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J Millet

Publications and source records attributed to J Millet.

At least 37 records · Page 2Linked to original sources
PubMed1996

Experimental venous thrombosis induced by homologous serum in the rat.

To investigate the effects of new drugs on the prevention of venous thrombosis, the Wessler stasis model is extensively employed usually in the rabbit. In this model, a hypercoagulable state is achieved by the administration of either heterologous serum (often human serum) or an activated coagulation factor followed by total stasis of the vein in which the thrombus is to be formed. Although classically the stasis model is performed in rabbits, it is possible to induce experimental venous thrombosis in a number of animal species, including rats. However, the use of homologous serum as the activating or thrombogenic agent has been previously described only in the dog and in the rabbit. The purpose of the present study was to compare the activities of heterologous and homologous serum as hypercoagulating agents in a rat stasis model of venous thrombosis using Wessler's technique. Heparin was employed as a reference antithrombotic compound in order to compare the two thrombogenic challenges.

Animals↗
PubMed1996

Lipophilicity determination of psoralens used in therapy through solubility and partitioning: comparison of theoretical and experimental approaches.

The aim of this study was to determine and to compare experimental and theoretical solubilities (S) as well as partition coefficients (PC) in an octanol/water system of psoralen (P), 8-methoxypsoralen (8-MOP), 5-methoxypsoralen (5-MOP) and 4,5',8-trimethylpsoralen (TMP). For each psoralen, experimental results were performed in triplicate with a spectrofluorimetric technique. The measurements were achieved 10 times for each solution. The obtained order of the solubilities in pure octanol was 5-MOP approximately TMP > P > 8-MOP, while in water-saturated octanol it was expressed as follows: TMP approximately 5-MOP > P > 8-MOP. However, the following order was found for hydrophobicity: TMP > 5-MOP > 8-MOP > P. The solubility ratios (SR) in pure octanol and water were assessed (mean +/- SD): 3.13 +/- 0.01 (P), 2.60 +/- 0.01 (8-MOP), 3.75 +/- 0.01 (5-MOP), and 5.11 +/- 0.01 (TMP). In saturated phases, they were 3.27 +/- 0.01, 2.63 +/- 0.01, 3.85 +/- 0.01, and 5.32 +/- 0.01, respectively. The PCs were determined with low concentrations according to the Dearden and Bresnen32 method and they were 1.67 +/- 0.01, 1.93 +/- 0.01, 2.00 +/- 0.01, and 3.14 +/- 0.01, respectively. Solubility parameters (delta), in Hildebrand unit (H) or in (cal/cm3)1/2, were evaluated. They confirmed the polarity of psoralens, previously expressed through the PC, although the positional isomers (5-MOP and 8-MOP) revealed no difference. Hildebrand's approach to the solubility of regular solutions and Yalkowsky's concept of the solubility of nonelectrolytes and weak electrolytes in an octanol/water system permitted a comparison of the theoretical and experimental results. The perspective of this work is to use the physicochemical properties of the psoralens in practice for insuring convenient experimental assays and the prediction, in vitro, of the percutaneous absorption of these compounds.

5-Methoxypsoralen↗
PubMed1996

Pharmacologic and biochemical profiles of new venous antithrombotic beta-D-xyloside derivatives: potential antiathero/thrombotic drugs.

Following a screening program for orally active antithrombotic drugs, it was found that a series of thioxyloside compounds presented with good venous antithrombotic properties. Of more than 500 compounds, LF 09-0055, LF 04-0212, and LF 05-0030 were the most active at inhibiting venous thrombus formation in the rat and rabbit Wessler model after intravenous and oral dosing. LF 05-0030 showed the greatest activity with an ED80 value of 6 mg/kg on oral administration in rats. This activity was maintained in several different models of venous thrombosis and shown to be devoid of anticoagulant effects or hemorrhage. Kinetic studies have demonstrated that maximal levels of activity, following either intravenous or oral dosing, occurred between 2 and 4 hours after administration. This may reflect the type of mechanism involved, since it has been well documented in the literature that xylosides are capable of initiating glycosaminoglycan (GAG) synthesis. Moreover, in vitro galactosyltransferase 1 (the second enzyme involved in GAG polymerization) enzymic assays showed that these thioxyloside derivatives were good acceptors for galactose transfer and therefore at initiating GAG formation. Further in vivo experimentation demonstrated that after treatment by these molecules an important elevation in circulating GAG occurred, with LF 05-0030 presenting the greatest activity, being five times higher than control levels. In addition it was found that dermatan sulfate levels, expressed as antithrombin activity by heparin cofactor II, were significantly increased over control values. As such, this dermatan sulfate moiety is believed to support the antithrombotic activity observed. Studies are underway to investigate the activity of these interesting molecules in atherosclerosis and other vessel wall diseases.

Administration, Oral↗
PubMed1996

The arterial antithrombotic activity of thioxylosides in a rat model of laser-induced thrombosis.

beta-D xylosides have been shown to have venous antithrombotic properties after simple oral administration. Therefore, the arterial antithrombotic effect of these compounds was investigated in vivo, using the experimental thrombosis model induced by laser injury. The products tested were administered orally, 4 h before the thrombosis induction. Two beta-D xylosides were tested (LF 09-0055 and LP 05-0030), either after a simple oral administration at 50, 100, 200, and 400 mg/kg, or after repetitive oral administration at 200 mg/kg twice daily during 5 days. These compounds increased significantly the number of laser shots required to induce arterial thrombosis and decreased the number of emboli and the duration of embolization. At single-dose or repeated administrations, these xylosides did not affect diluted thrombin time in platelet-poor plasma collected after thrombosis inductions. They induced a dermatan sulfate-like activity in the plasma of treated rats, as measured by heparin cofactor II-mediated thrombin inhibition assay. These data suggest that these xylosides are potent arterial antithrombotic agents after single or repetitive oral administrations. beta-D xylosides constitute a very promising therapeutic class of orally active antithrombotic drugs.

Administration, Oral↗
PubMed1996

In vitro study of percutaneous absorption, cutaneous bioavailability and bioequivalence of zinc and copper from five topical formulations.

Percutaneous absorption and cutaneous bioavailability of zinc and copper from zinc 2-pyrrolidone 5-carboxylate (ZnPC), zinc oxide (ZnO), zinc sulfate (ZnSO4), copper 2-pyrrolidone 5-carboxylate (CuPC) and copper sulfate (CuSO4) were compared using 5 formulations (3 emulsions and 2 ointments) that were applied topically on human skin in vitro. After application for 72 h, percutaneous absorption of zinc from ointments containing ZnO and ZnSO4 was found to be lower than that from a ZnPC-containing emulsion (0.36 and 0.34 versus 1.60% of applied dose). In the case of copper, the flux after a 72-hour treatment period showed that there had been minimal release from CuPC- and CuSO4-containing formulations (approximately 5 ng/cm2/h). All formulations used in this study effected an increase in zinc and copper concentrations in whole skin and epidermis. Bioequivalence of the 5 formulations based on pharmacokinetic results was assessed, and salt and vehicle effects were discussed.

Administration, Topical↗
PubMed1996

The antithrombotic potential of dalteparin in man assessed indirectly by Wessler's technique.

The aim of this study was to demonstrate the effectiveness of a new experimental approach for assessing the antithrombotic potential of low molecular weight heparins (LMWH) such as dalteparin. In this approach, sera obtained from healthy volunteers treated with various i.v. doses of dalteparin (30, 60 or 120 anti-Xa IU/kg) or placebo (physiological saline) were used as the thrombogenic challenge in a Wessler's stasis model of experimental venous thrombosis in the rat. Sera from placebo-treated volunteers showed a stable thrombogenic activity (0.25 ml/kg of serum producing thrombi of about 50 mg wet weight). Sera from healthy volunteers having previously received dalteparin however demonstrated dose- and time-related reductions in their thrombogenic activity. Half-lives of these effects were 300. 444 and > 480 min for 30, 60 and 120 anti-Xa IU/kg dalteparin respectively. These values were significantly higher than the corresponding anti-Xa and anti-IIa half-lives.

Animals↗
PubMed1995

The effect of the beta-D-xyloside naroparcil on circulating plasma glycosaminoglycans. An explanation for its known antithrombotic activity in the rabbit.

Beta-D-Xylosides are known to initiate or prime free glycosaminoglycan (GAG) chain synthesis in cell and tissue culture. As such, the effect of the venous antithrombotic beta-D-xyloside, naroparcil, was investigated on the plasma GAG profile in the rabbit after oral administration. Using dose-response experiments, we showed that antithrombin activity via antithrombin III and heparin cofactor II was increased in parallel with GAG plasma levels compared to control. A more detailed qualitative examination of plasma GAGs by cellulose acetate electrophoresis and ion-exchange chromatography, following oral administration of naroparcil at 400 mg/kg, revealed the presence of higher density charged molecules compared to control. The extracted GAGs were found to activate inhibition of thrombin by heparin cofactor II and contained approximately 25% of a dermatan sulfate-like compound (undetectable in control), which could be responsible for the antithrombotic effect. Using radiolabeled naroparcil, we found radiolabeled GAG fractions and the fact that naroparcil was a substrate for galactosyltransferase I, the second enzyme responsible for GAG chain polymerization, suggested that the compound could initiate in vivo the biosynthesis of antithrombotic free GAG chains. This is, to our knowledge, the first description of the in vivo effect of a beta-D-xyloside on GAG biosynthesis; furthermore, this is correlated with an antithrombotic action.

Animals↗
PubMed1995

A mutant of Azospirillum brasilense Sp7 impaired in flocculation with a modified colonization pattern and superior nitrogen fixation in association with wheat.

We report here significant phenotypic and genetic differences between Azospirillum brasilense Sp7 and spontaneous mutant Sp7-S and their related properties in association with wheat. In contrast to the wild-type strain of Sp7, colonies of Sp7-S stained weakly with Congo red when grown on agar media containing the dye and did not flocculate in the presence of fructose and nitrate. Scanning and transmission electron micrographs showed clearly that the Sp7-S strain lacked surface materials present as a thick layer on the surface of the wild-type Sp7 strain. Different patterns of colonization on wheat roots between Sp7 and Sp7-S, revealed by in situ studies using nifA-lacZ as a reporter gene, were related to a large increase in nitrogenase activity (acetylene reduction) with Sp7-S in association with normal and 2,4-dichlorophenoxyacetic acid-treated wheat for assays conducted under conditions in which the nitrogenase activity of free-living Azospirillum organisms was inhibited by an excess of oxygen. Randomly amplified polymorphic DNA analysis indicated the close genetic relationship of Sp7-S to several other sources of Sp7, by comparison to other recognized strains of A. brasilense. Genetic complementation of Sp7-S was achieved with a 9.4-kb fragment of DNA cloned from wild-type Sp7, restoring Congo red staining and flocculation.

2,4-Dichlorophenoxyacetic Acid↗
PubMed1995

Venous antithrombotic and anticoagulant activities of a fucoïdan fraction.

Fucoïdans catalyse thrombin inhibition by antithrombin (AT) and heparin cofactor II (HCII); their affinity for each serpin varies according to the seaweed species from which they are extracted, as well as their chemical composition and molecular weight. We extracted a homogeneous fucoïdan fraction from Ascophyllum nodosum, a brown seaweed, and tested its anticoagulant and antithrombotic activities. At a fucoïdan concentration of 3.75 micrograms/ml, thrombin inhibition mediated by AT showed an apparent second-order rate constant (kapp) of 2 x 10(8) M-1 min-1, compared to 1.5 x 10(6) M-1 min-1 for the uncatalyzed reaction. The kapp value of thrombin inhibition via HCII was 1.17 x 10(9) M-1 min-1 at a fucoïdan concentration of 50 micrograms/ml, compared to 1.72 x 10(5) M-1 min-1 for the uncatalyzed reaction. In a Wessler model of venous thrombosis, the fucoïdan fraction, injected intravenously to rabbits 10 min before thrombosis induction, exhibited antithrombotic activity: 1.8 mg/kg was the dose which inhibited F Xa-induced thrombus formation by 80% (ED80), compared to a heparin ED80 of 0.1 mg/kg. At this ED80 the antithrombotic effect of the fucoïdan persisted longer than that of heparin (30 min versus 15 min). The thrombin clotting time (TCT) was significantly prolonged (73 s versus control 29 s, compared to 53 s with heparin) 10 min after a fucoïdan bolus infusion giving a plasma fucoïdan concentration of 14.6 +/- 2.7 micrograms/ml. The bleeding time was slightly increased after fucoïdan infusion at the ED80. Fucoïdan extracted from marine flora thus shows promise as an antithrombotic drug.

Animals↗
PubMed1994

The venous antithrombotic profile of naroparcil in the rabbit.

The venous antithrombotic profile of naroparcil or (4-[4-cyanobenzoyl]-phenyl)-1.5-dithio-beta-D-xylopyranoside was investigated in the rabbit following single i.v. and oral administration. Naroparcil attenuated thrombus development in a Wessler stasis model of venous thrombosis (jugular vein) employing bovine factor Xa as a thrombogenic stimulus giving ED50 values of 21.9 mg/kg and 36.0 mg/kg after respectively i.v. and oral administration. Venous antithrombotic activity was maximal 2-3 h after i.v. administration and 4-8 h after oral administration. Four hours after the oral administration of maximal antithrombotic (Wessler model, factor Xa) doses (100 and 400 mg/kg), naroparcil had no significant effect on bleeding time. In platelet poor plasma obtained from animals treated 4 h previously with various doses (25 to 400 mg/kg) of naroparcil, there was no detectable anti-factor Xa nor antithrombin activity. Similarly, naroparcil had no effect on APTT nor on thrombin time. A sensitized thrombin time (to about 35 s) was modestly but significantly increased following oral administration of the compound at 400 mg/kg. However, thrombin generation by the intrinsic pathway was reduced in a dose-related manner, maximal reduction being 65% at 400 mg/kg. The same dose of naroparcil enhanced the formation of thrombin/heparin cofactor II complexes at the expense of thrombin/antithrombin III complexes in plasma incubated with (125I)-human alpha-thrombin and induced the appearance of dermatan sulfate-like material in the plasma of treated rabbits, as measured by a heparin cofactor II-mediated thrombin inhibition assay.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Oral↗
PubMed1993

Glycosylated derivatives of benzophenone, benzhydrol, and benzhydril as potential venous antithrombotic agents.

A series of glycosylated derivatives of benzophenone, benzhydrol, and benzhydril has been synthesized and evaluated for potential activity as venous antithrombotic agents. Studies on structure-activity relationships revealed that compounds having an electron-withdrawing group in the benzhydril or benzhydrol moiety, and specifically those having the beta-D-xylopyranosyl structure in the sugar moiety, were good antithrombotic agents in a rat model of venous thrombosis.

Administration, Oral↗
PubMed1992

Cellulose degradation by Clostridium thermocellum: from manure to molecular biology.

Clostridium thermocellum, a Gram-positive, thermophilic anaerobe produces a highly active cellulase system. This system, termed the cellulosome, is a complex composed of at least 14-18 different types of components organized around a large, cellulose-binding protein. Combining recombinant DNA technology and protein biochemistry has proved to be a successful approach in unravelling some important features of the system.

Amino Acid Sequence↗
PubMed1992

The venous antithrombotic effect of LF 1351 in the rat following oral administration.

The venous antithrombotic effects of a novel chemical entity, LF 1351, were investigated in rats following single oral administration, in comparison with i.v. administered heparin. LF 1351 demonstrated a dose-related antithrombotic effect in three models of venous thrombosis. The compound was approximately equipotent in two models involving complete stasis of the vena cava and administration of factor Xa or porcine serum, giving respective ED50 values of 48.7 mg/kg and 36.7 mg/kg. LF 1351 was less effective in a model involving partial stasis in the presence of an endothelial lesion. In this case, the antithrombotic effect did not exceed 60-65%, the ED50 being 150 mg/kg. Heparin (50-300 micrograms/kg; 7.5-45.0 U/kg) was effective in all three models. At the approximate ED80 value against factor Xa-induced thrombosis (75 mg/kg) the antithrombotic effect of LF 1351 persisted for 6 h. The antithrombotic effect of LF 1351 (300 mg/kg) occurred without significant changes in APTT or thrombin time.

Administration, Oral↗
PubMed1991

Identification of a histidyl residue in the active center of endoglucanase D from Clostridium thermocellum.

Diethylpyrocarbonate modification of endoglucanase D from Clostridium thermocellum, cloned in Escherichia coli, resulted in a rapid but partial (maximally 70-80%) loss of activity. The second-order rate constant of inactivation proved to be exceptionally high (3210 M-1.min-1). A 3-fold reduction of the kcat and a 2-fold increase of the Km for 2'-chloro-4'-nitrophenyl beta-cellobioside were observed. Spectrophotometric analysis indicate the presence of one rapidly (k = 0.45 min-1) and two slower (k = 0.23 min-1) reacting histidyl residues. In the presence of 50 mM methyl beta-cellotrioside, the rate of inactivation was reduced 16-fold, and the kinetics of modification were compatible with the protection of 1 histidyl residue. Since peptide analysis was inconclusive, identification of the critical residue was attempted by site-directed mutagenesis. Each of the 12 histidyl residues present in the endoglucanase D sequence was mutated into either Ala or Ser. Seven of the mutant enzymes had specific activities lower than 50% of the wild-type. Only in the case of the Ser-516 mutant, however, was the residual activity not affected by diethyl pyrocarbonate. These findings suggest an important functional or structural role for His-516 in the wild-type enzyme.

Amino Acid Sequence↗
PubMed1991

High activity of inclusion bodies formed in Escherichia coli overproducing Clostridium thermocellum endoglucanase D.

The formation of cytoplasmic inclusion bodies by Escherichia coli overproducing Clostridium thermocellum endoglucanase D (EGD) was investigated. EGD was found in inclusion bodies as a 68 kDa form, whereas the size of the cytoplasmic form was 65 kDa. Upon solubilization with urea followed by dialysis, the 68 kDa form was converted to the 65 kDa species. Proteolysis occurred within the COOH-terminal, reiterated region of the 68 kDa form, which is conserved among most C. thermocellum endoglucanases, but is not required for catalytic activity. The specific activity of the enzyme embedded in inclusion bodies was close to that of the purified protein. Thus, inclusion body formation does not involve denaturation of the catalytic domain of EGD, but, more likely, the participation of the reiterated, conserved region in intermolecular interactions.

Blotting, Western↗
PubMed1991

Peak blistering point: influence on fluid levels of 5-MOP in human skin in vivo after systemic administration.

The concentration of 5-methoxypsoralen (5-MOP) in suction blister fluid (SBF) after oral intake was determined in relation to the peak blistering point. Interstitial fluid was obtained from nine healthy male volunteers by applying mild suction (300-350 mmHg) to the skin of the volar aspect of the forearm. Blisters were raised at three different times: 18 h prior to drug administration (group I); 2 h prior to drug administration (group II); and during drug ingestion (group III). SBF levels of 5-MOP were determined in each group of blisters 2 h after oral administration of 1.2 mg/kg micronized 5-MOP. The results showed a statistically significant difference in the concentrations of 5-MOP between each group of blisters. The highest concentration of 5-MOP in the SBF was found in group III and the lowest in group II blisters. These findings suggest that the point of peak blistering in relation to drug administration needs to be known in each pharmacological study using suction blisters.

5-Methoxypsoralen↗