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A competitive enzyme-linked immunoadsorbent assay (ELISA) technique has been developed to facilitate quantitative analysis of the earliest step in the initiation of the extrinsic pathway of coagulation, i.e., complex formation of factor VII/VIIa with tissue factor. The ELISA measures the binding of biotinylated human plasma factor VII to relipidated recombinant human tissue factor. Quantitation of the relative affinity (expressed as IC50) of any factor VII molecular population or structural analogue for tissue factor can be determined by competitive binding. Subnanomolar concentrations of both wild-type recombinant human factor VII (rFVII) and rFVII(R152Q), a mutation at the FVII activation site, competed effectively with biotinylated plasma-derived factor VII in binding to tissue factor. In contrast, the affinity of rFVII(R79Q), a mutation in the first epidermal growth factor-like domain, was 12-fold lower. Following activation of rFVII(R79Q), its affinity for tissue factor and enzymatic activity increased 4-fold and 6-fold, respectively. For wild-type rFVII, enzymatic activity rose significantly following activation. However, its affinity for tissue factor was unchanged. We conclude that both the activation state of factor VII and the mutation of amino-acid residues within the first epidermal growth factor-like domain may alter the affinity of factor VII for tissue factor. 相似文献
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To examine the separate effects of viscosity and sweetness on astringency, aqueous solutions of grape seed tannin (GST) were thickened with carboxymethyl cellulose (CMC) from 2 to 45 cP (experiment 1) or sweetened with 0 to 1.8 g/L aspartame (experiment 2). Trained subjects continuously rated astringency and bitterness in duplicate. Subjects were categorized by the salivary flow induced by citric acid and ability to taste n-propyl thiouracil (PROP). In experiment 1, maximum intensity and total duration of astringency were significantly decreased as viscosity rose, although time to maximum intensity of astringency was not affected. Maximum intensity and total duration of bitterness were not significantly affected by increasing viscosity; however, the onset of bitterness was significantly delayed. In experiment 2, increasing sweetness had no affect on any astringency parameter, although maximum intensity of bitterness was significantly decreased. Neither PROP nor salivary flow-status had any effect on perception of bitterness or astringency in either experiment. 相似文献
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The effect of ancrod-induced defibrinogenation on thrombosis and bleeding time was determined in anesthetized rats. Functional plasma fibrinogen levels were reduced 42, 71, 94 and 93% by ancrod doses of 5, 10, 20 and 30 U/kg, respectively, while a 2.5 U/kg dose was without significant effect. Ancrod inhibited vena cava thrombosis induced by partial stasis of blood flow combined with mild vascular injury. Thrombus weight was decreased 85 and 93% by the 10 and 20 U/kg doses, but was unaffected at lower doses. In contrast, ancrod doses of up to 30 U/kg did not significantly decrease carotid artery thrombi formed in response to oxidative transmural vessel injury. Ancrod caused a dose-dependent increase in bleeding time measured by puncturing small mesenteric arteries with a hypodermic needle. The bleeding time increase was approximately 38% in response to the 2.5 and 5 U/kg doses, and 182% in response to the 10 U/kg dose. These studies demonstrate that ancrod-induced reductions in plasma fibrinogen more effectively inhibit venous compared to arterial thrombosis, although these activities require doses that also increase bleeding time in small arteries. 相似文献
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South Korea and Cuba are dissimilar in religion, economy, culture and attitudes toward premarital sexual relations. In 1960, Korea instituted a national family planning programme to combat rapid population growth. Cuba explicitly rejected Malthusian policies, but made family planning universally available in 1974 in response to health needs. Both countries have undergone rapid fertility declines and today have less than replacement level fertility. Both countries have also used a similar mixture of methods, including a high prevalence of female sterilisation. Abortion has played a major role in the fertility decline of both countries, rising in the first half of the fertility transition and then falling, although remaining a significant variable in the second half. It is concluded that access to contraception, voluntary sterilisation, and safe abortion has a direct impact on fertility and has been associated with a rapid fall in family size in two very different countries. 相似文献
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S Chasserot-Golaz P Hubert D Thiersé S Dirrig CJ Vlahos D Aunis MF Bader 《Canadian Metallurgical Quarterly》1998,70(6):2347-2356
Several lines of evidence suggest that phosphorylated products of phosphatidylinositol play critical functions in the regulation of membrane trafficking along the secretory pathway. To probe the possible involvement of phosphatidylinositol 3-kinase (PI 3-kinase) in regulated exocytosis, we have examined its subcellular distribution in cultured chromaffin cells by immunoreplica analysis and confocal immunofluorescence. We found that the PI 3-kinase heterodimer consisting of the regulatory and catalytic subunits was associated essentially with the subplasmalemmal cytoskeleton in both resting and nicotine-stimulated chromaffin cells. Attempts to immunoprecipitate PI 3-kinase with anti-phosphotyrosine antibodies failed, suggesting that the activity of PI 3-kinase was not modulated by tyrosine phosphorylation and/or physical interaction with SH2-containing proteins in stimulated chromaffin cells. LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one], a potent inhibitor of PI 3-kinase, produced a dose-dependent inhibition of catecholamine secretion evoked by various secretagogues. Furthermore, cytochemical experiments with rhodamine-labeled phalloidin revealed that LY294002 blocked the disassembly of cortical actin in chromaffin cells stimulated by a depolarizing concentration of potassium. Our results suggest that PI 3-kinase may be one of the important regulatory exocytotic components involved in the signaling cascade controlling actin rearrangements required for catecholamine secretion. 相似文献
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