Plasminogen Depletion during Streptokinase Treatment or Two-Chain Urokinase Incubation Correlates with Decreased Clot Lysability Ex Vivo and In Vitro

SummaryThe relationship between lytic state variables and ex vivo clot lysability was investigated in blood drawn from patients during streptokinase administration for acute myocardial infarction. A lytic state was already evident after 5 min of treatment and after 20 min the plasminogen concentration had decreased to 24%, antiplasmin to 7% and fibrinogen 0.2 g/1. Lysis of radiolabeled retracted clots in the patient plasmas decreased from 37 ± 8% after 5 min to 21 ± 8% at 10 min and was significantly lower (8 ± 9%, p <0.005) in samples drawn at 20, 40 and 80 min. Clot lysability correlated positively with the plasminogen concentration (r = 0.78, p = 0.003), but not with plasmin activity. Suspension of radiolabeled clots in normal plasma pre-exposed to 250 U/ml two-chain urokinase for varying time to induce an in vitro lytic state was also associated with decreasing clot lysability in direct proportion with the duration of prior plasma exposure to urokinase. The decreased lysability correlated with the time-dependent reduction in plasminogen concentration (r = 0.88, p <0.0005). Thus, clot lysability decreases in conjunction with the development of the lytic state and the associated plasminogen depletion. The lytic state may therefore limit reperfusion during thrombolytic treatment.

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Monitoring Anticoagulant Therapy by Activated Partial Thromboplastin Time: Hirudin Assessment

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648943 ◽

1994 ◽

Vol 72 (05) ◽

pp. 685-692 ◽

Cited By ~ 43

Author(s):

Michael T Nurmohamed ◽

René J Berckmans ◽

Willy M Morriën-Salomons ◽

Fenny Berends ◽

Daan W Hommes ◽

...

Keyword(s):

Whole Blood ◽

Partial Thromboplastin Time ◽

Ex Vivo ◽

Fold Increase ◽

Heparin Therapy ◽

Activated Partial Thromboplastin Time ◽

Recombinant Hirudin ◽

Interindividual Differences ◽

The Relationship

SummaryBackground. Recombinant hirudin (RH) is a new anticoagulant for prophylaxis and treatment of venous and arterial thrombosis. To which extent the activated partial thromboplastin time (APTT) is suitable for monitoring of RH has not been properly evaluated. Recently, a capillary whole blood device was developed for bed-side monitoring of the APTT and it was demonstrated that this device was suitable to monitor heparin therapy. However, monitoring of RH was not evaluated.Study Objectives. To evaluate in vitro and ex vivo the responsiveness and reproducibility for hirudin monitoring of the whole blood monitor and of plasma APTT assays, which were performed with several reagents and two conventional coagulometers.Results. Large interindividual differences in hirudin responsiveness were noted in both the in vitro and the ex vivo experiments. The relationship between the APTT, expressed as clotting time or ratio of initial and prolonged APTT, and the hirudin concentration was nonlinear. A 1.5-fold increase of the clotting times was obtained at 150-200 ng/ml plasma. However, only a 2-fold increase was obtained at hirudin levels varying from 300 ng to more than 750 ng RH/ml plasma regardless of the assays. The relationship linearized upon logarithmic conversion of the ratio and the hirudin concentration. Disregarding the interindividual differences, and presuming full linearity of the relationship, all combinations were equally responsive to hirudin.Conclusions. All assays were equally responsive to hirudin. Levels up to 300 ng/ml plasma can be reliably estimated with each assay. The manual device may be preferable in situations where rapid availability of test results is necessary.

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Neutrophil extracellular traps induce MCP-1 release from endothelial cells at the plaque rupture site in acute myocardial infarction

European Heart Journal ◽

10.1093/ehjci/ehaa946.3834 ◽

2020 ◽

Vol 41 (Supplement_2) ◽

Author(s):

A Ondracek ◽

T.M Hofbauer ◽

A Mangold ◽

T Scherz ◽

V Seidl ◽

...

Keyword(s):

Myocardial Infarction ◽

Acute Myocardial Infarction ◽

Endothelial Cells ◽

Plaque Rupture ◽

Ex Vivo ◽

Coronary Intervention ◽

Neutrophil Extracellular Traps ◽

Funding Source ◽

Extracellular Traps

Abstract Introduction Leukocyte-mediated inflammation is crucial in acute myocardial infarction (AMI). We recently observed that neutrophil extracellular traps (NETs) are increased at the culprit site, promoting activation and differentiation of fibrocytes, cells with mesenchymal and leukocytic properties. Fibrocyte migration is mediated by monocyte chemoattractant protein (MCP)-1 and C-C chemokine receptor type 2 (CCR2). We investigated the interplay between NETs, fibrocyte function, and MCP-1 in AMI. Methods Culprit site and femoral blood of AMI patients was drawn during percutaneous coronary intervention. We characterized CCR2 expression of fibrocytes by flow cytometry. MCP-1 and the NET marker citrullinated histone H3 (citH3) were measured by ELISA. Fibrocytes were treated in vitro with MCP-1. Human coronary arterial endothelial cells (hCAECs) were stimulated with isolated NETs, and MCP-1 was measured by ELISA and qPCR. The influence of MCP-1 on NET formation in vitro was assessed using isolated neutrophils. Results We have included 50 consecutive AMI patients into the study. NETs and concentrations of MCP-1 were increased at the CLS. NET stimulation of hCAECs induced MCP-1 on mRNA and protein level. Increasing MCP-1 gradient was associated with fibrocyte accumulation at the site of occlusion. In the presence of higher MCP-1 these fibrocytes expressed proportionally less CCR2 than peripheral fibrocytes. In vitro, MCP-1 dose-dependently decreased fibrocyte CCR2 and reduced ex vivo NET release of healthy donor neutrophils. Conclusions NETs induce endothelial MCP-1 release, presumably promoting a chemotactic gradient for leukocyte and fibrocyte migration. MCP-1 mediated inhibition of NET formation could point to a negative feedback loop. These data will shed light on vascular healing. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): Austrian Science Fund

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Considerations to Model Heart Disease in Women with Preeclampsia and Cardiovascular Disease

Cells ◽

10.3390/cells10040899 ◽

2021 ◽

Vol 10 (4) ◽

pp. 899

Author(s):

Clara Liu Chung Ming ◽

Kimberly Sesperez ◽

Eitan Ben-Sefer ◽

David Arpon ◽

Kristine McGrath ◽

...

Keyword(s):

Myocardial Infarction ◽

Cardiovascular Disease ◽

Ex Vivo ◽

Myocardial Damage ◽

Cardiovascular Disorder ◽

Model Systems ◽

Ex Vivo Model ◽

Disease Manifestation

Preeclampsia is a multifactorial cardiovascular disorder diagnosed after 20 weeks of gestation, and is the leading cause of death for both mothers and babies in pregnancy. The pathophysiology remains poorly understood due to the variability and unpredictability of disease manifestation when studied in animal models. After preeclampsia, both mothers and offspring have a higher risk of cardiovascular disease (CVD), including myocardial infarction or heart attack and heart failure (HF). Myocardial infarction is an acute myocardial damage that can be treated through reperfusion; however, this therapeutic approach leads to ischemic/reperfusion injury (IRI), often leading to HF. In this review, we compared the current in vivo, in vitro and ex vivo model systems used to study preeclampsia, IRI and HF. Future studies aiming at evaluating CVD in preeclampsia patients could benefit from novel models that better mimic the complex scenario described in this article.

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Considerations to Model Heart Disease in Women with Preeclampsia and Cardiovascular Disease

10.20944/preprints202104.0240.v1 ◽

2021 ◽

Author(s):

Clara Liu Chung Ming ◽

Kimberly Sesperez ◽

Eitan Ben-Sefer ◽

David Arpon ◽

Kristine McGrath ◽

...

Keyword(s):

Myocardial Infarction ◽

Cardiovascular Disease ◽

Ex Vivo ◽

Myocardial Damage ◽

Cardiovascular Disorder ◽

Model Systems ◽

Ex Vivo Model ◽

Disease Manifestation

Preeclampsia is a multifactorial cardiovascular disorder diagnosed after 20 weeks of gestation that is the leading cause of death for both mothers and babies in pregnancy. The pathophysiology remains poorly understood due to variability and unpredictability of disease manifestation when studied in animal models. After preeclampsia, both mothers and offspring have a higher risk of cardiovascular disease (CVD) including myocardial infarction or heart attack and heart failure (HF). Myocardial infarction is an acute myocardial damage that can be treated through reperfusion, however, that therapeutic approach leads to ischemic/reperfusion injury (IRI) often leading to HF. In this review, we compared the current in vivo, in vitro and ex vivo model systems used to study preeclampsia, IRI and HF. Future studies aiming at evaluating CVD in preeclampsia patients could benefit from novel models that better mimic the complex scenario described in this article.

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Ex vivo-expanded bone marrow CD34+ for acute myocardial infarction treatment: in vitro and in vivo studies

Cytotherapy ◽

10.3109/14653249.2011.597559 ◽

2011 ◽

Vol 13 (9) ◽

pp. 1140-1152 ◽

Cited By ~ 7

Author(s):

Monica Gunetti ◽

Alessio Noghero ◽

Fabiola Molla ◽

Lidia Irene Staszewsky ◽

Noeleen de Angelis ◽

...

Keyword(s):

Myocardial Infarction ◽

Acute Myocardial Infarction ◽

Bone Marrow ◽

Ex Vivo ◽

In Vivo Studies

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PPARβ/δ Priming Enhances the Anti-Apoptotic and Therapeutic Properties of Mesenchymal Stromal Cells in Myocardial Ischemia-Reperfusion Injury

10.21203/rs.3.rs-1139048/v1 ◽

2021 ◽

Author(s):

Charlotte Sarre ◽

Rafael Contreras Lopez ◽

Nitirut Nerpernpisooth ◽

Christian Barrere ◽

Sarah Bahraoui ◽

...

Keyword(s):

Oxidative Stress ◽

Myocardial Infarction ◽

Mesenchymal Stromal Cells ◽

Stromal Cells ◽

Ex Vivo ◽

Phase Iii ◽

Therapeutic Properties ◽

Cardioprotective Effects

Abstract Background: Mesenchymal Stromal Cells (MSC) have been widely used for their therapeutic properties in many clinical applications including myocardial infarction. Despite promising preclinical results and evidences of safety and efficacy in phases I/ II, inconsistencies in phase III trials have been reported. In a previous study, we have shown using MSC derived from the bone marrow of PPARβ/δ (Peroxisome proliferator-activated receptors β/δ) knockout mice that the acute cardioprotective properties of MSC during the first hour of reperfusion are PPARβ/δ-dependent but not related to the anti-inflammatory effect of MSC. However, the role of the modulation of PPARβ/δ expression on MSC cardioprotective and anti-apoptotic properties has never been investigated. Objectives: The aim of this study was to investigate the role of PPARβ/δ modulation (inhibition or activation) in MSC therapeutic properties in vitro and ex vivo in an experimental model of myocardial infarction.Methods and results: Naïve MSC and MSC pharmacologically activated or inhibited for PPARβ/δ were challenged with H202. Through specific DNA fragmentation quantification and qRT-PCR experiments, we evidenced in vitro an increased resistance to oxidative stress in MSC pre-treated by the PPARβ/δ agonist GW0742 versus naïve MSC. In addition, PPARβ/δ-priming allowed to reveal the anti-apoptotic effect of MSC on co-cultured cardiomyocytes. When injected during reperfusion in an ex vivo heart model of myocardial infarction, PPARβ/δ-primed MSC at a dose of 3.75x105 MSC/heart provided the same cardioprotective efficiency than 7.5x105 naïve MSC, identified as the optimal dose in our model. These enhanced short-term cardioprotective effects were associated with an increase in both anti-apoptotic effects and the number of MSC detected in the left ventricular wall at 1 hour of reperfusion. By contrast, inhibition of PPARβ/δ before their administration in post-ischemic hearts during reperfusion decreased their cardioprotective effects. Conclusion: Altogether these results revealed that PPARβ/δ-primed MSC exhibit an increased resistance to oxidative stress and enhanced anti-apoptotic properties on cardiac cells in vitro. PPARβ/δ-priming appears as an innovative strategy to enhance the cardioprotective effects of MSC and to decrease the injected doses. These results could be of major interest to improve MSC efficacy for the cardioprotection of injured myocardium in AMI patients.

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Effects of Sodium Nitroprusside on Platelet Adhesion, Subsequent Aggregation and Vasoconstriction

10.1055/s-0039-1682743 ◽

1977 ◽

Author(s):

H.R. Baumgartner

Keyword(s):

Myocardial Infarction ◽

Acute Myocardial Infarction ◽

Sodium Nitroprusside ◽

Platelet Adhesion ◽

Ex Vivo ◽

Balloon Catheter ◽

Strong Inhibitor ◽

Induced Aggregation

Sodium nitroprusside (SNP), a potent vasodilator, has shown beneficial effects in acute myocardial infarction. Since platelets may play an important role in the pathogenesis of myocardial infarction, the effect of SNP on their interaction with rabbit aorta subendothelium was investigated in vivo and under controlled blood flow conditions ex vivo and in vitro.One iliac artery and the abdominal aorta were denuded of endothelium by balloon catheter injury during infusion of glucose, SNP at 6 or 12 μg/kg/min in groups of 12, 6 and 7 rabbits respectively. The aorta and their branches were perfuse-fixed under controlled pressure 10 min after denudation. Morphometric evaluation showed dose-dependent and significant (2p < 0.01 or 0.001) inhibition of platelet spreading, adhesion and aggregation. The latter was abolished at the higher dose of SNP. Denudation and subsequent platelet adhesion caused strong vasoconstriction (2p < 0.001) which was inhibited by SNP (2p < 0.01).By exposure of subendothelium to either citrated blood or native blood in a flow chamber (2000 sec-1 shear rate) strong inhibition of spreading and adhesion-induced aggregation was again demonstrated at 6 and 12 μg/kg/min SNP. In vitro, adhesion-induced aggregation was completely abolished after the addition of SNP to rabbit (at 20 μg/ml) or human blood (2 μg/ml). 1 μg/ml PGE1 was needed to induce a similar inhibitory effect.Thus SNP is a strong inhibitor of platelet function and of injury + platelet induced vasoconstriction. These findings may explain its beneficial effect in acute myocardial infarction.

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Effect of creatine kinase-MM subtype composition on a CK-MB immunoinhibition assay.

Clinical Chemistry ◽

10.1093/clinchem/34.3.535 ◽

1988 ◽

Vol 34 (3) ◽

pp. 535-538 ◽

Cited By ~ 2

Author(s):

I M Morison ◽

K J Clayson ◽

J S Fine

Keyword(s):

Myocardial Infarction ◽

Creatine Kinase ◽

Human Serum ◽

Time Dependent ◽

Clinical Specimens ◽

In Vitro Incubation ◽

Dependent Change ◽

Time Dependent Change

Abstract Immunoinhibition (INH) by use of polyclonal anti-human CK-M antibody may be used to measure CK-MB in serum. Previous studies have shown that inhibiting antibodies prepared against purified muscle extracts may inhibit CK-MM by greater than 99%. Using patients' sera and muscle homogenates incubated with human serum, we studied the effect of CK-MM subtype composition on an INH assay. We found that with increasing time from the CK-releasing event, e.g., myocardial infarction, or with longer in vitro incubation, the proportion of CK-MM1 increased and the proportion of uninhibited CK-MM increased from 0.2% to 0.7-0.8%. As a consequence, CK-MB activity may be overestimated by as much as 1.6% of total CK when uncorrected INH results are used. Inhibition was maximal in samples containing 100% CK-MM3, the tissue subtype. Because of the time-dependent change in CK-MM subtypes, published results for INH from studies in which CK-MM purified from muscle was used may not be directly applicable to clinical specimens.

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Inhibition of Stimulated Interleukin-2 Production in Whole Blood: A Practical Measure of Cyclosporine Effect

Clinical Chemistry ◽

10.1093/clinchem/45.9.1477 ◽

1999 ◽

Vol 45 (9) ◽

pp. 1477-1484 ◽

Cited By ~ 67

Author(s):

C Michael Stein ◽

John J Murray ◽

Alastair JJ Wood

Keyword(s):

Whole Blood ◽

Ex Vivo ◽

Interleukin 2 ◽

Major Effect ◽

Biologically Relevant ◽

Maximum Inhibition ◽

Before And After ◽

Dose Dependent Fashion ◽

The Relationship

Abstract Background: Prediction of cyclosporine (CSA) efficacy and toxicity in individual patients is difficult. There is no practical, biologically relevant, pharmacodynamic measure of CSA effect. A major effect of CSA is to decrease interleukin-2 (IL-2) production; however, measurement of this effect in isolated lymphocytes as a marker of response to CSA has been problematic. Methods: CSA inhibition of phytohemagglutinin-P (PHA)-stimulated IL-2 production, measured by ELISA, was studied ex vivo in whole blood drawn before, and after subjects received 4 mg/kg oral CSA. Results: Four hours after CSA was administered, the mean (± SD) CSA concentration was 702 ± 196 μg/L and PHA-stimulated IL-2 production decreased by 68.7% ± 17.2% (P <0.0001; n = 17). Twenty-four hours after CSA was administered, concentrations were low (64 ± 24 μg/L), with no inhibition of IL-2 production. A rapid, concentration-dependent response occurred. Maximum CSA concentrations (944 ± 187 μg/L) and maximum inhibition of IL-2 production (86.9% ± 13.7%) occurred 90 min after subjects received CSA. In vitro, 32.5–1200 μg/L CSA also inhibited PHA-stimulated IL-2 production in whole blood in a dose-dependent fashion with a similar IC50 (∼300–400 μg/L) ex vivo and in vitro. Conclusion: In the search for a pharmacodynamic marker to better guide immunosuppressive therapy, the relationship between this simple, biologically relevant measure of CSA effect and clinical outcome should be determined.

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Effects of Magnesium on Platelet Aggregation and Adhesion

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648983 ◽

1994 ◽

Vol 72 (06) ◽

pp. 912-918 ◽

Cited By ~ 94

Author(s):

M Gawaz ◽

I Ott ◽

A J Reininger ◽

F-J Neumann

Keyword(s):

Myocardial Infarction ◽

Platelet Aggregation ◽

Platelet Function ◽

Platelet Adhesion ◽

Bleeding Time ◽

Magnesium Deficiency ◽

Ex Vivo ◽

Surface Expression ◽

Activated Platelets

SummaryMagnesium deficiency and its association with platelet hyperreactivity has been well recognised in a variety of diseases including myocardial infarction, preeclampsia, and diabetes. In order to investigate potential effects of intravenous Mg2+ supplementation, platelet function was studied by measurements of in vitro bleeding time (BT) and of fibrinogen (Fg)-mediated aggregation of washed platelets. In addition, the effect of Mg2+ on platelet adhesion onto immobilised Fg, on Fg binding to activated platelets, and on surface expression of GMP-140 or GP53 was evaluated. Mg2+(4 mM) prolonged in vitro BT by 30% and inhibited Fg-mediated aggregation significantly, independent of the agonist used to initiate platelet aggregation (ADP, collagen, epinephrine, thrombin, phorbol ester). Adhesion of resting platelets to immobilised Fg was reduced by 50% in the presence of 2 mM Mg2+. Moreover, Mg2+ reduced Fg binding to ADP- or collagen-stimulated platelets as well as surface expression of GMP-140 with an IC50 of approximately 3 mM. Intravenous administration of Mg2+ to healthy volunteers inhibited both ADP-induced platelet aggregation (p <0.05) by 40% and binding of Fg or surface expression of GMP-140 by 30% (p <0.05). Thus, pharmacological concentrations of Mg2+ effectively inhibit platelet function in vitro and ex vivo.

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