Increased Protein Synthesis by Human Platelets during Phagocytosis of Latex Particles in Vitro

1976 ◽  
Vol 35 (02) ◽  
pp. 350-357 ◽  
Author(s):  
Hana Bessler ◽  
Galila Agam ◽  
Meir Djaldetti
Keyword(s):  
Protein Synthesis ◽  
Fold Increase ◽  
Human Platelets ◽  
Polystyrene Latex ◽  
Latex Particles ◽  
The Third ◽  
Platelet Protein ◽  
Dose Dependent ◽  
Phagocytotic Activity

SummaryA three-fold increase of protein synthesis by human platelets during in vitro phagocytosis of polystyrene latex particles was detected. During the first two hours of incubation, the percentage of phagocytizing platelets and the number of latex particles per platelet increased; by the end of the third hour, the first parameter remained stable, while the number of latex particles per cell had decreased.Vincristine (20 μg/ml of cell suspension) inhibited platelet protein synthesis. This effect was both time- and dose-dependent. The drug also caused a decrease in the number of phagocytizing cells, as well as in their phagocytotic activity.

scholarly journals Loss of 111Indium as indicator of platelet injury

Blood ◽  
1981 ◽  
Vol 58 (2) ◽  
pp. 350-353 ◽  
Author(s):  
JH Joist ◽  
RK Baker
Keyword(s):  
Small Molecules ◽  
Adenine Nucleotides ◽  
Human Platelets ◽  
Metabolic Energy ◽  
Platelet Factor ◽  
Platelet Protein ◽  
Threshold Rate ◽  
Immune Injury ◽  
Triton X

Abstract We previously demonstrated that platelets can be labeled with 111Inoxine with high labeling efficiency and that 111In is not liberated from labeled platelets during the platelet release reaction or prolonged in vitro storage. In view of these findings, we examined the potential usefulness of loss of 111In from labeled platelets as an indicator or platelet damage by comparing the loss of 111In with that of 51Cr and LDH (in some experiments also with platelet factor 3 availability) under different conditions of platelet injury. When washed human platelets labeled with either 51Cr-chromate or 111In-oxine were exposed to increasing concentrations of detergents (Triton X-100, lysolecithin), threshold, rate, and extent of loss of 111In, 51Cr and, LDH were similar. In contrast, when labeled platelets were depleted of metabolic energy by incubation in glucose-free Tyrode albumin solution or glucose-depleted plasma in the presence of antimycin A and 2-deoxy-D- glucose, loss of 51Cr (and PF3a) occurred earlier and progressed at a faster rate than that of 111In or LDH. Similar results were obtained when platelets were exposed to increasing concentrations of PlA1 antibody, causing complement-mediated immune injury. The findings indicate that with certain agents that cause rapid platelet disruption (lysis), different platelet constituents are lost at similar rates. However, under conditions of more subtle or slowly progressive platelet injury, small molecules such as adenine nucleotides (51Cr) may escape earlier and at faster rates than larger molecules such as LDH or 111In- binding platelet protein. Thus, neither 111In loss nor LDH loss appear to be suitable indicators for sublytic or prelytic platelet injury.

Endotoxin-Induced Platelet Activation in Human Whole Blood In Vitro

1988 ◽  
Vol 59 (03) ◽  
pp. 378-382 ◽  
Author(s):  
Gyorgy Csako ◽  
Eva A Suba ◽  
Ronald J Elin
Keyword(s):  
Platelet Aggregation ◽  
Whole Blood ◽  
Human Platelet ◽  
Platelet Rich Plasma ◽  
Atp Release ◽  
Human Platelets ◽  
Lag Phase ◽  
Human Whole Blood ◽  
Dose Dependent

SummaryThe effect of purified bacterial endotoxin was studied on human platelets in vitro. In adding up to 1 μg/mL of a highly purified endotoxin, we found neither aggregation nor ATP release in heparinized or citrated human platelet-rich plasma. On the other hand, endotoxin at concentrations as low as a few ng/mL (as may be found in septic patients) caused platelet aggregation in both heparinized and citrated human whole blood, as monitored by change in impedance, free platelet count, and size. Unlike collagen, the platelet aggregation with endotoxin occurred after a long lag phase, developed slowly, and was rarely coupled with measurable release of ATP. The platelet aggregating effect of endotoxin was dose-dependent and modified by exposure of the endotoxin to ionizing radiation. Thus, the activation of human platelets by “solubilized” endotoxin in plasma requires the presence of other blood cells. We propose that the platelet effect is mediated by monocytes and/or neutrophils stimulated by endotoxin.

scholarly journals Effects of insulin in vitro on protein turnover in rat epitrochlearis muscle

1983 ◽  
Vol 210 (2) ◽  
pp. 323-330 ◽  
Author(s):  
W S Stirewalt ◽  
R B Low
Keyword(s):  
Protein Synthesis ◽  
Protein Degradation ◽  
Nitrogen Balance ◽  
Protein Metabolism ◽  
Serum Insulin ◽  
Specific Radioactivity ◽  
Physiological Concentration ◽  
Rat Serum ◽  
The Third

Rates of protein synthesis and degradation were measured in the isolated rat epitrochlearis muscle by radiotracer techniques, by using the specific radioactivity of tRNA-bound amino acid as precursor for protein synthesis. The tissue maintained linear rates of protein synthesis for 3 h of incubation in the presence of amino acids and glucose and in the absence of insulin. Under these conditions, however, the muscles were in negative nitrogen balance, with rates of protein degradation exceeding rates of protein synthesis. Under steady-state conditions of labelling, the specific radioactivities of tRNA-bound leucine, phenylalanine and valine were significantly less than their respective values in the incubation medium, at concentrations in the medium varying from 1 to 10 times those in normal rat serum. Insulin caused a dose- and time-dependent increase in tRNA-based protein synthesis rates, more than doubling rates at 5 and 50 ng of insulin/ml. At the lower, physiological, concentration of insulin, the stimulation of protein synthesis was not observed until the third hour of incubation with the hormone, whereas the rate of protein synthesis at the higher concentration was elevated during the second hour. There were no delays in the stimulation by insulin of glucose conversion into glycogen. The delayed stimulatory effects of insulin on the rate of protein synthesis brought the tissue to a nitrogen balance near zero. The presence of the hormone also prevented the increase in the rate of protein degradation seen in the third hour of incubation in the absence of the hormone. These studies demonstrate the viability of the incubated rat epitrochlearis muscle with respect to protein metabolism and sensitivity to the protein anabolic effects of physiological concentrations of insulin, and indicate that the preparation is a suitable experimental model for the study of the control of protein metabolism in fast-twitch skeletal muscle.

scholarly journals Hormonal stimulation in the exocrine pancreas results in coordinate and anticoordinate regulation of protein synthesis.

1984 ◽  
Vol 99 (5) ◽  
pp. 1569-1574 ◽  
Author(s):  
J Schick ◽  
H Kern ◽  
G Scheele
Keyword(s):  
Protein Synthesis ◽  
Fold Increase ◽  
Response Patterns ◽  
Hormonal Stimulation ◽  
Group Iii ◽  
Hormone Stimulation ◽  
Control Value ◽  
Group I ◽  
Separation Of Proteins

24-h intravenous caerulein infusion studies in the rat were combined with in vitro amino acid incorporation studies followed by high-resolution separation of proteins by two-dimensional isoelectric focusing and SDS gel electrophoresis to study the extent to which persistent changes in the biosynthesis of exocrine pancreatic proteins are regulated by cholecystokinin-like peptides. Beginning in the third hour of optimal hormone infusion at 0.25 microgram kg-1 h-1, changes were observed in the synthetic rates of 12 proteins, which progressed over the course of the 24-h study. Based on coordinate response patterns, exocrine proteins could be classified into four distinct groups. Group I (trypsinogen forms 1 and 2) showed progressive increases in synthetic rates reaching a combined 4.3-fold increase over control levels. Group II (amylase forms 1 and 2) showed progressive decreases in synthesis to levels 7.1- and 14.3-fold lower than control levels, respectively. Group III proteins (ribonuclease, chymotrypsinogen forms 1 and 2, procarboxypeptidase forms A and B, and proelastase 1) showed moderate increases in synthesis, 1.4-2.8-fold, and group IV proteins (trypsinogen 3, lipase, proelastase 2, and unidentified proteins 1-4) did not show changes in synthesis with hormone stimulation. Regulation of protein synthesis in response to caerulein infusion was specific for individual isoenzymic forms in the case of both trypsinogen and proelastase. The ratio of biosynthetic rates of trypsinogen forms 1 + 2 to amylase forms 1 + 2 increased from a control value of 0.56 to 24.4 after 24 h of hormonal stimulation (43.5-fold increase). Biosynthetic rates for an unidentified protein (P23) with an Mr = 23,000 and isoelectric point of 6.2 increased 14.2-fold, and the ratio of synthesis of P23 to amylase 2 increased 200-fold during caerulein infusion. During hormone stimulation the anticoordinate response in the synthesis of pancreatic glycosidases (decreased synthesis) and serine protease zymogens (increased synthesis) explain previous observations that showed little change in rates of total protein synthesis under similar conditions.

scholarly journals Citrulline directly modulates muscle protein synthesis via the PI3K/MAPK/4E-BP1 pathway in a malnourished state: evidence from in vivo, ex vivo, and in vitro studies

2017 ◽  
Vol 312 (1) ◽  
pp. E27-E36 ◽  
Author(s):  
Servane Le Plénier ◽  
Arthur Goron ◽  
Athanassia Sotiropoulos ◽  
Eliane Archambault ◽  
Chantal Guihenneuc ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Ex Vivo ◽  
Muscle Protein ◽  
Muscle Protein Synthesis ◽  
Fold Increase ◽  
Underlying Mechanism ◽  
Muscle Homeostasis

Citrulline (CIT) is an endogenous amino acid produced by the intestine. Recent literature has consistently shown CIT to be an activator of muscle protein synthesis (MPS). However, the underlying mechanism is still unknown. Our working hypothesis was that CIT might regulate muscle homeostasis directly through the mTORC1/PI3K/MAPK pathways. Because CIT undergoes both interorgan and intraorgan trafficking and metabolism, we combined three approaches: in vivo, ex vivo, and in vitro. Using a model of malnourished aged rats, CIT supplementation activated the phosphorylation of S6K1 and 4E-BP1 in muscle. Interestingly, the increase in S6K1 phosphorylation was positively correlated ( P < 0.05) with plasma CIT concentration. In a model of isolated incubated skeletal muscle from malnourished rats, CIT enhanced MPS (from 30 to 80% CIT vs. Ctrl, P < 0.05), and the CIT effect was abolished in the presence of wortmannin, rapamycin, and PD-98059. In vitro, on myotubes in culture, CIT led to a 2.5-fold increase in S6K1 phosphorylation and a 1.5-fold increase in 4E-BP1 phosphorylation. Both rapamycin and PD-98059 inhibited the CIT effect on S6K1, whereas only LY-294002 inhibited the CIT effect on both S6K1 and 4E-BP1. These findings show that CIT is a signaling agent for muscle homeostasis, suggesting a new role of the intestine in muscle mass control.

Hemostatic Properties of Infusible Trehalose-Stabilized Lyophilized Platelet Derivatives.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 834-834
Author(s):  
Keith A. Moskowitz ◽  
Josh Dee ◽  
Jason Barnidge ◽  
Ruth Sum ◽  
David Ho ◽  
...  
Keyword(s):  
Fluid Phase ◽  
Fold Increase ◽  
Submicron Particles ◽  
Cell Counting ◽  
Attractive Alternative ◽  
Dependent Manner ◽  
Clot Retraction ◽  
Drug Induced ◽  
Dose Dependent

Abstract Availability of platelet concentrates for treatment of bleeding associated with thrombocytopenia, trauma, or drug-induced coagulopathies is problematic due to the short 5 day platelet storage time and because platelets require controlled shaking at ambient temperature in order to remain viable, a condition which augments bacterial growth. To address the platelet availability problem we expanded upon trehalose cryo-preservation technology to create a lyophilized hemostatic platelet derivative. Washed platelets were stabilized by accumulation of 5–10 mM intracellular trehalose via fluid phase endocytosis then formulated with excipients and lyophilized. Lyophilized platelets were instantaneously rehydrated with > 90% recovery and were stable for at least 3–6 months at ambient temperatures. Rehydrated (RH) platelets responded quantitatively to α-and γ-thrombin and ristocetin by transmittance aggregometry and were partially agglutinated by collagen as judged by aggregometry and single cell counting using the Platelet Works® system. RH platelets co-aggregated in a dose dependent manner when mixed with fresh autologous platelets during collagen-induced activation. Aggregation response to low-dose thrombin and collagen was inhibited by the GPIIb/IIIa antagonist RGDS and by EGTA. RH platelets were quantitatively incorporated into fibrin clots and elicited platelet-dependent fibrin-clot retraction ~ 60% as well as fresh platelets. RH platelets were similar in size to fresh and had less than 25% submicron particles as judged by electronic particle counting and flow cytometry scatter profiles. RH platelets were partially activated upon rehydration as judged by anti P-selectin and anti-LAMP-3 binding, yet GPIIb/IIIa remained in a resting conformation, as judged by a lack of PAC-1 binding. GPIIb/IIIa receptors were present as judged by the binding of complex-dependent (clone 5B12) and function-blocking (clone P2) antibodies. RH platelets also contained intact GPIbα as judged by binding of the function-blocking MoAb AN51. Function of GPIIb/IIIa and collagen receptors on RH platelets was further demonstrated as RH platelets adhered to immobilized fibrinogen and collagen in the absence of added agonists and in a dose-dependent manner. Moreover, RH platelets exhibited a two-fold increase in platelet procoagulant activity in the presence of thrombin receptor agonist peptide SFLLRN as judged by Annexin-V binding. Procoagulant and hemostatic activity was further demonstrated as RH platelets accelerated the clotting of recalcified whole thrombocytopenic blood in a dose-dependent manner similarly to fresh platelets. Lastly, RH platelets corrected the coagulopathy induced by contact pathway inhibition with aprotinin during the recalcification of citrated whole blood. The technology has been scaled to single donor platelet aphaeresis units, equivalent to a standard transfusion dose. Preclinical animal models of safety, efficacy, and circulation persistence are currently being evaluated. In summary, trehalose- stabilized lyophilized platelet derivatives contain numerous in vitro hemostatic properties and may offer an attractive alternative to fresh platelet transfusions when the latter are indicated yet unavailable.

scholarly journals Platelet activation by immobilized monoclonal antibody: evidence for a CD9 proximal signal

Blood ◽  
1991 ◽  
Vol 78 (7) ◽  
pp. 1753-1759
Author(s):  
L Griffith ◽  
J Slupsky ◽  
J Seehafer ◽  
L Boshkov ◽  
AR Shaw
Keyword(s):  
Platelet Aggregation ◽  
Hla Class I ◽  
Human Platelets ◽  
Dense Granule ◽  
Polystyrene Latex ◽  
Cross Linking ◽  
Latex Beads ◽  
Dose Dependent ◽  
Aggregation Of Platelets ◽  
Granule Release

Anti-CD9 monoclonal antibodies (MoAbs) are reported to activate human platelets through stimulation of the Fc gamma II receptor. We show here that nonstimulatory F(ab')2 fragments of the anti-CD9 MoAb 50H.19 induce dense-granule release and dose-dependent platelet aggregation when attached to polystyrene latex beads. Cross-linking F(ab')2 fragments of MoAb 50H.19 by F(ab')2 fragments of goat anti-mouse IgG does not result in platelet aggregation unless the second antibody is bound to latex beads, indicating that immobilization, and not cross- linking of the stimulus, is critical to the initiation of the CD9 signal. In contrast, F(ab')2 fragments of the second antibody readily induce the aggregation of platelets treated with the anti-Fc gamma II receptor MoAb IV.3. Immobilization of MoAb per se is insufficient to induce an activation signal because intact and F(ab')2 fragments of nonstimulatory MoAb directed to glycoprotein Ib and HLA class I do not become stimulatory when attached to beads. CD9-induced activation requires cytoskeletal rearrangement because it is inhibited by cytochalasin B. Aggregation is blocked by inhibitors of the thromboxane pathway, indicating that CD9 activates phospholipase C indirectly through prior activation of phospholipase A2.\

The Inhibitory Effect of GR32191, a Thromboxane Receptor Blocking Drug, on Human Platelet Aggregation, Adhesion and Secretion

1989 ◽  
Vol 61 (03) ◽  
pp. 429-436 ◽  
Author(s):  
E J Hornby ◽  
M R Foster ◽  
P J McCabe ◽  
L E Stratton
Keyword(s):  
Platelet Aggregation ◽  
Thrombus Formation ◽  
Rabbit Aorta ◽  
Inhibitory Effect ◽  
Human Platelets ◽  
Camp Phosphodiesterase ◽  
Platelet Protein ◽  
Thromboxane Receptor ◽  
Serotonin Secretion

SummaryGR32191, a potent selective thromboxane receptor antagonist, has been shown to inhibit completely prostaglandin endoperoxide and thromboxane A2 (TxA2)-induced platelet aggregation, [14C]-serotonin secretion and β-thromboglobulin secretion. Deposition of human platelets onto damaged rabbit aorta in vitro is reduced in the presence of GR32191 which appears to inhibit aggregation of platelets but not direct adhesion of platelets to subendothelium. The effects of non-prostanoid platelet activating agents whose mode of action requires the biosynthesis of TxA2 are also inhibited by GR32191. Prostanoids which inhibit platelet function, such as prostacyclin or PGD2, retain their inhibitory properties in the presence of GR32191 which does not inhibit phospholipase A2, prostaglandin cyclooxygenase, thromboxane synthase, 12-lipoxygenase or cAMP phosphodiesterase activity. The inhibitory action of GR32191 on platelet aggregation, mural thrombus formation and platelet protein storage granule secretion suggests that it has potential in treatingthrombotic disease in man.

scholarly journals Platelet activation by immobilized monoclonal antibody: evidence for a CD9 proximal signal

Blood ◽  
1991 ◽  
Vol 78 (7) ◽  
pp. 1753-1759 ◽  
Author(s):  
L Griffith ◽  
J Slupsky ◽  
J Seehafer ◽  
L Boshkov ◽  
AR Shaw
Keyword(s):  
Platelet Aggregation ◽  
Hla Class I ◽  
Human Platelets ◽  
Dense Granule ◽  
Polystyrene Latex ◽  
Cross Linking ◽  
Latex Beads ◽  
Dose Dependent ◽  
Aggregation Of Platelets ◽  
Granule Release

Abstract Anti-CD9 monoclonal antibodies (MoAbs) are reported to activate human platelets through stimulation of the Fc gamma II receptor. We show here that nonstimulatory F(ab')2 fragments of the anti-CD9 MoAb 50H.19 induce dense-granule release and dose-dependent platelet aggregation when attached to polystyrene latex beads. Cross-linking F(ab')2 fragments of MoAb 50H.19 by F(ab')2 fragments of goat anti-mouse IgG does not result in platelet aggregation unless the second antibody is bound to latex beads, indicating that immobilization, and not cross- linking of the stimulus, is critical to the initiation of the CD9 signal. In contrast, F(ab')2 fragments of the second antibody readily induce the aggregation of platelets treated with the anti-Fc gamma II receptor MoAb IV.3. Immobilization of MoAb per se is insufficient to induce an activation signal because intact and F(ab')2 fragments of nonstimulatory MoAb directed to glycoprotein Ib and HLA class I do not become stimulatory when attached to beads. CD9-induced activation requires cytoskeletal rearrangement because it is inhibited by cytochalasin B. Aggregation is blocked by inhibitors of the thromboxane pathway, indicating that CD9 activates phospholipase C indirectly through prior activation of phospholipase A2.\

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