endothelial cell function, including its ability to activate the protein C anticoagulant pathway are strongly associated with this. Fibrin-driven tissue ischemia could contribute to the development of the tissue necrosis seen in BU lesions.

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Section: Methodsmentioning

confidence: 99%

Mycolactone-Dependent Depletion of Endothelial Cell Thrombomodulin Is Strongly Associated with Fibrin Deposition in Buruli Ulcer Lesions

et al. 2015

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“…Human platelet preparation and aggregation assays were performed using standard methods172122. Briefly, blood was obtained from aspirin-free, healthy human volunteers with written informed consent as approved by the University of Reading Research Ethics Committee.…”

Section: Methodsmentioning

confidence: 99%

Connexin40 regulates platelet function

et al. 2013

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The presence of multiple connexins was recently demonstrated in platelets, with notable expression of Cx37. Studies with Cx37-deficient mice and connexin inhibitors established roles for hemichannels and gap junctions in platelet function. It was uncertain, however, whether Cx37 functions alone or in collaboration with other family members through heteromeric interactions in regulation of platelet function. Here we report the presence and functions of an additional platelet connexin, Cx40. Inhibition of Cx40 in human platelets or its deletion in mice reduces platelet aggregation, fibrinogen binding, granule secretion and clot retraction. The effects of the Cx37 inhibitor 37,43Gap27 on Cx40−/− mouse platelets and of the Cx40 inhibitor 40Gap27 on Cx37−/− mouse platelets revealed that each connexin is able to function independently. Inhibition or deletion of Cx40 reduces haemostatic responses in mice, indicating the physiological importance of this protein in platelets. We conclude that multiple connexins are involved in regulating platelet function, thereby contributing to haemostasis and thrombosis.

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“…8,9,18,19 Detailed materials and methods are provided in supplemental Methods (see supplemental Data available on the Blood Web site).…”

Section: Methodsmentioning

confidence: 99%

EphB2 regulates contact-dependent and contact-independent signaling to control platelet function

Vaiyapuri

Sage

Rana

et al. 2015

Blood

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Key Points• EphB2 regulates initial platelet activation in the absence of ligand binding in a contactindependent manner. • EphB2-mediated signaling regulates thrombus formation and clot retraction.The Eph kinases, EphA4 and EphB1, and their ligand, ephrinB1, have been previously reported to be present in platelets where they contribute to thrombus stability. Although thrombus formation allows for Eph-ephrin engagement and bidirectional signaling, the importance specifically of Eph kinase or ephrin signaling in regulating platelet function remained unidentified. In the present study, a genetic approach was used in mice to establish the contribution of signaling orchestrated by the cytoplasmic domain of EphB2 (a newly discovered Eph kinase in platelets) in platelet activation and thrombus formation. We conclude that EphB2 signaling is involved in the regulation of thrombus formation and clot retraction. Furthermore, the cytoplasmic tail of this Eph kinase regulates initial platelet activation in a contact-independent manner in the absence of Eph-ephrin ligation between platelets. Together, these data demonstrate that EphB2 signaling not only modulates platelet function within a thrombus but is also involved in the regulation of the function of isolated platelets in a contact-independent manner. (Blood. 2015;125(4):720-730)

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Rhinocetin, a Venom-derived Integrin-specific Antagonist Inhibits Collagen-induced Platelet and Endothelial Cell Functions

Cited by 24 publications

References 54 publications

Mycolactone-Dependent Depletion of Endothelial Cell Thrombomodulin Is Strongly Associated with Fibrin Deposition in Buruli Ulcer Lesions

Mycolactone-Dependent Depletion of Endothelial Cell Thrombomodulin Is Strongly Associated with Fibrin Deposition in Buruli Ulcer Lesions

Connexin40 regulates platelet function

EphB2 regulates contact-dependent and contact-independent signaling to control platelet function

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