scholarly journals Polyregional aggregatometry of blood in patients with acute thrombosis as a potential model for preclicinal studies of new correctors of hemostasis system ex vivo

2017 ◽  
Vol 16 (1) ◽  
pp. 65-71
Author(s):  
A. L. Urakov ◽  
A. V. Samorodov ◽  
F. Kh. Kamilov ◽  
F. A. Khaliullin
Keyword(s):  
Ex Vivo ◽  
Blood Stream ◽  
Laboratory Animals ◽  
Coagulation System ◽  
Pharmacological Agents ◽  
Acute Thrombosis ◽  
Coronary Syndrome ◽  
Hemostasis System

Current recommendations to develop new pharmacological agents influencing hemostasis system are limited to the studies carried out on healthy volunteers in vitro, healthy lab animals and animals with model thrombosis in vivo. However, specific and seasonal peculiarities of hemostasis system in laboratory animals may not always adequately represent processes that occur during an event of cardiovascular accident. In this context, the main objective of this work is to study hemostasis system in patients with thrombotic events and to use the received data as a model for assessing the effectiveness of pharmacological agents as exemplified by aspirin and pentoxifylline in ex vivo. Experimental work is carried out on the blood of healthy male donors and patients with acute thrombosis. The findings show that completed thrombosis in one of the regions of coronary blood stream is not always accompanied by systemic tension of the coagulation system. The method of polyregional thromboelastography was used to study hemostasis system and to find out that hyperaggregation of platelets is a responsible part in the development of system hyperactivity of hemostasis in cases where it is registered. Pulmonary embolism, mesenteric ischemia and acute coronary syndrome is most often accompanied by systemic hyperaggegatioin of platelets, which can be used to assess the therapeutic efficacy of medicaments under conditions ex vivo. Aspirin and pentoxifylline are taken as examples to prove the necessity of preclinical studies for potential correctors of hemostatic system aimed to assess therapeutic effectiveness under conditions ex vivo in patients with completed thrombosis.

scholarly journals The MEMIC: An ex vivo system to model the complexity of the tumor microenvironment

2021 ◽  
Author(s):  
Libuše Janská ◽  
Libi Anandi ◽  
Nell C. Kirchberger ◽  
Zoran S. Marinkovic ◽  
Logan T. Schachtner ◽  
...  
Keyword(s):  
Tumor Microenvironment ◽  
Tumor Cells ◽  
Ex Vivo ◽  
Tumor Stroma ◽  
Stem Cell Differentiation ◽  
Blood Stream ◽  
Direct Access ◽  
Ex Vivo Model

There is an urgent need for accurate, scalable, and cost-efficient experimental systems to model the complexity of the tumor microenvironment. Here, we detail how to fabricate and use the Metabolic Microenvironment Chamber (MEMIC) – a 3D-printed ex vivo model of intratumoral heterogeneity. A major driver of the cellular and molecular diversity in tumors is the accessibility to the blood stream that provides key resources such as oxygen and nutrients. While some tumor cells have direct access to these resources, many others must survive under progressively more ischemic environments as they reside further from the vasculature. The MEMIC is designed to simulate the differential access to nutrients and allows co-culturing different cell types, such as tumor and immune cells. This system is optimized for live imaging and other microscopy-based approaches, and it is a powerful tool to study tumor features such as the effect of nutrient scarcity on tumor-stroma interactions. Due to its adaptable design and full experimental control, the MEMIC provide insights into the tumor microenvironment that would be difficult to obtain via other methods. As a proof of principle, we show that cells sense gradual changes in metabolite concentration resulting in multicellular spatial patterns of signal activation and cell proliferation. To illustrate the ease of studying cell-cell interactions in the MEMIC, we show that ischemic macrophages reduce epithelial features in neighboring tumor cells. We propose the MEMIC as a complement to standard in vitro and in vivo experiments, diversifying the tools available to accurately model, perturb, and monitor the tumor microenvironment, as well as to understand how extracellular metabolites affect other processes such as wound healing and stem cell differentiation.

188 DEVELOPMENT OF AN EFFECTIVE WHOLE-OVARY PERFUSION SYSTEM

2015 ◽  
Vol 27 (1) ◽  
pp. 185
Author(s):  
S. Maffei ◽  
G. Galeati ◽  
G. Pennarossa ◽  
T. A. L. Brevini ◽  
G. Gandolfi
Keyword(s):  
Cell Proliferation ◽  
Ex Vivo ◽  
Animal Health ◽  
Basal Medium ◽  
Laboratory Animals ◽  
Large Animal ◽  
Perfusion System ◽  
Whole Ovaries

The different structures of a mammalian ovary require complex 3-dimensional interactions to function properly. It is difficult to access the ovary in vivo and to study its physiology in vitro, it is necessary to dissect its different parts and culture them individually. Although informative, this approach prevents the understanding of the role played by their interactions. Perfusion systems are available for ovaries of laboratory animals while organs of larger species have been maintained in culture only for a few hours. This has prompted us to develop a system that can preserve the function of a whole sheep ovary for a few days ex vivo so that it is available for analysis in controlled conditions. Twenty-four sheep ovaries were collected at the local abattoir; 18 were assigned randomly to 3 experimental groups (media A, B, and C) and 6 were immediately fixed in 10% formaldehyde and used as fresh controls. Whole ovaries were cultured for up to 4 days using a semi-open perfusion system. Organs were perfused through the ovarian artery, at a flow rate of 1.5 mL min–1 with basal medium (M199, 25 mM HEPES, 2 mM l-glutamine and 100 µg mL–1 antibiotic-antimycotic solution) supplemented with 0.4% fatty acid free BSA (medium A); or 0.4% BSA heat shock fraction (medium B); or 10% FBS, 50 ng mL–1 IGF-1, and 50 mg bovine insulin (medium C). Ovaries were stimulated with FSH (Folltropin®-V, Bioniche Animal Health Inc., Belleville, Ontario, Canada) changing medium in a pulsatile manner (1 mg mL–1 for 2 h; 0.5 mg mL–1 for 2 h; 0 mg mL–1 for 20 h), with the same cycle repeated each day of culture. At every change, aliquots were collected for oestradiol (E2) and progesterone (P4) quantification. After culture, ovaries were examined for follicular morphology, cell proliferation, and apoptotic rate. Statistical analysis was performed using one-way ANOVA (SPSS 20, IBM, Armonk, NY, USA). In media A and B, all morphological parameters showed a small but significant decrease compared to fresh control, only after 3 days of culture. The different BSA in medium B did not affect follicle morphology but significantly increased cell proliferation (medium A, 28.59 ± 3.26%; medium B, 32.04 ± 2.67%) and decreased apoptosis (medium A, 32.51 ± 5.92%; medium B, 24.55 ± 2.55%). In both media, steroid concentration increased after FSH pulses (E2 range 1.95–10.50 pg mL–1; P4 range 0.34–3.08 ng mL–1), reaching levels similar to those measurable in peripheral plasma. The presence of FBS, IGF-1, and insulin in medium C allowed extension of the culture period to 4 days with a percentage of intact follicles comparable to that observed after 3 days in media A and B. Moreover, proliferation rates were comparable to fresh controls. Steroid pattern changed with P4 values dropping close to zero (range 0.03–1.18 ng mL–1) and E2 level (range 23.59–94.98 pg mL–1) increasing 10-fold, achieving a concentration similar to that measured in the ovarian vein around oestrous. Our data indicate that it is possible to support viability of large animal whole ovaries for up to 4 days, providing a physiologically relevant model for studying ovarian functions in vitro. Research was supported by AIRC IG 10376 and by the Carraresi Foundation.

scholarly journals Nanocurcumin-Loaded UCNPs for Cancer Theranostics: Physicochemical Properties, In Vitro Toxicity, and In Vivo Imaging Studies

Nanomaterials ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 2234
Author(s):  
Anbharasi Lakshmanan ◽  
Roman A. Akasov ◽  
Natalya V. Sholina ◽  
Polina A. Demina ◽  
Alla N. Generalova ◽  
...  
Keyword(s):  
Near Infrared ◽  
Therapeutic Potential ◽  
Ex Vivo ◽  
Laboratory Animals ◽  
In Vitro Toxicity ◽  
Upconversion Nanoparticles ◽  
Lewis Lung Cancer ◽  
Spectral Bands

Formulation of promising anticancer herbal drug curcumin as a nanoscale-sized curcumin (nanocurcumin) improved its delivery to cells and organisms both in vitro and in vivo. We report on coupling nanocurcumin with upconversion nanoparticles (UCNPs) using Poly (lactic-co-glycolic Acid) (PLGA) to endow visualisation in the near-infrared transparency window. Nanocurcumin was prepared by solvent-antisolvent method. NaYF4:Yb,Er (UCNP1) and NaYF4:Yb,Tm (UCNP2) nanoparticles were synthesised by reverse microemulsion method and then functionalized it with PLGA to form UCNP-PLGA nanocarrier followed up by loading with the solvent-antisolvent process synthesized herbal nanocurcumin. The UCNP samples were extensively characterised with XRD, Raman, FTIR, DSC, TGA, UV-VIS-NIR spectrophotometer, Upconversion spectrofluorometer, HRSEM, EDAX and Zeta Potential analyses. UCNP1-PLGA-nanocurcumin exhibited emission at 520, 540, 660 nm and UCNP2-PLGA-nanocurmin showed emission at 480 and 800 nm spectral bands. UCNP-PLGA-nanocurcumin incubated with rat glioblastoma cells demonstrated moderate cytotoxicity, 60–80% cell viability at 0.12–0.02 mg/mL marginally suitable for therapeutic applications. The cytotoxicity of UCNPs evaluated in tumour spheroids models confirmed UCNP-PLGA-nanocurcumin therapeutic potential. As-synthesised curcumin-loaded nanocomplexes were administered in tumour-bearing laboratory animals (Lewis lung cancer model) and showed adequate contrast to enable in vivo and ex vivo study of UCNP-PLGA-nanocurcumin bio distribution in organs, with dominant distribution in the liver and lungs. Our studies demonstrate promise of nanocurcumin-loaded upconversion nanoparticles for theranostics applications.

Abstract 17216: Intracoronary Dual-modal OCT/NIRF Structural-Molecular Imaging with a Clinical Dose of Indocyanine Green (ICG) for Detection of High-risk Coronary Plaques in Diabetic Swine Model

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Sunwon Kim ◽  
Min Woo Lee ◽  
Han Saem Cho ◽  
Joon Woo Song ◽  
Sunki Lee ◽  
...  
Keyword(s):  
High Risk ◽  
Near Infrared ◽  
Ex Vivo ◽  
Swine Model ◽  
Fully Integrated ◽  
Coronary Syndrome ◽  
Fibrous Cap ◽  
Nirf Imaging

Background: Acute coronary syndrome is frequently caused by rupture of macrophage abundant plaques with a large lipid-rich core. The present study aimed to investigate whether a fully integrated OCT/NIRF imaging combined with a clinically available near-infrared fluorescence (NIRF) enhancing ICG can detect the inflamed, lipid-rich plaques in swine coronary atheromata whose phenotype is similar to human vulnerable fibroatheroma. Methods and Results: Accelerated atherosclerosis was made by coronary balloon denudation in alloxan induced diabetic minipigs. A rapid coronary imaging (20 mm/sec pullback speed) using a fully integrated OCT/NIRF catheter was safely performed 30 minutes after I.V. injection of ICG (2.0 mg/kg) just under contrast purge. OCT clearly identified the lipid-rich plaques with fibrous cap. Simultaneously acquired, distance-calibrated NIRF imaging detected lipid-laden macrophage signals in OCT-proven plaques (figure). The in vivo plaque target-to-background ratio (pTBR) was significantly higher in ICG-injected swine compared to non-diabetic swines or saline-injected controls (p<0.05), which was validated on ex vivo fluorescence reflectance imaging (FRI) (figure). The in vivo and ex vivo peak pTBRs correlated significantly (p<0.05). In vitro experiments, and histopathology including fluorescence microscopic imaging and immunostaining of the plaque sections corroborated the findings in vivo . Conlusions: An OCT/NIRF imaging with a clinical use of ICG accurately identified macrophage abundant, lipid-rich coronary plaques in diabetic atheromatous minipigs. This highly translatable dual-modal molecular-structural imaging could be relevant for clinical intracoronary detection of high-risk plaques.

Antithrombotic drug candidate ALX-0081 shows superior preclinical efficacy and safety compared with currently marketed antiplatelet drugs

Blood ◽  
2011 ◽  
Vol 118 (3) ◽  
pp. 757-765 ◽  
Author(s):  
Hans Ulrichts ◽  
Karen Silence ◽  
Anne Schoolmeester ◽  
Peter de Jaegere ◽  
Stefaan Rossenu ◽  
...  
Keyword(s):  
Molecular Size ◽  
Therapeutic Window ◽  
Drug Candidate ◽  
Efficacy And Safety ◽  
Acute Thrombosis ◽  
Coronary Syndrome ◽  
Platelet Receptor ◽  
Surgical Bleeding

Abstract Neutralizing the interaction of the platelet receptor gpIb with VWF is an attractive strategy to treat and prevent thrombotic complications. ALX-0081 is a bivalent Nanobody which specifically targets the gpIb-binding site of VWF and interacts avidly with VWF. Nanobodies are therapeutic proteins derived from naturally occurring heavy-chain–only Abs and combine a small molecular size with a high inherent stability. ALX-0081 exerts potent activity in vitro and in vivo. Perfusion experiments with blood from patients with acute coronary syndrome on standard antithrombotics demonstrated complete inhibition of platelet adhesion after addition of ALX-0081, while in the absence of ALX-0081 residual adhesion was observed. In a baboon efficacy and safety model measuring acute thrombosis and surgical bleeding, ALX-0081 showed a superior therapeutic window compared with marketed antithrombotics. Pharmacokinetic and biodistribution experiments demonstrated target-mediated clearance of ALX-0081, which leads to a self-regulating disposition behavior. In conclusion, these preclinical data demonstrate that ALX-0081 combines a high efficacy with an improved safety profile compared with currently marketed antithrombotics. ALX-0081 has entered clinical development.

scholarly journals Modern view on hemostasis system: cell theory

2019 ◽  
pp. 72-77
Author(s):  
I. V. Schastlivtsev ◽  
K. V. Lobastov ◽  
S. N. Tsaplin ◽  
D. S. Mkrtychev
Keyword(s):  
Coagulation Factors ◽  
Cascade Model ◽  
Coagulation System ◽  
Cell Theory ◽  
Cascade Theory ◽  
Plasma Coagulation ◽  
Hemostasis System ◽  
Thrombin Generation Assay

For many years, there has been no model capable of explaining the complex processes of interaction between various bloodclotting factors leading to a stop of bleeding. One of the most successful models able to partially reflect the mechanisms of hemostasis for a long time was the cascade theory. The cascade model perfectly explains the processes occurring during coagulation in vitro, but was completely inadequate in attempts to evaluate the processes occurring in vivo. A significant drawback of the cascade model is the impossibility to trace the interaction of cells carrying the tissue factor, platelets and plasma coagulation factors on their surface, since these conditions cannot be imitated. The cell theory, which has replaced the cascade theory, pays attention not only to the interaction of plasma coagulation factors, but also takes into account the role of platelets as important participants of coagulation processes. It is based on a four-stage reaction cascade that includes the following stages: initiation, amplification, propagation, and termination.The cell theory of hemostasis is able to reflect the complex process of interaction of all the links of hemostasis and answer questions related to the problems in patients with disorders of the coagulation system. The cell theory of hemostasis allows to reflect more precisely the processes of hemostasis in vivo and to interpret correctly the results of tests and pathophysiological mechanisms of disorders of the coagulation system. Global tests (thrombin generation assay, thromboelastography, thrombodynamics) used for hemostasis system evaluation are more complimentary with cell theory of hemostasis.

Biochemical and Pharmacologic Profiling of a Novel Dual Acting Antithrombin Agent with Antiprotease and Antiplatelet Actions. Hematologic Implications.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1876-1876
Author(s):  
Jawed Fareed ◽  
D. Hoppensteadt ◽  
W. Haque ◽  
J. Diakur ◽  
W. Jeske ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Dose Range ◽  
Coagulation System ◽  
Antithrombotic Drugs ◽  
Platelet Activity ◽  
Titration Assay ◽  
Sites Of Action ◽  
Dose Dependent

Abstract Background: The pathogenesis of thrombosis involves both cellular and humoral processes. Most antithrombotic drugs exhibit either anti-protease or anti-platelet effects. A combination of anti-protease and anti-platelet drugs provides better efficacy in the management of thrombotic disorders. A series of synthetic low molecular weight serine protease inhibitors with varying anti-platelet effects (Medicure Inc.) are being assessed for antithrombotic properties. Materials and Methods: This investigation reports on a compound with low antithrombin/high anti-platelet activity MC 45301 (A) and a compound with high antithrombin/low anti-platelet activity MC 45308 (B) activity in in-vitro and in-vivo settings used to profile antithrombotic drugs. Results: A exhibited strong anti-platelet actions as measured using ADP as an agonist (IC50=1.1 g/ml), whereas B had a higher IC50 (9.4 g/ml). In the antithrombin titration assay A (>100 μg/ml) showed a relatively higher IC50 than B (45 μg/ml). In the global anticoagulant assays, A exhibited somewhat weaker effects than B. In the Xa generation assay, both compounds exhibited similar effects. However, in the thrombin generation assays B exhibited stronger effects. In whole blood assays both compounds produced anticoagulant and anti-platelet effects. Intravenous administration of these compounds to rabbits over a dose range of 50–500 g/kg produced strong dose dependent antithrombotic actions. In comparison to direct antithrombin agents such as argatroban, at a comparable dose, B produced identical antithrombotic actions, which were disproportional to the systemic anticoagulant effects. A produced modest antithrombotic actions with minimal ex vivo clotting effects. This data is highly suggestive that compounds with dual targets are able to produce stronger antithrombotic actions relative to monotherapeutic agents. Additional studies in arterial thrombosis may provide newer insights into the antithrombotic actions of compounds with dual sites of action. Moreover, these agents may be more effective in thrombotic conditions where both platelets and the coagulation system are involved.

Effects of human cytomegalovirus infection on the coagulation system

2005 ◽  
Vol 93 (03) ◽  
pp. 403-410 ◽  
Author(s):  
Victor Gerdes ◽  
Harry Büller ◽  
Alessandro Squizzato
Keyword(s):  
Case Reports ◽  
Acute Infection ◽  
In Vivo Studies ◽  
Coagulation System ◽  
Cmv Infection ◽  
Acute Thrombosis ◽  
Human Cytomegalovirus Infection ◽  
Human Cmv

SummaryPathophysiological mechanisms of acute vascular thrombosis are not fully understood. It has been suggested that different infectious pathogens are responsible agents of thrombotic disorders. The infection hypothesis is supported by an increasing number of reports on the interaction between acute infection and coagulation. Cytomegalovirus (CMV) is supposed to play an important role in apparently unprovoked thrombosis. We reviewed all human in vitro and in vivo studies on the influence of human CMV infection on the coagulation system, as well as all case reports of acute thrombosis during acute human CMV infection. In the published literature there is mounting evidence that human CMV may play a role in thrombotic disorders. Definitive conclusions, however, cannot be drawn, although the in vitro studies are convincing and offer insight in the pathogenesis.

scholarly journals The Comparative Evaluation of γ-Linolenic Acid and Dihomo-γ-Linolenic Acid, their Esters and Possible Precursors as Potential Antithrombotic/Thrombolytic Agents

1977 ◽  
Author(s):  
A.K. Sim ◽  
A.P. McCraw
Keyword(s):  
Linolenic Acid ◽  
Unsaturated Fatty Acids ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Plasma Cholesterol ◽  
Laboratory Animals ◽  
Prostaglandin Precursors ◽  
Dose Levels

γ-linolenic acid, dihomo-γ-linolenic acid and Naudicelle, a naturally-occurring rich source of essential unsaturated fatty acids have been evaluated as potential antithrombotic agents. A sequential series of tests was used to compare each substance in vitro (human plasma), in vivo using an artificially induced thrombus in the microcirculation of small laboratory animals and ex vivo in non-human primates with a spontaneous pathological thrombotic tendency. Finally the test substances were compared in man after oral administration. Particular reference was made to the effects of the compounds on platelet function, the fibrinolytic system, thrombus formation and on circulating blood lipids. The compounds which are possible prostaglandin precursors, have been shown to have a positive (antithrombotic) effect in the test systems used. Platelet aggregation was inhibited over 24 hours in non-human primates and in man at oral dose levels of 1.5 mg/kg. In addition plasma cholesterol and triglycerides were decreased by up to 50% on the same dose regime.

scholarly journals Tannins as Hemostasis Modulators

2022 ◽  
Vol 12 ◽  
Author(s):  
Natalia Marcińczyk ◽  
Anna Gromotowicz-Popławska ◽  
Michał Tomczyk ◽  
Ewa Chabielska
Keyword(s):  
Coagulation System ◽  
Thromboembolic Events ◽  
Plant Origin ◽  
Hemostasis System ◽  
Pathological Conditions ◽  
Process Studies ◽  
Special Mention ◽  
Promising Source

The hemostasis system is often affected by complications associated with cardiovascular diseases, which results in thromboembolic events. Compounds of plant origin and plant extracts are considered as a promising source of substances that could modulate the functioning of the hemostasis system and thus reduce the risk of thromboembolism. Among them, tannins, which are plant-origin compounds with potential effects in hemostasis, deserve a special mention. This paper describes the hemostasis-modifying ability of three groups of tannins, namely ellagitannins, gallotannins, and procyanidins. The review highlights the desirable as well as undesirable influence of tannins on specific components of hemostasis, namely platelets, coagulation system, fibrinolysis system, and endothelium, and the multidirectional effect of these compounds on the thrombotic process. Studies performed under normal and pathological conditions such as diabetes or hypercoagulation are described, and the pathophysiology-dependent action of tannins is also highlighted. Most of the studies presented in the paper were performed in vitro, and due to the low bioavailability of tannins more studies should be conducted in the future to understand their actual activity in vivo.

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