Species Specific Immunoassays to Measure Blood Platelet and Coagulation Activation in the Rat

1996 ◽  
Vol 76 (06) ◽  
pp. 1090-1095 ◽  
Author(s):  
C Ravanat ◽  
M Freund ◽  
S Schuhler ◽  
P Grunert ◽  
L Meyer ◽  
...  
Keyword(s):  
Antithrombin Iii ◽  
Plasma Concentrations ◽  
Simultaneous Detection ◽  
Blood Platelet ◽  
Coagulation Activation ◽  
Platelet Factor ◽  
Prethrombotic State ◽  
Low Levels ◽  
Species Specific ◽  
Higher Sensitivity

SummaryThe purpose of this study was to develop specific and sensitive immunoassays to detect early indices of hypercoagulability in the rat. Rat platelet factor 4 (rPF4) and rat fibrinopeptide A (rFPA) assays, tools for the detection of activation of platelets and coagulation respectively, were designed using antibodies raised against purified rPF4 and against synthetic rFPA. The relevance of these new assays and of the commercially available ELISA kit for thrombin-antithrombin III (TAT) complexes was demonstrated in a rat model of a prethrombotic state induced by intravenous infusion of varying doses of thromboplastin (90 to 2400 μl/kg/h). In this model, the immunoassays allowed simultaneous detection of low levels of rFPA and rPF4 which were correlated with fibrinogen and platelet consumption and TAT generation and further proved to be of higher sensitivity than the classical methods of platelet count or measurement of fibrinogen levels. Plasma concentrations of rFPA, rPF4 and TAT were dependent on infusion time and thromboplastin dose, while hirudin (1 mg/kg) prevented their appearance. Thus the new specific immunoassays for rPF4 and rFPA and the commercial human TAT assay represent useful tools for pathophysiological studies or the screening of antithrombotic drugs in rats.

Platelet and Coagulation Function in Patients with Abnormal Cardiac Valves Treated with Sulphinpyrazone

1981 ◽  
Vol 46 (04) ◽  
pp. 743-746 ◽  
Author(s):  
C A Ludlam ◽  
N Allan ◽  
R B blandford ◽  
R Dowdle ◽  
N J Bentley ◽  
...  
Keyword(s):  
Heart Disease ◽  
Antithrombin Iii ◽  
Plasma Concentrations ◽  
Consumption Coagulopathy ◽  
Elevated Plasma ◽  
Cardiac Valves ◽  
Platelet Factor ◽  
Coagulation Function ◽  
Specific Proteins ◽  
Rheumatic Heart

SummaryEight patients on warfarin with rheumatic heart disease and prosthetic cardiac valves were selected for study on the basis of persistently elevated plasma β-thromboglobulin (β-tg) and platelet factor 4 (PF4) concentrations. Platelet mean lifespan and fibrinogen half life were short, and positively correlated, and both were inversely related to the Plasma concentrations of the platelet specific proteins. Antithrombin III (ATIII) levels were also reduced. Treatment with sulphinpyrazone resulted in lengthening of both platelet and fibrinogen survival, a rise in ATIII but no change in the βtg or PF4 concentrations. It is concluded that patients with abnormal cardiac valves and raised plasma levels of βtg or PF4 have, despite warfarin, a consumption coagulopathy that can be inhibited by sulphinpyrazone.

Immunologic Studies of Antithrombin III Heparin Cofactor in the Newborn

1978 ◽  
Vol 39 (03) ◽  
pp. 624-630 ◽  
Author(s):  
W E Hathaway ◽  
L L Neumann ◽  
C A Borden ◽  
L J Jacobson
Keyword(s):  
Gestational Age ◽  
Antithrombin Iii ◽  
Newborn Infants ◽  
Term Infant ◽  
Sick Newborn ◽  
At Iii ◽  
Thrombotic Complications ◽  
Low Levels ◽  
Collection Methods ◽  
Made In

SummarySerial quantitative immunoelectrophoretic (IE) measurements of antithrombin III heparin cofactor (AT III) were made in groups of well and sick newborn infants classified by gestational age. Collection methods (venous vs. capillary) did not influence the results; serum IE measurements were comparable to AT III activity by a clotting method. AT III is gestational age-dependent, increasing from 28.7% of normal adult values at 28-32 weeks to 50.9% at 37-40 weeks, and shows a gradual increase to term infant levels (57.4%) by 3-4 weeks of age. Infants with the respiratory distress syndrome (RDS) show lower levels of AT III in the 33-36 week group, 22% vs. 44% and in the 37-40 week group, 33.6% vs. 50.9%, than prematures without RDS. Infants of 28-32 week gestational age had only slight differences, RDS = 24%, non-RDS = 28.7%. The lowest levels of AT III were seen in patients with RDS complicated by disseminated intravascular coagulation and those with necrotizing enterocolitis. Crossed IE on representative infants displayed a consistent pattern which was identical to adult controls except for appropriate decreases in the amplitude of the peaks. The thrombotic complications seen in the sick preterm infant may be related to the low levels of AT III.

Neutralization of Antithrombin VI (Fibrinogen Breakdown Products) with Platelet Antiheparin Factor (Platelet Factor 4)*

1965 ◽  
Vol 14 (03/04) ◽  
pp. 490-499 ◽  
Author(s):  
S Niewiarowski ◽  
R Farbiszewski ◽  
A Popławski
Keyword(s):  
Zinc Acetate ◽  
Antithrombin Iii ◽  
Platelet Factor 4 ◽  
Breakdown Product ◽  
Platelet Factor ◽  
Chromatography Column ◽  
Antiheparin Activity

SummaryIt has been found that fibrinogen breakdown product – antithrombin VI – is neutralized by the purified preparation of platelet factor 4, obtained by means of zinc acetate precipitation and DEAE chromatography column. It has been suggested that antiheparin activity of platelet factor 4 and its ability to neutralize antithrombin VI may be related to the same protein.The purified preparation of platelet factor 4 does not influence the fibrinogen – fibrin conversion by thrombin. This means that platelet factor 2 and platelet factor 4 are not the same substance.Crude platelet extracts neutralize antithrombin III and V. However, the purified product did not interferes with the action of these antithrombins.

scholarly journals A New Multiplex Real-Time RT-PCR for Simultaneous Detection and Differentiation of Avian Bornaviruses

Viruses ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1358
Author(s):  
Brigitte Sigrist ◽  
Jessica Geers ◽  
Sarah Albini ◽  
Dennis Rubbenstroth ◽  
Nina Wolfrum
Keyword(s):  
Real Time ◽  
Simultaneous Detection ◽  
Epidemiological Studies ◽  
Virus Species ◽  
Rt Pcr ◽  
P Gene ◽  
Field Samples ◽  
Causative Agents ◽  
Species Specific ◽  
Proventricular Dilatation Disease

Avian bornaviruses were first described in 2008 as the causative agents of proventricular dilatation disease (PDD) in parrots and their relatives (Psittaciformes). To date, 15 genetically highly diverse avian bornaviruses covering at least five viral species have been discovered in different bird orders. Currently, the primary diagnostic tool is the detection of viral RNA by conventional or real-time RT-PCR (rRT-PCR). One of the drawbacks of this is the usage of either specific assays, allowing the detection of one particular virus, or of assays with a broad detection spectrum, which, however, do not allow for the simultaneous specification of the detected virus. To facilitate the simultaneous detection and specification of avian bornaviruses, a multiplex real-time RT-PCR assay was developed. Whole-genome sequences of various bornaviruses were aligned. Primers were designed to recognize conserved regions within the overlapping X/P gene and probes were selected to detect virus species-specific regions within the target region. The optimization of the assay resulted in the sensitive and specific detection of bornaviruses of Psittaciformes, Passeriformes, and aquatic birds. Finally, the new rRT-PCR was successfully employed to detect avian bornaviruses in field samples from various avian species. This assay will serve as powerful tool in epidemiological studies and will improve avian bornavirus detection.

scholarly journals Internal exposure dynamics drive the Adverse Outcome Pathways of synthetic glucocorticoids in fish

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Luigi Margiotta-Casaluci ◽  
Stewart F. Owen ◽  
Belinda Huerta ◽  
Sara Rodríguez-Mozaz ◽  
Subramanian Kugathas ◽  
...  
Keyword(s):  
Predictive Power ◽  
Adverse Outcome ◽  
Plasma Concentrations ◽  
Internal Exposure ◽  
Adverse Outcome Pathway ◽  
Conceptual Tool ◽  
Fish Model ◽  
Species Specific

Abstract The Adverse Outcome Pathway (AOP) framework represents a valuable conceptual tool to systematically integrate existing toxicological knowledge from a mechanistic perspective to facilitate predictions of chemical-induced effects across species. However, its application for decision-making requires the transition from qualitative to quantitative AOP (qAOP). Here we used a fish model and the synthetic glucocorticoid beclomethasone dipropionate (BDP) to investigate the role of chemical-specific properties, pharmacokinetics, and internal exposure dynamics in the development of qAOPs. We generated a qAOP network based on drug plasma concentrations and focused on immunodepression, skin androgenisation, disruption of gluconeogenesis and reproductive performance. We showed that internal exposure dynamics and chemical-specific properties influence the development of qAOPs and their predictive power. Comparing the effects of two different glucocorticoids, we highlight how relatively similar in vitro hazard-based indicators can lead to different in vivo risk. This discrepancy can be predicted by their different uptake potential, pharmacokinetic (PK) and pharmacodynamic (PD) profiles. We recommend that the development phase of qAOPs should include the application of species-specific uptake and physiologically-based PK/PD models. This integration will significantly enhance the predictive power, enabling a more accurate assessment of the risk and the reliable transferability of qAOPs across chemicals.

Microparticles from Patients with Multiple Organ Dysfunction Syndrome and Sepsis Support Coagulation through Multiple Mechanisms

2001 ◽  
Vol 85 (05) ◽  
pp. 810-820 ◽  
Author(s):  
Karin Joop ◽  
René Berckmans ◽  
Rienk Nieuwland ◽  
Johanna Berkhout ◽  
Fred Romijn ◽  
...  
Keyword(s):  
Organ Dysfunction ◽  
Multiple Organ Dysfunction Syndrome ◽  
Plasma Concentrations ◽  
Annexin V ◽  
Multiple Organ ◽  
Factor Xii ◽  
Healthy Controls ◽  
Multiple Organ Dysfunction ◽  
Coagulation Activation

Summary Aim. We investigated the occurrence and thrombin generating mechanisms of circulating microparticles (MP) in patients with multiple organ dysfunction syndrome (MODS) and sepsis. Methods. MP, isolated from blood of patients (n = 9) and healthy controls (n = 14), were stained with cell-specific monoclonal antibodies (MoAbs) or anti-tissue factor (anti-TF) MoAb and annexin V, and analyzed by flow cytometry. To assess their thrombin-generating capacity, MP were reconstituted in normal plasma. The coagulation activation status in vivo was quantified by plasma prothrombin fragment F1+2- and thrombin-antithrombin (TAT) measurements. Results. Annexin V-positive MP in the patients originated predominantly from platelets (PMP), and to a lesser extent from erythrocytes, endothelial cells (EMP) and granulocytes (GMP). Compared to healthy controls, the numbers of annexin V-positive PMP and TF-exposing MP were decreased (p = <0.001 for both), EMP were decreased (E-selectin, p = 0.003) or found equal (CD144, p = 0.063), erythrocyte-derived MP were equal (p = 0.726), and GMP were increased (p = 0.008). GMP numbers correlated with plasma concentrations of elastase (r = 0.70, p = 0.036), but not with C-reactive-protein or interleukin-6 concentrations. Patient samples also contained reduced numbers of annexin V-negative PMP, and increased numbers of erythrocyte-derived MP and GMP (p = 0.005, p = 0.021 and p <0.001, respectively). Patient MP triggered thrombin formation, which was reduced compared to the healthy controls (p = 0.008) and strongly inhibited by an anti-factor XII MoAb (two patients), by anti-factor XI MoAb (eight patients) or by anti-TF MoAb (four patients). Concentrations of F1+2 and TAT were elevated (p = 0.005 and p = 0.001, respectively) and correlated inversely with the number of circulating MP (and r = –0.51, p = 0.013, and r = –0.65, p = 0.001, respectively) and their thrombin generation capacity (F1+2: r = –0.62, p = 0.013). Conclusions. In patients with MODS and sepsis relatively low numbers of MP are present that differ from controls in their cellular origin, numbers and coagulation activation mechanisms.

Seasonal changes of steroid concentrations in seminal plasma of a European wild boar

1984 ◽  
Vol 107 (3) ◽  
pp. 425-427 ◽  
Author(s):  
D. Schopper ◽  
J. Gaus ◽  
R. Claus ◽  
H. Bader
Keyword(s):  
Wild Boar ◽  
Seminal Plasma ◽  
Seasonal Pattern ◽  
Plasma Concentrations ◽  
Boar Taint ◽  
Steroid Production ◽  
European Wild Boar ◽  
Clear Seasonal Pattern ◽  
Boar Semen ◽  
Low Levels

Abstract. The influence of season on testicular steroid production as a parameter of testicular function has been studied in a wild boar. Semen was collected once weekly while it served the dummy. In seminal plasma concentrations of the following steroids were determined by radioimmunoassay: unconjugated testosterone, conjugated testosterone, unconjugated total oestrogens, conjugated total oestrogens and 5α-androst-16-en-3-one ('boar-taint steroid'). All steroids showed a clear seasonal pattern with highest concentrations in autumn and early winter and low levels from January to July. Maxima during the rutting season were 10–25 times greater than average values out of season. During a 2-month-period (mid-July until mid-September) libido was abolished and the wild boar refused to mount the dummy. These results indicate that the seasonal variation in testicular steroid production by the wild boar, regulated by photoperiod, are similar to those of the domestic boar.

Differential binding of gold-labeled zona pellucida glycoproteins mZP2 and mZP3 to mouse sperm membrane compartments

Development ◽  
1991 ◽  
Vol 113 (1) ◽  
pp. 141-149 ◽  
Author(s):  
S. Mortillo ◽  
P.M. Wassarman
Keyword(s):  
Plasma Membrane ◽  
Zona Pellucida ◽  
Transmission Electron ◽  
Acrosomal Membrane ◽  
Membrane Compartments ◽  
Sperm Membrane ◽  
Differential Binding ◽  
Inner Acrosomal Membrane ◽  
Low Levels ◽  
Species Specific

Egg zona pellucida glycoproteins mZP3 and mZP2 serve as primary and secondary sperm receptors, respectively, during initial stages of fertilization in mice [Wassarman (1988) A. Rev. Biochem. 57, 415–442]. These receptors interact with complementary egg-binding proteins (EBPs) located on the sperm surface to support species-specific gamete adhesion. Results of whole-mount autoradiographic experiments suggest that purified egg mZP3 and mZP2 bind preferentially to acrosome-intact (AI) and acrosome-reacted (AR) sperm heads, respectively [Bleil and Wassarman (1986) J. Cell Biol. 102, 1363–1371]. Here, we used purified egg mZP2, egg mZP3 and fetuin, which were coupled directly to colloidal gold (‘gold-probes’), to examine binding of these glycoproteins to membrane compartments of AI and AR sperm by transmission electron microscopy. mZP3 gold-probes were found associated primarily with plasma membrane overlying the acrosomal and post-acrosomal regions of AI sperm heads. They were also found associated with plasma membrane overlying the post-acrosomal region of AR sperm heads. mZP2 gold-probes were found associated primarily with inner acrosomal membrane of AR sperm heads, although some gold was associated with outer acrosomal membrane of AI sperm that had holes in plasma membrane overlying the acrosome. Fetuin gold-probes, used to assess background levels of binding, were bound at relatively low levels to plasma membrane and inner acrosomal membrane of AI and AR sperm, respectively. None of the gold-probes exhibited significant binding to sperm tails, or to red blood cells and residual bodies present in sperm preparations. These results provide further evidence that mZP2 and mZP3 bind preferentially to heads of AR and AI sperm, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Elevation Of Antithrombin III By Sulfinpyrazone Therapy In Chronic Renal Failure

1981 ◽  
Author(s):  
M Bern ◽  
J Green
Keyword(s):  
Renal Failure ◽  
Chronic Renal Failure ◽  
Antithrombin Iii ◽  
Chronic Hemodialysis ◽  
Protective Effects ◽  
Platelet Factor ◽  
At Iii ◽  
Before And After ◽  
Artery Disease ◽  
And Function

Sulfinpyrazone can reduce the incidence of thrombosis of A-V shunts in chronic renal failure. The drug is also reported to prevent acute deaths from coronary artery disease. This study was to determine mechanisms for these protective effects.Patients on chronic hemodialysis served as the study models. Six patients on dialysis three times per week for 6 or more months received sulfinpyrazone 200 mgm t.i.d. p.o. for 14 days. Blood samples were obtained before dialysis was begun before and after the 14 days of drug therapy.Results are shown as mean ± standard error of mean.AT III levels rose significantly by functional and immune assays. Functional levels (by von Kaulla technique) rose 24.5 ± 3.1 sec. to 47.3 + 5.5 sec. (P>.005) Plasma protein AT III (by radial immunodiffusion) rose 31.2 ± 2.17 mg/dl to 37.9 ± 2.1 mgm/dl. (P>.01) Platelet factor 4 (by Abbot radioimmunology assay) fell from 46.4 + 13.6 ngm/ml to 9.5 ± 1.1 ngm/ml.(P>.005) The concentration of thrombin-anti-thrombin complex (by R. Rosenberg, Harvard Medical School, Boston) rose from 4.2 ± .09 to 8.4 ± 1.0 (P>.005)Thus it appears that sulfinpyrazone elevates antithrombin concentration and function while simultaneously suppressing platelet release. These two effects may or may not be mutually dependent. The clinical efficacy of sulfinpyrazone may relate in part to the elevation of antithrombin III, probably by inhibiting its consumption, while also inhibiting platelet function.

Antithrombin III Infusion During Fulminant Hepatic Failure

1981 ◽  
Author(s):  
S Braude ◽  
J Arias ◽  
R D Hughes ◽  
J Canalese ◽  
A E S Gimson ◽  
...  
Keyword(s):  
Fulminant Hepatic Failure ◽  
Hepatic Failure ◽  
Hepatic Coma ◽  
Antithrombin Iii ◽  
Platelet Destruction ◽  
At Iii ◽  
Heparin Anticoagulation ◽  
Low Levels ◽  
Initial Bolus

The antithrombin III (ATIII) levels in 17 patients with fulminant hepatic failure due to viral hepatitis or paracetamol overdose were found to be 25.8%±SD 12.80 of normal on admission. The levels did not correlate with eventual survival or death and remained essentially unchanged for up to 7 days.In an attempt to assess the role of the low levels of AT III during the course of hepatic failure and in relation to treatment by charcoal haemoperfusion we have infused patients with commercially purified ATIII. Preliminary measurements of ATIII (chromogenic substrate method) were made and ATIII infused to achieve a plasma concentration of 50 to 70%. Infusion was by an initial bolus of 1500-2000 units followed by up to 500 units every 6 hours. To date 3 patients in Grade IV hepatic coma have been treated, one died 1 day after admission and the other two survived. In the latter the return of the prothrombin time to normal was similar to that in patients without the addition of ATIII. In one of the survivors the platelet count did not fall, suggesting ATIII may have had a protective effect on platelet consumption. There was also an indication, that there was a more uniform and better response to heparin anticoagulation during haemoperfusion than found previously without ATIII infusion.Further patients will be treated to evaluate whether AT III substitution can reduce the consumptive coagulopathy and platelet destruction which occurs in the course of fulminant hepatic failure.

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