Inhibitory effects of hydrogen on in vitro platelet activation and in vivo prevention of thrombosis formation

Abstract Background: Soluble CD40 ligand (sCD40L) was suggested as a novel biomarker of cardiovascular risk. We examined the effect of preanalytical variation on the measurement of sCD40L concentration. Methods: From healthy control individuals (n = 20) and patients with acute coronary syndrome (ACS) (n = 20) or sepsis (n = 20), we obtained blood drawn into 5 tubes containing citrate or a mixture of citrate, theophylline, adenosine, and dipyridamole (CTAD). The tubes were incubated for 30 min at room temperature or 0 °C before a single or double centrifugation (15 min, 2500g) at room temperature or 4 °C, respectively. sCD40L, β-thromboglobulin (βTG), and platelet factor 4 (PF4) concentrations were measured using immunoassays. Results: Concentrations of sCD40L were very low in all CTAD and citrated samples maintained at 0 °C (median ≤0.076 μg/L). Although increased βTG and PF4 confirmed disease-related in vivo platelet activation, sCD40L was not higher in patients than in controls. In contrast, if the samples were processed at room temperature, sCD40L was significantly higher in ACS patients than in controls (P <0.02 in CTAD and citrated plasma at room temperature). Moreover, the βTG:PF4 ratio decreased in patient but not control CTAD samples, suggesting a greater susceptibility of patient platelets to in vitro activation. Conclusions: Increased sCD40L concentrations resulted from in vitro platelet activation during sample preparation. Disease-related in vivo activation did not contribute to sCD40L concentrations in plasma. Therefore, published studies of sCD40L demand cautious interpretation, because their preanalytical conditions were not standardized.

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Submission for Special Issue: The Role of Platelet Activation in the Pathophysiology of HIV, Tuberculosis, and Pneumococcal Disease. Bedaquiline Suppresses ADP-Mediated Activation of Human Platelets In Vitro via Interference With Phosphatidylinositol 3-Kinase

Frontiers in Immunology ◽

10.3389/fimmu.2020.621148 ◽

2021 ◽

Vol 11 ◽

Author(s):

Gregory R. Tintinger ◽

Annette J. Theron ◽

Helen C. Steel ◽

Moloko C. Cholo ◽

Jan G. Nel ◽

...

Keyword(s):

Platelet Activation ◽

Platelet Rich Plasma ◽

Statistical Significance ◽

Human Platelets ◽

Phosphatidylinositol 3 Kinase ◽

Inhibitory Effects ◽

Isolated Cells ◽

Phosphatidylinositol 3

Although bedaquiline has advanced the treatment of multidrug-resistant tuberculosis (TB), concerns remain about the cardiotoxic potential of this agent, albeit by unexplored mechanisms. Accordingly, we have investigated augmentation of the reactivity of human platelets in vitro as a potential mechanism of bedaquiline-mediated cardiotoxicity. Platelet-rich plasma (PRP) or isolated cells prepared from the blood of healthy, adult humans were treated with bedaquiline (0.625–10 µg/ml), followed by activation with adenosine 5’-diphosphate (ADP), thrombin or the thromboxane A2 receptor agonist (U46619). Expression of platelet CD62P (P-selectin), platelet aggregation, Ca2+ fluxes and phosphorylation of Akt1 were measured using flow cytometry, spectrophotometry, fluorescence spectrometry, and by ELISA procedures, respectively. Exposure to bedaquiline caused dose-related inhibition of ADP-activated, but not thrombin- or U46619-activated, expression of CD62P by platelets, achieving statistical significance at a threshold concentration of 5 µg/ml and was paralleled by inhibition of aggregation and Ca2+ mobilization. These ADP-selective inhibitory effects of bedaquiline on platelet activation were mimicked by wortmannin, an inhibitor of phosphatidylinositol 3-kinase (PI3-K), implicating PI3-K as being a common target of both agents, a contention that was confirmed by the observed inhibitory effects of bedaquiline on the phosphorylation of Akt1 following activation of platelets with ADP. These apparent inhibitory effects of bedaquiline on the activity of PI3-K may result from the secondary cationic amphiphilic properties of this agent. If operative in vivo, these anti-platelet effects of bedaquiline may contribute to ameliorating the risk of TB-associated cardiovascular disease, but this remains to be explored in the clinical setting.

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Combination of a novel growth hormone releasing hormone Antagonist JMR 132 and Tamoxifen enhances the inhibitory effects on growth of ZR 75 and T47D estrogen dependent human breast cancer cells in vitro and in vivo

Geburtshilfe und Frauenheilkunde ◽

10.1055/s-0029-1225201 ◽

2009 ◽

Vol 69 (05) ◽

Author(s):

S Buchholz ◽

AV Schally ◽

A Krishan ◽

A Papadia ◽

O Ortmann ◽

...

Keyword(s):

Breast Cancer ◽

Growth Hormone ◽

Human Breast Cancer ◽

Breast Cancer Cells ◽

Growth Hormone Releasing Hormone ◽

Human Breast Cancer Cells ◽

Human Breast ◽

Inhibitory Effects

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Inhibitory effects of epimedium herb water extract on the inflammatory response in vitro and in vivo

Planta Medica ◽

10.1055/s-0036-1596615 ◽

2016 ◽

Vol 81 (S 01) ◽

pp. S1-S381

Author(s):

YC Oh ◽

YH Jeong ◽

WK Cho ◽

SJ Lee ◽

JY Ma

Keyword(s):

Inflammatory Response ◽

Water Extract ◽

Inhibitory Effects

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The Effects of Brinolase (Fibrinolytic Enzyme from Aspergillus Oryzae) on Platelet Aggregation of Dog and Man

Thrombosis and Haemostasis ◽

10.1055/s-0038-1649039 ◽

1972 ◽

Vol 28 (01) ◽

pp. 031-048 ◽

Cited By ~ 4

Author(s):

W. H. E Roschlau ◽

R Gage

Keyword(s):

Platelet Aggregation ◽

Aspergillus Oryzae ◽

Degradation Products ◽

Blood Platelet ◽

Human Platelets ◽

Fibrinolytic Enzyme ◽

Inhibitory Effects ◽

Blood Platelet Aggregation

SummaryInhibition of blood platelet aggregation by brinolase (fibrinolytic enzyme from Aspergillus oryzae) has been demonstrated with human platelets in vitro and with dog platelets in vivo and in vitro, using both ADP and collagen as aggregating stimuli. It is suggested that the optimal inhibitory effects of brinolase occur indirectly through the generation of plasma fibrinogen degradation products, without compromising platelet viability, rather than by direct proteolysis of platelet structures.

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Defibrotide Inhibits Platelet Activation by Cathepsin G Released From Stimulated Polymorphonuclear Leukocytes

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648519 ◽

1992 ◽

Vol 67 (06) ◽

pp. 660-664 ◽

Cited By ~ 17

Author(s):

Virgilio Evangelista ◽

Paola Piccardoni ◽

Giovanni de Gaetano ◽

Chiara Cerletti

Keyword(s):

Platelet Activation ◽

Polymorphonuclear Leukocytes ◽

Direct Interaction ◽

Cathepsin G ◽

In Vivo Test ◽

Cell Functions ◽

Test Models ◽

Antithrombotic Effects

SummaryDefibrotide is a polydeoxyribonucleotide with antithrombotic effects in experimental animal models. Most of the actions of this drug have been observed in in vivo test models but no effects have been reported in in vitro systems. In this paper we demonstrate that defibrotide interferes with polymorphonuclear leukocyte-induced human platelet activation in vitro. This effect was not related to any direct interaction with polymorphonuclear leukocytes or platelets, but was due to the inhibition of cathepsin G, the main biochemical mediator of this cell-cell cooperation. Since cathepsin G not only induces platelet activation but also affects some endothelial cell functions, the anticathepsin G activity of defibrotide could help to explain the antithrombotic effect of this drug.

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Inhibitory Effects of a Reengineered Anthrax Toxin on Canine Oral Mucosal Melanomas

Toxins ◽

10.3390/toxins12030157 ◽

2020 ◽

Vol 12 (3) ◽

pp. 157 ◽

Cited By ~ 2

Author(s):

Adriana Tomoko Nishiya ◽

Marcia Kazumi Nagamine ◽

Ivone Izabel Mackowiak da Fonseca ◽

Andrea Caringi Miraldo ◽

Nayra Villar Scattone ◽

...

Keyword(s):

Tumor Cells ◽

Evaluation Criteria ◽

Anthrax Toxin ◽

Treatment Modalities ◽

Inhibitory Effects ◽

Upa Receptor ◽

New Treatment ◽

Oral Malignancy

Canine oral mucosal melanomas (OMM) are the most common oral malignancy in dogs and few treatments are available. Thus, new treatment modalities are needed for this disease. Bacillus anthracis (anthrax) toxin has been reengineered to target tumor cells that express urokinase plasminogen activator (uPA) and metalloproteinases (MMP-2), and has shown antineoplastic effects both, in vitro and in vivo. This study aimed to evaluate the effects of a reengineered anthrax toxin on canine OMM. Five dogs bearing OMM without lung metastasis were included in the clinical study. Tumor tissue was analyzed by immunohistochemistry for expression of uPA, uPA receptor, MMP-2, MT1-MMP and TIMP-2. Animals received either three or six intratumoral injections of the reengineered anthrax toxin prior to surgical tumor excision. OMM samples from the five dogs were positive for all antibodies. After intratumoral treatment, all dogs showed stable disease according to the canine Response Evaluation Criteria in Solid Tumors (cRECIST), and tumors had decreased bleeding. Histopathology has shown necrosis of tumor cells and blood vessel walls after treatment. No significant systemic side effects were noted. In conclusion, the reengineered anthrax toxin exerted inhibitory effects when administered intratumorally, and systemic administration of this toxin is a promising therapy for canine OMM.

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Trichostatin A downregulates bromodomain and extra-terminal proteins to suppress osimertinib resistant non-small cell lung carcinoma

Cancer Cell International ◽

10.1186/s12935-021-01914-y ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Yuting Meng ◽

Xixi Qian ◽

Li Zhao ◽

Nan Li ◽

Shengjie Wu ◽

...

Keyword(s):

Trichostatin A ◽

Small Cell Lung Carcinoma ◽

Cell Types ◽

Inhibitory Effects ◽

Cell Lung Carcinoma ◽

Growth Inhibitory ◽

Terminal Proteins ◽

P Cells

Abstract Background The third-generation epithelial growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) have shown significant therapeutic effects on patients with non-small cell lung carcinoma (NSCLC) who carry active EGFR mutations, as well as those who have developed acquired resistance to the first-generation of EGFR-TKIs due to the T790M mutation. However, most patients develop drug resistance after 8–10 months of treatment. Currently, the mechanism has not been well clarified, and new therapeutic strategies are urgently needed. Methods Osimertinib resistant cell lines were established by culturing sensitive cells in chronically increasing doses of osimertinib. The anticancer effect of reagents was examined both in vitro and in vivo using the sulforhodamine B assay and a xenograft mouse model. The molecular signals were detected by western blotting. The combination effect was analyzed using CompuSyn software. Results We found that bromodomain and extra-terminal proteins (BETs) were upregulated in osimertinib resistant (H1975-OR) cells compared with those in the paired parental cells (H1975-P), and that knockdown of BETs significantly inhibited the growth of H1975-OR cells. The BET inhibitor JQ1 also exhibited stronger growth-inhibitory effects on H1975-OR cells and a greater expression of BETs and the downstream effector c-Myc than were observed in H1975-P cells. The histone deacetylase (HDAC) inhibitor trichostatin A (TSA) showed stronger growth suppression in H1975-OR cells than in H1975-P cells, but vorinostat, another HDAC inhibitor, showed equal inhibitory efficacy in both cell types. Consistently, downregulation of BET and c-Myc expression was greater with TSA than with vorinostat. TSA restrained the growth of H1975-OR and H1975-P xenograft tumors. The combination of TSA and JQ1 showed synergistic growth-inhibitory effects in parallel with decreased BET and c-Myc expression in both H1975-OR and H1975-P cells and in xenograft nude mouse models. BETs were not upregulated in osimertinib resistant HCC827 cells compared with parental cells, while TSA and vorinostat exhibited equal inhibitory effects on both cell types. Conclusion Upregulation of BETs contributed to the osimertinib resistance of H1975 cells. TSA downregulated BET expression and enhanced the growth inhibitory effect of JQ1 both in vitro and in vivo. Our findings provided new strategies for the treatment of osimertinib resistance.

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