Contact with surfaces results in activation of formed blood elements. This study demonstrates that during extracorporeal circulation, release of lysosomal enzymes occurs concomitantly with platelet ...granule secretion. Fresh, heparinized human blood was recirculated for 2 hours at 1,000 ml/min at 37 degrees C in silicone rubber circuits containing a membrane oxygenator (0.85 m2). The plasma levels of a platelet-specific protein, low affinity platelet factor 4 (LA-PF4), rose from less than 0.5 to 16 +/- 3 SEM microgram/ml plasma, indicating extensive release of platelet alpha granule contents. Concurrently, plasma activity of acid phosphatase and N-acetyl-beta-glucosaminidase increased nearly fivefold. Platelet inhibition prevented release of LA-PF4 and reduced acid phosphatase levels, but failed to alter the levels of N-acetyl-beta-glucosaminidase. Lidocaine (10 to 20 microgram/ml), however, prevented the rise in N-acetyl-beta-glucosaminidase activity and diminished acid phosphatase levels without altering secretion of LA-PF4. The failure of platelet inhibitors to block the increase in N-acetyl-beta-glucosaminidase, and the efficacy of lidocaine to do so without altering LA-PF4 secretion, suggest that leukocytes, not platelets, are the primary source of this lysosomal enzyme. Although acid phosphatase activity measured in plasma my be derived from leukocytes, platelet membranes, and not lysosomes, appear to be a more likely source. Release of hydrolytic enzymes from formed blood elements during cardiopulmonary bypass may cause endothelial cell injury and thus contribute to the increased vascular permeability associated with extracorporeal circulation.
We evaluated the ability of the calcium antagonist, verapamil, to alter human platelet function. With the concentrations tested (20 ng to 0.1 mg/ml of whole blood), verapamil inhibited ...epinephrine-induced aggregation and release of 14C-serotonin; produced a dose-dependent inhibition of 14C-serotonin uptake and prevented aggregation dependent release of thromboxane B2. The action of verapamil could be overcome by higher concentrations of both epinephrine and calcium. Furthermore, verapamil-induced inhibition could be reversed by gel-filtering platelets suggesting that verapamil's anti-platelet activity does not outlast its presence in plasma. Verapamil was relatively ineffective as an inhibitor of ADP-induced aggregation. As with epinephrine-induced platelet activation, the effects of verapamil on ADP-induced 14C-serotonin and thromboxane release correlated with its effects on secondary aggregation. Finally, verapamil failed to alter calcium ionophore-induced platelet aggregation. Thus, verapamil at the concentrations tested, appears to be functioning as a reversible, relatively specific inhibitor of epinephrine-induced platelet activation. Our findings suggest that the actions of verapamil in this regard are complex; there may be competitive inhibition of epinephrine binding as well as a blockade of epinephrine-induced calcium flux.
The corpus luteum of the rat stores large quantities of cholesteryl ester which is synthesized by acyl coenzyme A:cholesterol acyltransferase (ACAT), a microsomal enzyme. In previous studies of ACAT, ...assays that were probably substrate limited were employed. To reevaluate the regulation of ACAT in the ovaries of PMSG-hCG-primed rats, we used an improved assay in which exogenous cholesterol is provided as a dispersion in Triton WR-1339. Assayed in the absence of exogenous cholesterol, ACAT activity increased between days 1--8 post-hCG treatment and then declined between days 8--15. When exogenous cholesterol was included, a similar pattern of ACAT activity was found, but rates of cholesteryl ester formation were 2.5- to 4.5-fold greater. These changes in ACAT paralleled the previously reported levels of ovarian cholesteryl esters. Treatment of rats with 4-aminopyrazolopyrimidine, a drug that lowers blood cholesterol levels and reduces ovarian sterol ester stores, resulted in a lowered ACAT activity measured in the absence of exogenous sterol. However, enzyme activity was similar to that in controls when assayed in the presence of cholesterol. Inhibitors of steroidogenesis (aminoglutethimide and cycloheximide) promoted, within 4 h, ovarian sterol ester storage and resulted in increased ACAT activities measured in the absence of cholesterol. However, in the presence of exogenous sterol, the ACAT activities of controls were equal to those of drug-treated animals. When PMSG-hCG-primed animals received iv injections of hCG on day 8 post-hCG, ovarian sterol ester stores were markedly depleted within 2 h. The ovarian ACAT activity of hCG-treated rats measured without cholesterol was significantly lower than that of controls. With cholesterol, ACAT activities of hCG-treated rats were similar to those in controls. Our findings indicate that the entry of cholesterol into the ACAT substrate pool may be a major factor controlling the rates of cholesteryl ester synthesis in the rat corpus luteum.
Platelet secretion induced by certain soluble aggregating agents is associated with thromboxane formation. Activation of platelets by artificial surfaces may involve similar biochemical pathways. ...Therefore, we monitored platelet release and formation of thromboxane B2. the stable end-product of thromboxane A2, during simulated extracorporeal circulation where contact between blood and synthetic surfaces is extensive. Fresh heparinized human blood was recirculated for 2 hr at 37°C at 1 liter/min in circuits (0.95 sqm) constructed of standard silicone rubber components (0.1 sqm) and a spiral coil membrane oxygenator (0.85 sqm). Within 2 min of recirculation, plasma levels of the platelet-specific protein, low-affinity platelet factor 4 (LA-PF4) rose from <1.5% to 11 % ±3% SEM of that which was released by triton X-100, indicating significant release of platelet α-granule contents. This occurred despite the absence of detectable thromboxane B2 in plasma (<0.5 pmole/ml). Between 2 and 15 min, plasma levels of LA-PF4 increased from 11% to 42% ± 3%, and plasma thromboxane B2 concentrations rose to 3.2 pmole/ml. Subsequently, thromboxane B2 levels continued to rise to 9.8 pmole/ml by 2 hr. Thromboxane B2 was not detected in blood from donors who had ingested aspirin, and the extent of platelet secretion was reduced by about 50% (25% at 2 hr). Thus, although a component of platelet granule release during extracorporeal recirculation appears to be associated with thromboxane synthesis, considerable release of LA-PF4 may occur by thromboxanβ-indβpen-dent pathways.
The presence of 92 kDa type-IV collagenase (MMP-9) was documented in human amniochorion collected from women with active labor or during caesarean sections of term pregnancies. MMP-9 immunoreactive ...protein and mRNA were detected mainly in amnion epithelium and decidual cells, but also some of the trophoblast cells of the spongy layer depicted positivity. Gelatin substrate gels from amniochorion extracts of samples collected after delivery, showed the presence of at least 5 different gelatinases ranging from 150 to 68 kDa. The main activity appeared in a 92 kDa band. No activity was detected in fetal membranes collected before onset of labor, however, Western blot analyses confirmed the presence of MMP-9 in these samples. The latent (92 kDa) and active (83 kDa) forms were detected In extracts of amniochorion collected after delivery. Activation of the latent MMP-9 enzyme with stromelysin (MMP-3) was observed in both groups using gelatin gels. We conclude that the existence of a labor-related potentially physiological mediator of chorioamniotic membranes rupture can be confirmed in this study. The presence of 92 kDa type IV collagenase in amnion epithelium, chorion decidual and trophoblast cells from term fetal membranes suggests their role in extracellular matrix components degradation as a mechanism of chorio-amnios weakening during both physiological and pathological conditions. Our results suggest that this process is linked to the activation of previously existent latent MMP-9, possibly by stromelysin
Temporary inhibition of platelet function with prostaglandin E1 (PGE1) prevents platelets loss and functional alterations during extracorporeal circulation with a membrane oxygenator. This study ...evaluated the ability of PGE1 to prevent platelet injury in circuits containing a bubble oxygenator. During in vitro recirculation of human blood, the circulating platelet count, expressed as a percent of initial levels, decreased to 29%; platelets became insensitive to adenosine diphosphate (ADP) and to epinephrine; and plasma levels of low-affinity platelet factor 4 (LA-PF4) progressively rose to 7.4 microgram per milliliter. With PGE1 (1.2 micron), platelet counts remained stable at 92%; platelet reactivity remained normal for 1 hour; and plasma levels of LA-PF4 rose to only 3.3 microgram per milliliter. In rhesus monkeys that underwent cardiopulmonary bypass using a bubble oxygenator without PGE1, platelet counts, expressed as a percent of the prebypass platelet count, declined to 38%; platelets became insensitive to ADP; plasma levels of LA-PF4 progressively rose to 8.4 microgram per milliliter; and the mean postoperative bleeding time was 4.6 minutes. In monkeys that received PGE1, platelet counts declined to only 65%; platelets remained threefold more sensitive to ADP; platelets demonstrated delayed release of LA-PF4 and the mean postoperative bleeding time was 2.7 minutes. This report demonstrates that in a bubble oxygenator system, PGE1 reduces platelet loss, mitigates platelet injury, and shortens postoperative bleeding times following extracorporeal circulation.
The National Centers of Excellence in Women's Health Program (CoE) represents a new model for women's health in academic health centers that unites women's health research, teaching, clinical care, ...public education and outreach, and career advancement for women in the health sciences. Lessons learned from the first 3 years of implementation indicate that this type of model requires a transformation from the traditionally fragmented set of activities in academic health centers to an integrated system united around the goal of advancing women's health. The transformation requires institutional commitment, dedicated players, and an ability to build on existing resources and bring added value to the institution. Challenges and strategies to link women's health activities and increase collaboration are also discussed.
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