Sandwich Immunoassay for Soluble Glycoprotein VI in Patients with Symptomatic Coronary Artery Disease

Bigalke, Boris; Pötz, Oliver; Kremmer, Elisabeth; Geisler, Tobias; Seizer, Peter; Püntmann, Valentina O.; Phinikaridou, Alkystis; Chiribiri, Amedeo; Nagel, Eike; Botnar, René M.; Joos, Thomas; Gawaz, Meinrad

doi:10.1373/clinchem.2010.158527

Cited by 25 publications

(25 citation statements)

References 31 publications

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“…by guest www.bloodjournal.org From than, shedding induced by GPVI ligands, calmodulin inhibitors, thiol-modifying reagents, and anti-GPVI monoclonal antibodies. 10,41 We also measured plasma sGPVI levels in a group of patients chronically exposed to elevated intracoronary shear rates that were 3-fold higher than sGPVI in healthy donors, 42 consistent with other reports of elevated sGPVI in SAP, 22 suggesting that elevated shear contributes to shedding of GPVI in vivo. Shearinduced GPVI shedding was significantly inhibited by an ADAM10-selective inhibitor GI254023, 23 suggesting that ADAM10 may play an important role in this process ( Figures 1D-E and 2C).…”

Section: Shear-induced Shedding Of Gpvi 4317supporting

confidence: 86%

“…48 Potentially, sGPVI measured in plasma could be a useful marker of pathologic conditions associated with high shear rate, including vascular atherosclerosis and thrombosis, prosthetic devices, graft junction, and vascular abnormalities. 17,22,42 As an activator of metalloproteinase-dependent GPVI shedding, elevated shear had very few prerequisites. There was no requirement for engagement of GPIb␣ or platelet aggregation, and shear-induced shedding was independent of intracellular signaling pathways as pathway inhibitors that blocked platelet aggregation did not block release of sGPVI.…”

Section: Shear-induced Shedding Of Gpvi 4317mentioning

confidence: 99%

“…20 In this regard, elevated surface levels of GPVI on circulating platelets correlate with increased incidence of stroke and acute coronary syndrome, 21 and elevated plasma levels of sGPVI have been reported in a stable angina pectoris (SAP) cohort. 22 We investigated the ability of shear stress, in the absence of GPVI ligands, to induce metalloproteinase-dependent GPVI shedding and release of sGPVI into plasma. We found that GPVI shedding occurred within 1 minute of exposure to shear, continued after cessation of shear, and was independent of VWF binding to GPIb␣, ␣ IIb ␤ 3 engagement, and platelet activation.…”

Section: Introductionmentioning

confidence: 99%

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Pathologic shear triggers shedding of vascular receptors: a novel mechanism for down-regulation of platelet glycoprotein VI in stenosed coronary vessels

Al‐Akchar

Tan

Qiao

et al. 2012

Blood

101

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Ligand-induced ectodomain shedding of glycoprotein VI (GPVI) is a metalloproteinase-dependent event. We examined whether shear force, in the absence of GPVI ligand, was sufficient to induce shedding of GPVI. Human-citrated platelet-rich plasma or washed platelets were subjected to increasing shear rates in a cone-plate viscometer, and levels of intact and cleaved GPVI were examined by Western blot and ELISA. Pathophysiologic shear rates (3000-10 000 seconds ؊1 ) induced platelet aggregation and metalloproteinase-dependent appearance of soluble GPVI ectodomain, and GPVI platelet remnant. Shedding of GPVI continued after transient exposure to shear. Blockade of ␣ IIb ␤ 3 , GPIb␣, or intracellular signaling inhibited shear-induced platelet aggregation but minimally affected shearinduced shedding of GPVI. Shearinduced GPVI shedding also occurred in platelet-rich plasma or washed platelets isolated from a von Willebrand disease type 3 patient with no detectable VWF, implying that shear-induced activation of platelet metalloproteinases can occur in the absence of GPVI and GPIb␣ ligands. Significantly elevated levels of sGPVI were observed in 10 patients with stable angina pectoris, with well-defined single vessel coronary artery disease and mean intracoronary shear estimates at 2935 seconds ؊1 (peak shear, 19 224 seconds ؊1 ). Loss of GPVI in platelets exposed to shear has potential implications for the stability of a forming thrombus at arterial shear rates. IntroductionPlatelet activation and accumulation at sites of vascular injury play a critical role in thrombus formation. This complex process is mainly initiated by 3 different but overlapping pathways: (1) exposure of subendothelial matrix proteins, including collagen and VWF, which activate the platelet adhesion-signaling receptors glycoprotein VI (GPVI) and GPIb␣ of the GPIb-IX-V complex, respectively; (2) exposure of tissue factor, which activates the coagulation cascade resulting in formation of active thrombin facilitating fibrin deposition as well as enhancing platelet activation; and (3) disturbed blood flow because of narrowing of the vascular lumen, which modulates the adhesive function of platelets and accelerates platelet activation and thrombus growth. 1 Indeed, changes in blood flow rates and hydrodynamic force are now recognized to play a more critical role in thrombus formation, especially at sites of vascular occlusion, as indicated by the ability of elevated (pathologic) shear stress to induce stable platelet aggregate formation without the requirement for platelet activation and adhesion 2 or for soluble agonists. 3 Human platelets normally circulate in a resting state and are exposed to shear rates within a physiologic range (ϳ 20-2000 seconds Ϫ1 ). Platelets may encounter shear rates beyond 10 000 seconds Ϫ1 under pathologic conditions, for example, in a stenosed atherosclerotic artery, and become activated and begin to aggregate. [3][4][5] Shear-dependent platelet activation is initiated by binding of plasma VWF to platelets primarily throu...

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Section: Shear-induced Shedding Of Gpvi 4317supporting

confidence: 86%

Section: Shear-induced Shedding Of Gpvi 4317mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Pathologic shear triggers shedding of vascular receptors: a novel mechanism for down-regulation of platelet glycoprotein VI in stenosed coronary vessels

Al‐Akchar

Tan

Qiao

et al. 2012

Blood

101

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“…28,29 Elevated fluid shear stress and active factor X (FXa) also trigger GPVI shedding, quantified by detection of the soluble ectodomain fragment (sGPVI) in plasma by an enzyme-linked immunosorbent assay. 30,31 Plasma sGPVI reflects platelet activation in thrombotic conditions including microangiopathy, 32 stroke, 33 DIC, 33 and Alzheimer's disease 34 and rheumatoid arthritis. 28,35 However, elevated plasma sGPVI levels in these patient groups is surprising, as only a fraction of platelets would be exposed to collagen, FXa, or elevated shear, and neither FcgRIIA or CLEC-2 plays major roles in hemostasis/thrombosis.…”

Section: Introductionmentioning

confidence: 99%

Soluble GPVI is elevated in injured patients: shedding is mediated by fibrin activation of GPVI

Montague

Delierneux

Lecut³

et al. 2018

Blood Advances

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• Soluble GPVI is elevated in patients with thermal injury with sepsis, and sGPVI levels augment severity score prediction of mortality.• The GPVI ligand, fibrin, induces GPVI shedding without requirement for platelet activation or signaling Soluble glycoprotein VI (sGPVI) is shed from the platelet surface and is a marker of platelet activation in thrombotic conditions. We assessed sGPVI levels together with patient and clinical parameters in acute and chronic inflammatory conditions, including patients with thermal injury and inflammatory bowel disease and patients admitted to the intensive care unit (ICU)for elective cardiac surgery, trauma, acute brain injury, or prolonged ventilation. Plasma sGPVI was measured by enzyme-linked immunosorbent assay and was elevated on day 14 after thermal injury, and was higher in patients who developed sepsis. sGPVI levels were associated with sepsis, and the value for predicting sepsis was increased in combination with platelet count and Abbreviated Burn Severity Index. sGPVI levels positively correlated with levels of D-dimer (a fibrin degradation product) in ICU patients and patients with thermal injury. sGPVI levels in ICU patients at admission were significantly associated with 28-and 90-day mortality independent of platelet count. sGPVI levels in patients with thermal injury were associated with 28-day mortality at days 1, 14, and 21 when adjusting for platelet count. In both cohorts, sGPVI associations with mortality were stronger than D-dimer levels. Mechanistically, release of GPVI was triggered by exposure of platelets to polymerized fibrin, but not by engagement of G protein-coupled receptors by thrombin, adenosine 59-diphosphate, or thromboxane mimetics. Enhanced fibrin production in these patients may therefore contribute to the observed elevated sGPVI levels. sGPVI is an important platelet-specific marker for platelet activation that predicts sepsis progression and mortality in injured patients.

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“…Due to low diagnostic sensitivity and technical requirements, the determination of pGPVI expression with flow cytometry also does not seem to be feasible in clinical routine. Previously, we developed a sandwich immunoassay for a reliable measurement of sGPVI, which has been successfully validated in a large group of patients with symptomatic coronary artery disease (CAD) [13]. In this study, we examined the levels of sGPVI using our novel sandwich immunoassay in patients showing symptoms of acute ischemic stroke.…”

Section: Introductionmentioning

confidence: 99%

Plasma levels of soluble glycoprotein VI (sGPVI) are associated with ischemic stroke

et al. 2012

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Platelet collagen receptor glycoprotein VI (pGPVI) is elevated in patients with acute coronary syndrome (ACS) and ischemic stroke. Recently, we developed a novel bead-based sandwich immunoassay to determine soluble GPVI (sGPVI), which has been validated in ACS patients. This study aimed to evaluate the plasma levels of sGPVI and pGPVI expression in patients with suspected stroke. We consecutively evaluated 176 patients, who were admitted to the stroke unit. Surface expression of pGPVI was determined by flow cytometry, sGPVI concentrations were determined using our sandwich immunoassay. Unlike patients with TIA, patients with stroke showed significantly decreased plasma levels of sGPVI compared to patients with non-ischemic (NI) events (TIA: mean [µg/L] ± standard deviation): 6.1 ± 2.1 vs. NI: 8 ± 4; p = 0.192; stroke: 5.9 ± 2.3 vs. NI; p = 0.013), whereas for pGPVI, patients with TIA and ischemic stroke revealed a significantly increased platelet surface expression compared to NI patients (TIA: mean fluorescence intensity [MFI] ± standard deviation): 20.9 ± 5.4 vs. NI: 17.6 ± 5.2; p = 0.021; stroke: 20.3 ± 6.2 vs. NI; p = 0.016). Using logistic regression analysis, both sGPVI (p = 0.002) and pGPVI (p = 0.012) are independently associated with ischemic stroke compared to other laboratory markers. To predict the individual risk for ischemic stroke using the plasma levels of sGPVI, receiver operating characteristic (ROC) analysis determined an optimal cutoff value of sGPVI at 6.5 µg/l, thus, patients with decreased plasma levels (<6.5 µg/l) have a 1.5-fold adjusted odds ratio (95%confidence interval, 1.4-2.7). Lower plasma levels of sGPVI are associated with the slightly elevated risk of stroke and may be a promising novel biomarker.

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Sandwich Immunoassay for Soluble Glycoprotein VI in Patients with Symptomatic Coronary Artery Disease

Abstract: BACKGROUND: Platelet glycoprotein VI (pGPVI) expression is increased in acute coronary syndrome (ACS), reflecting platelet activation. There is no reliable method available to measure pGPVI. Our aim was to develop a bead-based sandwich immunoassay to measure soluble GPVI (sGPVI).

Cited by 25 publications

References 31 publications

Pathologic shear triggers shedding of vascular receptors: a novel mechanism for down-regulation of platelet glycoprotein VI in stenosed coronary vessels

Pathologic shear triggers shedding of vascular receptors: a novel mechanism for down-regulation of platelet glycoprotein VI in stenosed coronary vessels

Soluble GPVI is elevated in injured patients: shedding is mediated by fibrin activation of GPVI

Plasma levels of soluble glycoprotein VI (sGPVI) are associated with ischemic stroke

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