Blood separation with two different autotransfusion devices: effects on blood cell quality and coagulation variables

The successful synthesis of 17 nitrosimines of 2 H-1,3,4-thiadiazines is reported. They are best characterized by their electronic spectra (lambda max = 504-522 nm). Some of the compounds were able to inhibit the aggregation of blood platelets in the Born-test (inducer collagen). The most active compound was the 3-nonyl-5-phenyl-derivative 10c which showed an IC50 = 6.5 mumol.l-1. In mesenteric arterioles of rats after p.o. administration (60 mg/kg) a 13% inhibition of thrombus formation is observed. The reason for the rather small effects is the high chemical stability of the title compounds and the fact that decomposition occurs mainly by the evolution of molecular nitrogen. Only small yields of NO are formed.     

7-Deazapurine 2'-deoxyribofuranosides are noncleavable competitive inhibitors of Escherichia coli purine nucleoside phosphorylase (PNP)

A series of 7-deazapurine 2'-deoxyribofuranosides were synthesized according to already known procedures and their substrate and inhibitor properties with purified E. coli purine nucleoside phosphorylase were examined. In agreement with previous findings, substrate activity was not detected for any of the compounds tested. Most of the nucleosides showed weak inhibition in the preliminary screening, i.e. at a concentration of about 100 microM. However some combinations of 6-chloro, 6-amino or 6-methoxy substituents with bulky hydrophobic groups at position 7 of the base and/or chloro, amino, methoxy or methylthio group at position 2 markedly enhanced affinity of such modified nucleosides for the E. coli enzyme. The most potent inhibition was observed for two nucleosides: 6-chloro- and 2-amino-6-chloro-7-deazapurine 2'-deoxyribofuranosides that show inhibition constants Ki = 2.4 and 2.3 microM, respectively. Several other compounds were also found to be good inhibitors, with inhibition constants in the range 5-50 microM. In all instances the inhibition was competitive vs. the nucleoside substrate 7-methylguanosine. Inhibition constants for 7-deazapurine nucleosides are in general several-fold lower than those observed for their purine counterparts. Therefore 7-deaza modification together with substitutions at positions 2, 6 and 7 of the base is a very promising approach to obtain competitive noncleavable inhibitors of E. coli PNP that may bind to the enzyme with inhibition constants in the microM range.     

New NO-donors with antithrombotic and vasodilating activities, Part 14. 1,3,4-Triazol-1-oles

Five 1,3,4-triazol-1-oles (5a-f) with different alkyl, aryl, and arylalkyl substituents in 2,5-position were synthesized and tested for their antithrombotic properties. The 2,5-dimethyl derivative 5a was most active. 2 h after administration of 60 mg/kg to rats thrombus formation by a laser beam was inhibited by 42% in arterioles and by 33% in venules. At the same dose the blood pressure of SHR rats was slightly (5%) but significantly decreased even 4 h after application of 5a. This pattern of activities suggests a nitric oxide mediated mechanism of action. 1,1'-Azo-bis-ethanone oxime(7)-the synthetic precursor of 5a-inhibited the aggregation of blood platelet (Born test) with an IC50 = 15 mumol/L.     

Potent cyano and carboxamido side-chain analogues of 1', 1'-dimethyl-delta8-tetrahydrocannabinol

The synthesis and pharmacological profile of several cyano (1a-e) and carboxamido (2a-h) side-chain-substituted analogues of 1', 1'-dimethyl-Delta8-THC are described. Commercially available cyano compound 3 was transformed to the resorcinol 6 in a three-step sequence. Condensation of 6 with p-menth-2-ene-1,8-diol formed the THC 7a which, with sodium cyanide/DMSO, gave 1b. Protection of the phenol in 7a as the MOM derivative provided the common intermediate 8 for the synthesis of 1a,c,e. Compound 1d was also synthesized from 7a via the aldehyde 9a. Base hydrolysis of 1b gave the acid 10 which, via its acid chloride and subsequent treatment with the appropriate amine, formed the target compounds 2a-h. The pharmacological profile indicated that the cyano analogues 1a-e had very high CB1 binding affinity (0.36-13 nM) and high in vivo potency as agonists. Two analogues (1a,b) had extremely high potency in the mouse tetrad tests. The dimethylcarboxamido analogue 2a showed a similar profile to 1a,b. The high potency was also retained in analogue 2c. In contrast the sulfonamide analogue 2d was unique as it had greater affinity than Delta9-THC, yet it was practically devoid of agonist effects. This study suggests that the incorporation of a cyano or an amide substituent in the side chain of Delta8-THC-DMH can enhance potency and can also lead to compounds with a unique profile which have high binding affinity and are practically devoid of agonist effects.     

Structure-activity relationships of N2-aryl-3-(isoxazolylsulfamoyl)-2-thiophenecarboxamides as selective endothelin receptor-A antagonists

We report here that N2-aryl-3-(isoxazolylsulfamoyl)-2-thiophenecarboxamides are potent and selective small molecule ETA receptor antagonists. The aryl group was subjected to extensive structural modification. With monosubstitution, the para position was most useful in increasing potency, with methyl being preferred. With disubstitution, 2,4-disubstitution further enhanced activity with methyl or cyano groups being preferred at the 2-position. In this series, a benzo-[d][1,3]dioxole group is equivalent to a 4-methyl group in in vitro activity and afforded the compounds with both in vivo activity and moderate half-lives.     

Altered microvascular responses to C5a in rat striated muscle during two-kidney, one clip renovascular hypertension

OBJECTIVE: To determine how two-kidney, one clip (2-K,1C) renovascular hypertension alters microvascular responses in rat striated muscle to complement C5a, one of the most important inflammatory mediators. METHODS: 2-K,1C hypertension was induced in male Sprague-Dawley rats. Under anesthesia with pentobarbital (50 mg/kg, intraperitoneally) the cremaster muscle microcirculatory preparation with intact neurovascular connections was studied in vivo by closed-circuit videomicroscopy. Recombinant human C5a was applied topically in the tissue bath at concentrations of 10(-12), 10(-10) and 10(-8) mol/l, consecutively. Changes in the microvessel diameters in small arterioles, large arterioles and venules were measured. RESULTS: In normotensive rats complement C5a induces a significant dilation in small arterioles at low bath concentrations (10(-12) or 10(-10) mol/l), but the dilation is attenuated at a higher concentration (10(-8) mol/l). In contrast, in 2-K,1C hypertensive rats C5a constricts small arterioles at low concentrations (< 10(-10) mol/l) but dilates them at a higher concentration (10(-8) mol/l). Large arterioles and venules have minimal responses to C5a in either normotensive or 2-K,1C hypertensive rats. CONCLUSION: 2-K,1C hypertension dramatically alters C5a-induced microvascular responses in small arterioles. The alteration might be attributable to the enhanced vasoconstrictor mechanisms and impaired vasodilator mechanisms during 2-K,1C renovascular hypertension.     

Synthesis and biological investigations of 1-(tetrahydropyran-2'-yl)- and 1-(tetrahydrofuran-2'-yl)benzimidazoles and 1/2-(tetrahydropyran-2'-yl)- and 1/2-(tetrahydrofuran-2'-yl)benzotriazoles

Sets of benzimidazole and benzotriazole derivatives bearing on position 1 or 2 a tetrahydrofuranyl or tetrahydropyranyl moieties were prepared through the addition of the suitable benzazoles on 2,3-dihydrofuran and 3,4-dihydro-2H-pyran. The reactions were carried on either without solvent or in carbon tetrachloride solution. In the last case some peculiar chlorinated side products were isolated and characterized. Twenty compounds were screened for in vitro antitumoral and anti-HIV-1 activities and found poorly active or completely inactive. On the other hand several compounds exhibited good tracheal relaxant activity in vitro; compound 8, 11, 16, 24 and 26 resulted more active than theophylline in this test, while compound 11 was comparable to amrinone till the concentration of 3 micrograms/ml. Finally, compound 5 resulted endowed with a strong diuretic and saluretic activity at the dose of 3 mg/Kg, thus representing a new lead for discovering new diuretic agents.     

Nitric oxide reduces tumor cell adhesion to isolated rat postcapillary venules

Adhesion of circulating tumor cells to microvascular endothelium plays an important role in tumor metastasis to distant organs. The purpose of this study was to determine whether nitric oxide (NO) would attenuate tumor cell adhesion (TCA) to naive or lipopolysaccharide (LPS)-treated postcapillary venules. A melanoma cell line, RPMI 1846, was shown to be much more adhesive to postcapillary venules isolated from rat mesentery than to corresponding precapillary arterioles. Although venules exposed to LPS for 4 h demonstrated an increased adhesivity for the melanoma cells, TCA to LPS-treated arterioles was not altered. Isolated venules exposed to DETA/NO (1 mM), an NO donor, for 30 min prior to tumor cell perfusion prevented the increment in adhesion induced by LPS and attenuated TCA to naive postcapillary venules. While L-arginine (100 microM), an NO precursor, failed to decrease TCA to naive postcapillary venules, this treatment abolished LPS-stimulated TCA to postcapillary venules. The effect of L-arginine was reversed by administration of N(omega)-nitro-L-arginine methyl ester (L-NAME, 100 microM), an NO synthase (NOS) inhibitor. These observations indicate that both exogenous and endogenous NO modulate TCA to postcapillary venules. To assess the role of NO-induced activation of cGMP in the reduction in TCA produced by DETA/NO, two additional series of experiments were conducted. In the first series, LY-83583 (10 microM), a guanylyl cyclase inhibitor, was shown to completely reverse the effect of DETA/NO on TCA to both naive and LPS-activated postcapillary venules. On the other hand, administration of 8-bromoguanosine 3',5'-cyclic monophosphate (8-B-cGMP) (1 mM), a cell permeant cGMP analog, mimicked the effect of DETA/NO and reduced TCA to LPS-stimulated postcapillary venules. These data suggest that (a) tumor cells are more likely to adhere to postcapillary venules than to corresponding precapillary arterioles, (b) LPS enhances TCA to postcapillary venules, (c) both exogenously applied (DETA/NO) and endogenously generated (L-arginine) NO attenuate the enhanced adhesion induced by LPS, but only DETA/NO reduced TCA to naive postcapillary venules, and (d) the NO-induced reduction in TCA to LPS-activated postcapillary venules occurs by a cGMP-dependent mechanism.     

Dihydrobenzofuran analogues of hallucinogens. 3. Models of 4-substituted (2,5-dimethoxyphenyl)alkylamine derivatives with rigidified methoxy groups

Tetrahydrobenzodifuran functionalities were employed as conformationally restricted bioisosteres of the aromatic methoxy groups in prototypical hallucinogenic phenylalkylamines 1 and 2. Thus, a series of 8-substituted 1-(2,3,6,7-tetrahydrobenzo[1,2-b:4,5-b']difuran-4-yl)-2-aminoal kanes (7a-e) were prepared and evaluated for activity in the two-lever drug discrimination paradigm in rats trained to discriminate saline from LSD tartrate (0.08 mg/kg) and for the ability to displace [3H]ketanserin from rat cortical homogenate 5-HT2A receptors and [3H]-8-OH-DPAT from rat hippocampal homogenate 5-HT1A receptors. In addition, 1-(8-bromo-2,3,6,7-tetrahydrobenzo[1,2-b:4,5-b']difuran-4-yl)-2-am inopropane (7b), which was found to be extremely potent in the rat in vivo assays, was evaluated for its ability to compete with [125I]DOI and [3H]ketanserin binding to cells expressing cloned human 5-HT2A, 5-HT2B, and 5-HT2C receptors. All of the dihydrofuranyl compounds having a hydrophobic substituent para to the alkylamine side chain had activities in both the in vitro and in vivo assays that equaled or surpassed the activity of the analogous conformationally flexible parent compounds. For example, 7b substituted for LSD in the drug discrimination assay with an ED50 of 61 nmol/kg and had Kj values in the nanomolar to subnanomolar range for the displacement of radioligand from rat and human 5-HT2 receptors, making it one of the most potent hallucinogen-like phenylalkylamine derivatives reported to date. The results suggest that the dihydrofuran rings in these new analogues effectively model the active binding conformations of the methoxy groups of the parent compounds 1 and 2. In addition, the results provide information about the topography and relative orientation of residues involved in agonist binding in the serotonin 5-HT2 receptors.     

A case of primary splenic large cell lymphoma with a t(9;14)(p13;q32)

Appropriately substituted 2,3-dihydro-7H-thiazolo[3,2-a]pyrimidin-7-ones 9-12 and 18 were considered as annulated analogues of HEPT (1-[(2-hydroxyethoxy)methyl]-6-(phenylthio)-thymine), and some of these compounds were also found active against HIV-1, the most active one being 2,3-dihydro-5-[(3,5-dimethylphenyl)methyl]-3-ethoxy-6-ethyl-7H- thiazolo[3,2-a]pyrimidin-7-one (10b). S-Alkylation of 5-alkyl-6-(arylmethyl)-2-thiouracils 1-4 was performed with 2-bromoacetaldehyde acetals to furnish the S-[bis(alkoxy)ethyl] derivatives 5-8 and with allyl bromide to furnish S-allyl derivatives 17. The target compounds 9-12 were obtained by an N1 regioselective intramolecular cyclization reaction of silylated 5-8 using trimethylsilyl trifluoromethanesulfonate (TMS triflate) as the catalyst. Treatment of the S-allyl derivatives 17 with bromine in dry methylene chloride afforded the 3-(bromomethyl) derivatives 18.     

Synthesis and antimicrobial activity of some novel 2,5- and/or 6-substituted benzoxazole and benzimidazole derivatives

A new series of 2,5- and/or 6-substituted benzoxazoles (7a-f), benzimidazoles (8a-g) holding cyclohexyl or cyclopentyl moieties at position 2 and 5- or 6-substituted-2-cyclohexylaminomethylbenzoxazoles (9a, b) was synthesized in order to determine their antimicrobial activities and feasible structure-activity relationships. The synthesized compounds were tested in vitro against three Gram-positive, two Gram-negative bacteria and the yeast Candida albicans in comparison with several control drugs. Microbiological results showed that the synthesized compounds were possessing a broad spectrum of antibacterial activity against the tested microorganisms. 5-Chloro-2-(2-cyclohexylethyl)benzimidazole (8g) was found as the most active compound against the screened Gram-positive bacteria strains at a minimum inhibitory concentration (MIC) value of 12.5 microg/ml. However, it exhibited lower antibacterial potency than the compared control drugs. On the other side, compounds 7-9 indicated significant antibacterial activity against the Gram-negative enterobacter Pseudomonas aeruginosa having MIC values of 50 microg/ml, providing either the same effect as tetracycline or higher activity than streptomycin, but showing less potency than the compared control drug gentamycin. Moreover, the synthesized compounds also possessed antimycotic activity against the yeast C. albicans showing MIC values between 25-50 microg/ml.     

Carbocyclic analogues of the potent cytidine deaminase inhibitor 1-(beta-D-ribofuranosyl)-1,2-dihydropyrimidin-2-one (zebularine)

Three carbocylic analogues of the potent cytidine deaminase inhibitor (CDA) zebularine [1-(beta-D-ribofuranosyl)-1, 2-dihydropyrimidin-2-one, 1a] were synthesized. The selected pseudosugar templates correspond, respectively, to the cyclopentenyl moiety of neplanocin A (compound 4), the cyclopentyl moiety of aristeromycin (compound 5), and a newly designed, rigid bicyclo[3.1. 0]hexane moiety (compound 6). These three carba-nucleoside versions of zebularine were fashioned to overcome the inherent instability of the parent drug. Each target compound was approached differently using either convergent or linear approaches. The immediate precursor to the cyclopentenyl analogue 4 was obtained by a Mitsunobu coupling of pseudosugar 7 with 2-hydroxypyrimidine. The cyclopentyl analogue 5 was linearly constructed from carbocyclic amine 17, and the final target 6 was similarly constructed from the carbobicyclic amine 27. Of the three target compounds, only 5 showed a significant level of inhibition against human CDA, but it was 16 times less potent than zebularine (Ki = 38 microM vs Ki(apparent) = 2.3 microM). Although these carbocyclic analogues appeared to be more stable than zebularine, replacement of the electronegative CO4' oxygen for the less electronegative carbon in 4-6 presumably reduces the capacity of the pyrimidin-2(1H)-one ring to form a covalent hydrate, a step considered crucial for the compound to function as a transition-state inhibitor of the enzyme.     

Rapid reversed-phase high-performance liquid chromatography method for quantitation, at high pH, of the recombinant apolipoprotein A-IMilano in Escherichia coli fermentation broth

In an effort to develop potent agents for reducing the levels of the active estrogen, estradiol, we developed a new category of 17beta-hydroxysteroid dehydrogenase (17beta-HSD) type 1 inhibitors. The compounds described possess a butyl methyl alkylamide side chain linked to the C6 position of estradiol by a thioether. With a series of epimeric mixtures, an optimal side-chain length of five methylene groups (between the amide group and steroid part) was first determined. Thereafter, both C6 epimers of optimized mixture were obtained after high-pressure liquid chromatography separation. 1H and 13C NMR experiments were performed to confirm the stereochemistry of each epimer. The 6beta-orientation of the side-chain was found to be crucial for enzymatic inhibition. Indeed, for the optimized side-chain length, the compound with a beta-orientation (5: N-butyl,N-methyl 7-(3',17'beta-dihydroxy-1',3',5'( 10')-estratriene-6'beta-yl)-7-thiaheptanamide) was 70-fold more potent than the 6alpha-analog. Compound 5 did not inactivate 17beta-HSD type 1, suggesting a reversible inhibitor. In addition, it was found to be a more potent inhibitor than the substrate estrone itself or a panel of three known inhibitors.     

Anthelmintic 1-cinnamamido-2,4-imidazolidinediones

A series of 1-(substituted cinnamamido)-2,4-imidazolidinediones has been prepared from the corresponding cinnamoyl chlorides and 1-amino-2,4-imidazolidinedione hydrochloride in pyridine. These compounds possess a significant degree of anthelmintic activity against the mouse pinworm Syphacia obvelata. The most active compounds are those substituted with halogen or cyano groups.     

Pulmonary arteriole hypertrophy in broilers with pulmonary hypertension syndrome (ascites)

Two experiments were conducted to determine the effect of low ventilation or cool temperature environments on pulmonary arteriole hypertrophy. Male broilers were maintained under control or low ventilation conditions in Experiment 1, whereas male broiler breeder by-product chicks were exposed to cool temperature conditions in Experiment 2. Birds were randomly selected for histological evaluation of lung tissue in both experiments. In Experiment 1, birds that had pulmonary hypertension syndrome (PHS+) exhibited a greater degree of inflammation of lung tissue at 5 and 7 wk of age than controls or birds that did not have PHS (PHS-). These PHS+ birds also had higher numbers of cartilaginous osseous nodules at 3 and 7 wk of age than controls. Morphometric analyses revealed that PHS+ birds in Experiment 1 had a thicker medial layer associated with 100 to 200 microns diameter pulmonary arterioles at 7 wk of age, and 50 to 100 microns arterioles at 3 and 7 wk of age than PHS- or control birds. In Experiment 2, PHS+ birds exhibited a thicker medial layer in pulmonary arterioles at 7 wk of age than did PHS- birds, but there were no differences in medial layer thickness at 5 wk of age nor were there differences in the degree of inflammation or amount of osseous nodule formation between PHS+ and PHS- birds at 5 and 7 wk of age. Thus, pulmonary arteriole hypertrophy was observed in birds having PHS in response to both low ventilation and cool temperature environments and this hypertrophy occurred with or without a coincident inflammatory response in lung tissue.     

Condensed heterocyclic compounds: synthesis and antiinflammatory activity of novel thiazolo[3,2-alpha] pyrimidines

In this study, thirty six new 2-benzylidene-7-methyl-3-oxo-5- phenyl-2,3-dihydro-5H-thiazolo[3,2-alpha]pyrimidine-6-carboxylic acid methyl esters were synthesized and characterized by spectral, crystallographic, and elemental analysis. The antiinflammatory activity of the compounds was tested by the carrageenan hind paw edema test. It was found that compound 6a having a 2-meth-oxyphenyl group at position 5 and a benzylidene group at position 2 was the most potent compound in this series. All the compounds that were tested for ulcer activity gave positive results.     

In vivo observation of subendocardial microvessels of the beating porcine heart using a needle-probe videomicroscope with a CCD camera

We developed a portable needle-probe videomicroscope with a charge-coupled device (CCD) camera to visualize the subendocardial microcirculation. In 12 open-chest anesthetized pigs, the sheathed needle probe with a doughnut-shaped balloon and a microtube for flushing away the intervening blood was introduced into the left ventricle through an incision in the left atrial appendage via the mitral valve. Images of the subendocardial microcirculation of the beating heart magnified by 200 or 400 on a 15-in. monitor were obtained. The phasic diameter change in subendocardial arterioles during cardiac cycle was from 114 +/- 46 microns (mean +/- SD) in end diastole to 84 +/- 26 microns in end systole (p < 0.001, n = 13, ratio of change = 24%) and that in venules from 134 +/- 60 microns to 109 +/- 45 microns (p < 0.001, n = 15, ratio of change = 17%). In contrast, the diameter of subepicardial arterioles was almost unchanged (2% decrease, n = 5, p < 0.01), and the venular diameter increased by 19% (n = 8, p < 0.001) from end diastole to end systole. Partial kinking and/or pinching of vessels was observed in some segments of subendocardial arterioles and venules. The percentage of systolic decrease in the diameter from diastole in the larger (> 100 microns) subendocardial arterioles and venules was greater than smaller (50-100 microns) vessels (both p < 0.05). In conclusion, using a newly developed microscope system, we were able to observe the subendocardial vessels in diastole and systole.(ABSTRACT TRUNCATED AT 250 WORDS)     

Synthesis and calcium channel modulating effects of isopropyl 1,4-dihydro-2,6-dimethyl-3-nitro-4-(thienyl)-5-pyridinecarboxylates

A group of racemic isopropyl 1,4-dihydro-2,6-dimethyl-3-nitro-4-(thienyl)-5-pyridinecarboxylates++ + 7a-f were prepared using a modified Hantzsch reaction that involved the condensation of a thienylcarboxaldehyde 4a-f with isopropyl 3-aminocrotonate 5 and nitroacetone 6. In vitro calcium channel antagonist activities were determined using a guinea pig ileum longitudinal smooth muscle (GPILSM) assay. Compounds 7a-f exhibited weaker calcium channel antagonist activity (IC50 = 10(-5) to 10(-7) M range) than the reference drug nifedipine (IC50 = 1.43 x 10(-8) M). The point of attachment of the C-4 thienyl ring system was a determinant of antagonist activity [3-thienyl (7b) > 2-thienyl (7a)]. A 5-substituent in the 2-thienyl moiety influenced antagonist activity where the potency order was 5-bromo-2-thienyl 7f > or = 5-methyl-2-thienyl 7c > 2-thienyl 7a. Although the 5-methyl-2-thienyl 7c and 3-methyl-2-thienyl 7d isomers are equipotent antagonists, the 5-bromo-2-thienyl compound 7f appears to be marginally more active than the 4-bromo-2-thienyl isomer 7e. The 2-thienyl compound 7a, unlike the 3-thienyl isomer 7b, exhibited an agonist effect on GPILSM in the absence of the muscarinic agonist carbachol. Effects of the 2-thienyl 7a and 3-thienyl 7b isomers on the magnitude of calcium current were determined in guinea pig ventricular myocytes with voltage clamp techniques. Results showed that 2-thienyl 7a inhibited calcium current (antagonist) when voltage steps were made from a potential of -40 mV. However, when voltage steps were made from -60 mV, 7a enhanced calcium current (agonist). The 3-thienyl isomer 7b had little, if any, effect on calcium current.     

Osteochondrodysplasia occurring in transgenic mice expressing interferon-gamma

Arterial thrombi are primarily composed of platelets. Platelets are bound to injured endothelial cells, sub-endothelial matrices, and other platelets by a range of adhesive proteins. Some of these reactions are governed by shear forces. The role of adhesive proteins in the pathogenesis of arterial thrombosis is not fully understood. The aim of this study was to examine the involvement of von Willebrand factor (vWF), fibrinogen (Fg), and fibronectin (FN) in the formation of microvascular thrombi in vivo using a helium-neon laser-induced thrombosis method. Transmission electron microscopy demonstrated that laser irradiation resulted in platelet-rich thrombosis in arterioles and venules, and revealed that this occurred in the absence of endothelial denudation. The mean wall shear rates in mesenteric arterioles and venules were 641 +/- 40 and 280 +/- 20 s-1, respectively. Shear rates increased approximately fivefold in arterioles and tenfold in venules during the formation of occlusive thrombi. Antibody to vWF inhibited thrombosis in arterioles and venules. Antibodies to Fg and FN inhibited thrombosis in venules but not in arterioles. These results confirm that vWF, Fg and FN were involved in thrombogenesis in vivo and demonstrated that significantly higher shear rates were required for the reactions involving vWF than those involving either Fg or FN.