scholarly journals Uncomplicated Plasmodium vivax malaria: Mapping the proteome from circulating platelets

2021 ◽  
Author(s):  
Diana Fernández ◽  
Cesar Segura ◽  
Mònica Arman ◽  
Suzzane McGill ◽  
Richard Burchmore ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Platelet Activation ◽  
Plasma Levels ◽  
Vivax Malaria ◽  
Endothelial Activation ◽  
Molecular Pathways ◽  
Healthy Controls ◽  
Cytoskeletal Remodeling ◽  
Plasmodium Vivax Malaria ◽  
Platelet Proteome

Abstract Background: Thrombocytopenia is frequent in uncomplicated Plasmodium vivax malaria. Contribution of platelets to pathogenesis is unknown and poorly understood. Our study explores the platelet proteome from uncomplicated P. vivax malaria patients to fingerprint molecular pathways in relation to platelet function. Also, plasma levels of platelet activation (Platelet factor 4 – PF4/CXCL4) and endothelial activation (Von Willebrand factor – VWf) markers, in conjunction with some in vitro interactions between platelets and P. vivax infected erythrocytes ( Pv -IEs) were measured to explore platelet responses during infection and their effect on parasite development. Methods: This study was performed in a cohort of 48 patients and 25 healthy controls. Platelets were purified from a subgroup of 5 patients and 5 healthy controls to be analyzed by LC-MS/MS. In all participants enrolled in this study, PF4/CXCL4 and VWf plasma levels were measured. Finally, a subsample of 10 P. vivax isolates were co-cultured with platelets to measure P v- IE schizonts inhibition as well as platelet activation due to their interaction. Results: In total 28 out of 215 proteins were significantly abundant in the proteomes from patients. The most significantly decreased protein was PF4/CXCL4 followed by other proteins related to platelet activation, cytoskeletal remodeling, and adhesion to endothelial cells. In contrast, acute phase proteins including SERPINs and Amyloid Serum A 1 (SAA1) were increased. High VWf plasma levels in patients suggested endothelial activation. Interestingly, PF4/CXCL4 plasma levels were similar between patients and controls, but high levels of this protein were found in co-cultures, and platelets inhibited Pv -IEs development to schizonts. Conclusions: Platelet proteome from patients with uncomplicated P. vivax malaria suggests platelet degranulation, platelet activation, cytoskeletal remodeling, and adhesion to endothelial cells. According to the evidenced endothelial activation our study plus the suggested specific localization of PF4/CXCL4 during P. vivax infection due to the normal levels in plasma, and the inhibition of Pv -IE schizonts development; our study suggest that platelets are active players during the response to P . vi vax infection. Future studies are needed to further investigate the molecular pathways of interaction between altered platelet proteins and host response; which could affect parasite control as well as disease progression.

scholarly journals Uncomplicated Plasmodium vivax malaria: mapping the proteome from circulating platelets

2022 ◽  
Vol 19 (1) ◽  
Author(s):  
Diana Fernández ◽  
Cesar Segura ◽  
Mònica Arman ◽  
Suzanne McGill ◽  
Richard Burchmore ◽  
...  
Keyword(s):  
Plasmodium Vivax ◽  
Plasma Levels ◽  
Acute Phase Proteins ◽  
Intracellular Localization ◽  
Endothelial Activation ◽  
Parasite Development ◽  
Molecular Pathways ◽  
Healthy Controls ◽  
Platelet Factor

Abstract Background Thrombocytopenia is frequent in Plasmodium vivax malaria but the role of platelets in pathogenesis is unknown. Our study explores the platelet (PLT) proteome from uncomplicated P. vivax patients, to fingerprint molecular pathways related to platelet function. Plasma levels of Platelet factor 4 (PF4/CXCL4) and Von Willebrand factor (VWf), as well as in vitro PLTs—P. vivax infected erythrocytes (Pv-IEs) interactions were also evaluated to explore the PLT response and effect on parasite development. Methods A cohort of 48 patients and 25 healthy controls were enrolled. PLTs were purified from 5 patients and 5 healthy controls for Liquid Chromatography–Mass spectrometry (LC–MS/MS) analysis. Plasma levels of PF4/CXCL4 and VWf were measured in all participants. Additionally, P. vivax isolates (n = 10) were co-cultured with PLTs to measure PLT activation by PF4/CXCL4 and Pv-IE schizonts formation by light microscopy. Results The proteome from uncomplicated P. vivax patients showed 26 out of 215 proteins significantly decreased. PF4/CXCL4 was significantly decreased followed by other proteins involved in platelet activation, cytoskeletal remodeling, and endothelial adhesion, including glycoprotein V that was significantly decreased in thrombocytopenic patients. In contrast, acute phase proteins, including SERPINs and Amyloid Serum A1 were increased. High levels of VWf in plasma from patients suggested endothelial activation while PF4/CXCL4 plasma levels were similar between patients and controls. Interestingly, high levels of PF4/CXCL4 were released from PLTs—Pv-IEs co-cultures while Pv-IEs schizont formation was inhibited. Conclusions The PLT proteome analyzed in this study suggests that PLTs actively respond to P. vivax infection. Altogether, our findings suggest important roles of PF4/CXCL4 during uncomplicated P. vivax infection through a possible intracellular localization. Our study shows that platelets are active responders to P. vivax infection, inhibiting intraerythrocytic parasite development. Future studies are needed to further investigate the molecular pathways of interaction between platelet proteins found in this study and host response, which could affect parasite control as well as disease progression.

Homozysteinspiegel von Patienten mit Raynaud-Phänomen

VASA ◽  
2002 ◽  
Vol 31 (2) ◽  
pp. 87-90 ◽  
Author(s):  
Al-Awami ◽  
Schillinger ◽  
Maca ◽  
Gschwandtner ◽  
Bieglmayer ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Plasma Levels ◽  
Serum Levels ◽  
Plasma Homocysteine ◽  
Serum Folate ◽  
Vitamin B ◽  
Healthy Controls ◽  
Healthy Individuals ◽  
Homocysteine Concentration ◽  
Age And Sex

Background: Patients with Raynaud’s phenomenon (RP) have vasomotor dysregulation, mainly caused by dysfunction of the endothelium. Since homocysteine has been found to be damaging to endothelial cells, we investigated the concentrations of plasma homocysteine, folate and vitamin B12 in patients with primary or secondary RP compared to healthy individuals. Patients and methods: We measured the concentrations of plasma fasting homocysteine, folate and vitamin B12 in a group of healthy individuals (n = 45) and in patients with primary (n = 26) or secondary RP (n = 42). Results: Median homocysteine levels in healthy controls and in patients with primary RP, secondary RP were 7.9 (IQR 4.1 to 11.8) 9.8 (IQR 5.1 to14.4), and 10.6 (6.0 to15.3) mumol/L, respectively. Patients with primary and secondary RP had significantly higher homocysteine concentration compared to healthy controls (Kruskal Wallis p = 0.01). After matching for age and sex, patients with either primary or secondary RP showed significantly higher homocysteine levels (Wilcoxon p < 0.0001). No significant differences between the three groups were found concerning serum levels of vitamin B12 (p = 0.9 ) and serum folate levels (p = 0.2). Conclusion: These data demonstrate that patients with RP have higher plasma levels of homocysteine. No significant differences in folate and vitamin B12 levels were found between patients with primary RP, secondary RP, and healthy individuals.These data suggest that homocysteine may play a role in RP and may provide new clues in understanding of the vasomotor dysregulation.

scholarly journals ELEVATED PLASMA LEVELS OF TNF-ALPHA, INF-GAMMA, IL-10 AND TGF-BETA IN MALARIA PATIENTS FROM TWO MALARIA NON-ENDEMIC REGIONS IN KARNATAKA, INDIA

2020 ◽  
Vol 4 (2) ◽  
Author(s):  
Mukthayakka G ◽  
Annapurna G Sajjan ◽  
Ragini Ananth Kashid  ◽  
Mohd Shannawaz ◽  
Thejaswini  HS ◽  
...  
Keyword(s):  
Plasmodium Vivax ◽  
Plasma Levels ◽  
Vivax Malaria ◽  
Infection Type ◽  
Receiver Operating Curve ◽  
Research Centre ◽  
Elevated Plasma ◽  
Tnf Α ◽  
Plasmodium Vivax Malaria ◽  
Ifn Γ

Purpose: In India, Plasmodium vivax malaria is endemic and accounts for 50-55% of the total malaria burden in the country. There has been limited sero-epidemiological data available from malaria non-endemic regions in Karnataka state. In this study, we aimed to evaluate the plasma levels of Tumor necrosis factor-α (TNF-α), Interferon- γ (IFN-γ), Interleukin-10 (IL-10), and Transforming growth factor- β (TGF-β) and correlate with malaria parasitaemia and infection type in vivax and falciparum malaria cases reported from two study centres. Methods: This hospital-based cross sectional observational study was conducted at BLDEU SHRI B.M. Patil Medical College, Hospital and Research Centre Vijayapur, Karnataka and BGS Global Institute of Medical Sciences, Bengaluru, Karnataka during 2016 to 2019. A total number of 45 microscopy positive and molecularly confirmed malaria cases were included in the study. Plasma samples were analyzed for the concentrations of four cytokines by Enzyme-linked immunosorbent assay (ELISA). 20 uninfected healthy volunteers were used as controls. Correlation of cytokines and parasitemia was done using Pearson correlation analysis. Results: The results show an overall significant elevation of plasma TNF-α (p<0.05), IFN-γ (p<0.005), IL-10 (p<0.001), and TGF-β (p<0.001) in malaria patients compared to healthy controls. Except TNF-α (p<0.001), there was no significant difference in infection type specific immune responses. No significant correlation was seen among all the four cytokines with parasite load. A Receiver operating curve (ROC) was generated and showed that TNF-α, IL-10, and IFN-γ were the best individual predictors of malaria. Conclusions: We conclude that significantly elevated plasma concentrations of TNF-α-, IL-10, IFN-γ and TGF-β in both P. vivax and P. falciparum cases suggest their active involvement in mounting defensive immune response against malaria infection. Keywords: Malaria, Plasmodium vivax malaria, TNF-α, INF-γ, IL-10, TGF-β, Karnataka

scholarly journals Association of Endothelial Biomarkers with Cerebral MRI Perfusion Analysis in Patients with Sickle Cell Disease

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1000-1000
Author(s):  
Liza Afzali-Hashemi ◽  
Lena Vaclavu ◽  
Erfan Nur ◽  
Aart J Nederveen ◽  
Bart J. Biemond
Keyword(s):  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Adhesion Molecule ◽  
Plasma Levels ◽  
Cerebral Perfusion ◽  
Endothelial Activation ◽  
Healthy Controls ◽  
Standard Laboratory ◽  
Cell Disease ◽  
Laboratory Parameters

Introduction Sickle cell disease (SCD) is associated with silent cerebral infarcts (SCI) which are related to neurocognitive damage. One of the major causes of SCI is the impaired cerebral oxygenation, which makes cerebral blood flow (CBF) an essential parameter to measure. Previous hemodynamic studies have shown elevated CBF and reduced cerebrovascular reserve (CVR) in patients with SCD (Helton 2015; Václavů 2018). CVR is known as the increase of CBF in response to vasoactive stimulus, relative to the baseline. The reduced CVR renders SCD patients susceptible to cerebral ischemia, especially under hypotensive or hypoxic conditions. Possible factors contributing to microvascular damage and thus reduced CVR include hemoglobin S polymerization, neutrophil activation and endothelial activation and adhesion. Previous studies examined the role of these factors in patients with SCD (Sins 2017; Al Najjar 2017). However, the association between the endothelial biomarkers and hemodynamic parameters is unknown in adult patients with SCD. In this study, we investigated the correlation between CBF and CVR and the adhesion molecules including sVCAM-1, sP-selectin, VWF-Ag and ADAMTS13. Additionally, we studied the association of these endothelial biomarkers with standard laboratory parameters. Methods This study was performed in accordance with the Declaration of Helsinki and was approved by the Review Board of Amsterdam UMC. For this study, 33 steady state patients with SCD (mean age 32.1 ± 10.7, 64% male, 29 HbSS, 4 HbSß) and 10 healthy volunteers (mean age 36,4 ± 15.9, 60% male, 2 HbAS, 8 HbAA) were included. Hematologic laboratory parameters were assessed using standard laboratory procedures. Plasma levels of sVCAM-1 and sP-selectin were determined using ELISA (R&D Systems, USA) and ADAMTS13 and VWF-Ag were measured with the INNOVANCE assay (Siemens Healthcare Diagnostics). For the CBF measurements, pseudo-continuous arterial spin labelling (pCASL) was acquired at 3T MRI (Philips Healthcare, The Netherlands). CBF was measured before and after acetazolamide (vasoactive stimulus) administration and subsequently CVR was calculated using the following equation: CVR = (CBFafter - CBFbefore) / CBFbefore x 100% Plasma levels of endothelial biomarkers were compared between groups using ANOVA test. Correlation between the parameters were measured using single regression model where p<0.05 was considered as statistically significant. Results sVCAM-1 levels and VWF-Ag were significantly higher in SCD patients compared to healthy controls (p < 0.01 and p = 0.01). ADAMTS13 and sP-selectin were not significantly different between the two groups (p = 0.06 and p = 0.33). sVCAM-1 was significantly associated with CBF, and parameters of hemolysis LDH and bilirubin in SCD patients (Fig. 1A and 2C). Negative correlation was observed between sVCAM-1 and hemoglobin (Fig. 1B). The relationship between sVCAM-1 and hemodynamic and laboratory parameters are shown in Table 1. No significant correlation was found between sVCAM-1, hemodynamic and standard laboratory parameters in healthy controls. VWF-Ag, ADAMTS13 and sP-selectin were not significantly associated with hemodynamic MRI parameters. Discussion Our results show elevated sVCAM-1 levels in sickle cell patients, strongly related to CBF. sVCAM-1 is an adhesion molecule and elevated plasma levels are found in patients with endothelial activation due to inflammation or atherosclerosis (Cook-Mills 2013; Cybulsky 2001). Previous studies showed elevated levels in sickle cell disease but no correlation with parameters of cerebral perfusion have been demonstrated yet (Antwi-Boasiako 2018; Kato 2005). The strong correlation with CBF is mostly related to the chronic hemolysis given the association found with hemoglobin levels and markers of hemolysis like LDH and bilirubin. In contrast to sVCAM-1, no such relation was found with other markers of endothelial activation such as VWF activity and sP-selectin levels. No correlation between endothelial markers and the CVR was demonstrated, suggesting that endothelial damage itself may not related to the impaired cerebral vasodilatation in response to a vasoactive stimulus. Conclusion Endothelial adhesion molecule sVCAM-1 showed a strong correlation with CBF and parameters of hemolysis, suggesting a relation between the hemolytic damage of endothelial cells and impaired cerebral perfusion. Disclosures Nur: Novartis Pharmaceuticals: Consultancy.

scholarly journals Nitric Oxide–Dependent Endothelial Dysfunction and Reduced Arginine Bioavailability in Plasmodium vivax Malaria but No Greater Increase in Intravascular Hemolysis in Severe Disease

2016 ◽  
Vol 214 (10) ◽  
pp. 1557-1564 ◽  
Author(s):  
Bridget E. Barber ◽  
Timothy William ◽  
Matthew J. Grigg ◽  
Kim A. Piera ◽  
Youwei Chen ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Dysfunction ◽  
Endothelial Function ◽  
Plasmodium Vivax ◽  
Vivax Malaria ◽  
Severe Disease ◽  
Healthy Controls ◽  
Intravascular Hemolysis ◽  
Plasmodium Vivax Malaria ◽  
Arginine Level

Background Pathogenesis of severe Plasmodium vivax malaria is poorly understood. Endothelial dysfunction and reduced nitric oxide (NO) bioavailability characterize severe falciparum malaria, but have not been assessed in severe vivax malaria. Methods In patients with severe vivax malaria (n = 9), patients with nonsevere vivax malaria (n = 58), and healthy controls (n = 79), we measured NO-dependent endothelial function by using reactive hyperemia–peripheral arterial tonometry (RH-PAT) and assessed associations with arginine, asymmetric dimethylarginine (ADMA), and hemolysis. Results The L-arginine level and the L-arginine to ADMA ratio (a measure of L-arginine bioavailability) were reduced in patients with severe vivax malaria and those with nonsevere vivax malaria, compared with healthy controls (median L-arginine level, 65, 66, and 98 µmol/mL, respectively [P = .0001]; median L-arginine to ADMA ratio, 115, 125, and 187, respectively [P = .0001]). Endothelial function was impaired in proportion to disease severity (median RH-PAT index, 1.49, 1.73, and 1.97 in patients with severe vivax malaria, those with nonsevere vivax malaria, and healthy controls, respectively; P = .018) and was associated with the L-arginine to ADMA ratio. While the posttreatment fall in hemoglobin level was greater in severe vivax malaria as compared to nonsevere vivax malaria (2.5 vs 1 g/dL; P = .0001), markers of intravascular hemolysis were not higher in severe disease. Conclusions Endothelial function is impaired in nonsevere and severe vivax malaria, is associated with reduced L-arginine bioavailability, and may contribute to microvascular pathogenesis. Severe disease appears to be more associated with extravascular hemolysis than with intravascular hemolysis.

Increased Circulating Nucleosomes and Neutrophil Activation As a Measure of the Formation of Neutrophil Extracellular Traps (NETs) During Sickle Cell Painful Crisis

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 821-821
Author(s):  
Marein Schimmel ◽  
Erfan Nur ◽  
Sacha Zeerleder ◽  
Gerard J van Mierlo ◽  
Shabnam Solati ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Steady State ◽  
Sickle Cell ◽  
Plasma Levels ◽  
Neutrophil Extracellular Traps ◽  
Neutrophil Activation ◽  
Acute Chest Syndrome ◽  
Healthy Controls ◽  
Extracellular Traps ◽  
Painful Crisis

Abstract Abstract 821 Introduction: Sickle cell disease (SCD) is characterized by recurrent acute vaso-occlusive painful crisis frequently leading to SCD related complications such as acute chest syndrome, stroke, multi-organ failure and even sudden death. The complex pathophysiology of the vaso-occlusive painful crisis is mediated by activation of endothelial cells, adhesion of sickled erythrocytes and neutrophils, oxidative stress, coagulation activation and increased release of inflammatory mediators, resulting in ischemic organ damage. Recently, neutrophils have been demonstrated to form neutrophil extracellular traps (NETs) upon activation. Nucleosomes and histones exposed together with neutrophil proteases, such as elastase on these NETs have been shown to kill efficiently bacteria. NET formation has been shown to propagate coagulation in sepsis and in deep venous thrombosis. In addition, nucleosomes and histones exposed on NETs have been shown to be strongly cytotoxic to endothelial cells. Beside the exposure on NETs, nucleosomes can be actively released into the circulation from dead cells. Circulating nucleosomes detected in sepsis have been reported to correlate with severity of inflammation, organ dysfunction and mortality. However, no studies are available yet on the dynamics of nucleosomes and NETs in sickle cell patients suffering from painful crisis. The aim of this case-control study was to assess plasma levels of circulating nucleosomes and human neutrophil elastase–α1-antitrypsin (EA) complexes as measure of systemic neutrophil activation, in sickle cell patients during steady state and painful crisis. Methods: Plasma levels of nucleosomes and EA as a measure of neutrophil activation were measured in 74 patients in asymptomatic state (49 HbSS/HbSβ0-thalassemia, and 25 HbSC/HbSβ+-thalassemia), 70 painful crises (53 HbSS/HbSβ°-thalassemia and 17 HbSC/HbSβ+-thalassemia) in 49 patients and in 24 HbAA healthy controls using Enzyme-Linked Immunosorbent Assay (ELISA). Results: Plasma levels of nucleosomes in both HbSS/HbSβ°-thalassemia and HbSC/HbSβ+-thalassemia patients were significantly higher during painful crisis (median; interquartile range, 20.2; 8.9 – 129.0 U/ml, P < 0.0001 and 11.7; 5.1 – 67.7 U/ml, P = 0.045 respectively) as compared to patients in steady state (6.0; 3.0 – 9.8 U/ml and 7.1; 4.6 – 9.6 U/ml respectively). Nucleosomes levels in healthy controls were just above the detection limit of the assay (5.0; 5.0 – 6.5) U/ml). Plasma levels of EA in HbSS/HbSβ°-thalassemia patients were significantly increased during painful crisis as compared to steady state (75.1; 56.5 – 102.4 vs. 45.7; 34.7 – 59.7 ng/ml, P < 0.0001). Also in HbSC/HbSβ+-thalassemia patients, EA levels were higher during painful crisis than in steady state, though the difference did not reach statistical significance (62.0; 48.0 – 96.7 vs. 50.2; 33.3 – 67.7, P = 0.051). Plasma levels of EA in healthy controls (39.9; 31.5 – 62.2 ng/ml) were comparable with those in steady state patients. In a paired analysis of 36 patients, included both during steady state and painful crisis, significant increments were observed during painful crisis in levels of both nucleosomes (from 5.0; 3.0 – 10.8 to 20.2; 6.8 – 94.3 U/ml, P < 0.0001) and EA (from 47.9; 36.0 – 67.6 to 70.6; 55.9 – 101.4 ng/ml, P < 0.0001), as compared to steady state. During painful crisis, EA levels were strongly correlated with levels of nucleosomes in both HbSS/HbSβ°-thalassemia (Spearman's rank (Sr)=0.55, P<0.0001) and HbSC/HbSβ+-thalassemia patients (Sr=0.90, P=<0.0001). In steady state the correlation was significant only in HbSC/HbSβ+-thalassemia patients (Sr=0.63, P=0.001) Four patients who developed an acute chest syndrome during painful crisis were among the patients with the highest nucleosome (359, 130, 128 and 100 U/ml) and EA levels (121, 87, 92 and 64 ng/ml respectively). Conclusion: Sickle cell painful crisis is associated with increased levels of nucleosome and stronger neutrophil activation. This might point to a crucial role of NET formation in the pathogenesis of painful crisis. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Endothelial glycocalyx degradation and disease severity in Plasmodium vivax and Plasmodium knowlesi malaria

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Bridget E. Barber ◽  
Matthew J. Grigg ◽  
Kim A. Piera ◽  
Youwei Chen ◽  
Timothy William ◽  
...  
Keyword(s):  
Falciparum Malaria ◽  
Disease Severity ◽  
Vivax Malaria ◽  
Severe Disease ◽  
Kidney Injury ◽  
Endothelial Activation ◽  
Endothelial Glycocalyx ◽  
Healthy Controls ◽  
Angiopoietin 2 ◽  
Urinary Glycosaminoglycans

AbstractDegradation of the endothelial glycocalyx is associated with mortality in adult falciparum malaria. However, its role in the pathogenesis of non-falciparum malaria is unknown. In Malaysian patients with knowlesi (n = 200) and vivax (n = 61) malaria, and in healthy controls (n = 50), we measured glycocalyx breakdown products plasma syndecan-1 and urinary glycosaminoglycans, and evaluated correlations with biomarkers of disease severity. Urinary glycosaminoglycans were increased in patients with knowlesi and vivax malaria compared to healthy controls, and in knowlesi malaria were highest in those with severe disease. In knowlesi malaria, plasma syndecan-1 was also highest in those with severe disease, and correlated with markers of endothelial activation (angiopoietin-2, osteoprotegerin, ICAM-1), asymmetric dimethylarginine (ADMA) and impaired microvascular reactivity. Syndecan-1 also correlated with endothelial activation (ICAM-1, angiopoietin-2) and ADMA in vivax malaria. In knowlesi malaria increased syndecan-1 was associated with acute kidney injury, after controlling for age and parasitemia. In knowlesi malaria, the difference in median syndecan-1 between severe and non-severe disease was more marked in females than males. Endothelial glycocalyx degradation is increased in knowlesi and vivax malaria, and associated with disease severity and acute kidney injury in knowlesi malaria. Agents that inhibit glycocalyx breakdown may represent adjunctive therapeutics for severe non-falciparum malaria.

scholarly journals Increased Phosphatidylserine-Exposing Microparticles and Their Originating Cells Are Associated with the Coagulation Process in Patients with Oral Squamous Cell Carcinoma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1222-1222
Author(s):  
Yingmiao Liu ◽  
Cong Zhang ◽  
Yan Kou ◽  
Lili Zou ◽  
Hui Liang ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Squamous Cell Carcinoma ◽  
Endothelial Cells ◽  
Cell Carcinoma ◽  
Binding Sites ◽  
Blood Cells ◽  
Plasma Levels ◽  
Stage Iv ◽  
Radical Resection ◽  
Healthy Controls

Abstract Introduction:Oral squamous cell carcinoma (OSCC) is the most common cancer of the head and neck area, and the incidence remains high.Despite advances in diagnosis and treatment, the poor prognosis of OSCC is characterized by a high rate of local recurrence and the overall five-year survival rate remains at approximately 50%. Therefore, the mechanisms underlying the development of OSCC still need to be clarified. Patients with cancer tend to develop a hypercoagulable state which predisposes them to thromboembolic events. Cancer increases the risk of venous thrombosis several fold with varying degree of relative risks (range 4-7). A recent study has reported that microparticles (MPs) increased procoagulant activity (PCA) in OSCC. MPs are small membrane vesicles of 0.1-1 µm containing negatively charged, procoagulant phosphatidylserine (PS), which plays an important role in thrombosis. The definitive role of PS in the hypercoagulable state in patients with OSCC remains unclear. Our objectives were to measure the PS exposure on MPs, blood cells, and endothelium, and to evaluate their PCA in different stages of OSCC. Methods: OSCC patients (n = 57) and healthy controls (n = 26) were included in our study. Blood samples were obtained from controls and OSCC patients within 1 day before surgery and 2-week after surgery. Human umbilical vein endothelial cells (HUVECs) were incubated in growth media containing 20% of pool serum obtained from either OSCC patients or healthy donors at room temperature for 24 h, respectively. Exposed PS was analyzed with flow cytometry and confocal microscopy. Lactadherin was used to quantify PS exposure on MPs and their original cells. PCA of MPs and these cells was evaluated using clotting time, purified coagulation complex, and fibrin formation assays. Meanwhile, the inflammation-related cytokines were detected by enzyme-linked immunosorbent assay. Results: Using flow cytometry, plasma levels of PS+ blood cells and MPs in OSCC patients with stage III/IV were significantly higher than those in stage I/II patients or healthy controls (all P < 0.05). However, we only found a significant difference between stage I or II and controls (P < 0.05) in total PS+ MPs and PMPs. Similarly, we found that the endothelial cells (ECs) treated with OSCC serum in vitro exposed more PS than those with healthy serum. Moreover, in OSCC patients with stage IV, MPs primarily originated from platelets (53.9 ± 3.2%) followed by leukocytes (21.8 ± 2.1%, including MPs from neutrophils, monocytes and lymphocytes), erythrocytes (6.3 ± 0.6%) and ECs (6.9 ± 0.8%). Additionally, PS+ blood cells, MPs and OSCC serum-cultured ECs markedly promoted shortened coagulation time and significantly increased FXa/thrombin/fibrin generation in stage IV or III OSCC patients compared with controls (all P < 0.05). Interestingly, confocal imaging of MPs or OSCC serum-treated ECs showed binding sites for FVa and FXa to form prothrombinase. Furthermore, blockade of PS on MPs/blood cells/ECs with lactadherin inhibited PCA by approximately 80%. Most importantly, we found treatment with radical resection significantly reduced the amount of PS+ blood cells, ECs and MPs, and prolonged the clotting times of those cells and MPs compared with presurgery patients. Lastly, the correlation analysis revealed that the plasma levels of interleukin 6, interleukin 8 and tumor necrosis factor α were positively correlated with the levels of total PS+ MPs and PS+ platelets in OSCC patients. Conclusions: Our results suggested that PS+ blood cells and MPs play a prominent role in inducing the hypercoagulable and prothrombotic state especially in advanced OSCC. Interestingly, we found treatment with radical resection could attenuate this effect. Moreover, the released inflammatory cytokines may contribute to PS exposure on platelets and MPs and the increased procoagulant activity in patients with OSCC. Notably, our findings also show that PS provides binding sites for FXa and prothrombinase complexes and promotes thrombin formation. Therefore, directly targeting FXa and prothrombinase complexes might decrease thrombotic risk OSCC patients. This study shows that future research should focus on the application of PS inhibitors as a novel therapeutic strategy in OSCC patients when coagulation is abnormally enhanced. Disclosures No relevant conflicts of interest to declare.

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