Intake of total omega-3 fatty acids, eicosapentaenoic acid and docosahexaenoic acid and risk of coronary heart disease in the Spanish EPIC cohort study.

BACKGROUND AND AIMS: The evidence about the benefits of omega-3 fatty acid intake on coronary heart disease (CHD) is not consistent. We thus aimed to assess the relation between dietary intake of total omega-3 fatty acids (from plant and marine foods) and marine polyunsaturated fatty acids (PUFAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on the risk of CHD in the Spanish cohort of the European Prospective Investigation into Cancer and Nutrition (EPIC). METHODS AND RESULTS: The analysis included 41,091 men and women aged 20-69 years, recruited from 1992 to 1996 and followed-up until December 2004. Omega-3 fatty acid intake was estimated from a validated dietary questionnaire. Only participants with definite incident CHD event were considered as cases. Cox regression models were used to assess the association between the intake of total omega-3 fatty acids, EPA or DHA and CHD. A total of 609 participants (79% men) had a definite CHD event. Mean intakes of total omega-3 fatty acids, EPA and DHA were very similar in the cases and in the cohort, both in men and women. In the multivariate adjusted model, omega-3 fatty acids, EPA and DHA were not related to incident CHD in either men or women. The hazard ratios (HR) for omega-3 were 1.23 in men (95% CI 0.94-15.9, p = 0.20); and 0.77 in women (95% CI 0.46-1.30, p = 0.76). CONCLUSION: In the Spanish EPIC cohort, with a relatively high intake of fish, no association was found between EPA, DHA and total omega-3 fatty acid intake and risk of CHD.

A comparative study of the sensitivity of the 3-induction and 9-induction Buehler test procedures for assessing skin sensitisation potential.

Assessment of skin sensitization potential is a mandatory requirement for the registration or notification of most types of chemicals and products. Until recently, two methods using the guinea pig as test model were the most widely accepted; the guinea pig maximisation test and the Buehler test. In the case of agrochemical formulations, which constitute the final end use product in contact with operators, industry and also some regulatory authorities consider the Buehler method more appropriate as the methodology is more relevant to likely exposure in the field. However, certain European regulatory authorities have become concerned about the sensitivity of the Buehler test for this purpose and have requested that a modified method is used in which additional applications of test materials are used during the induction phase of the protocol (a total of 9 rather than the normal 3). This study was designed to assess whether this modification was justified. Six reference substances (formaldehyde, alpha-hexylcinnamaldehyde, fragrance mix, thimerosal, mercaptobenzothiazole and phthalic anhydride); all mild to moderate skin sensitizing chemicals, were assessed in a study, which compared the use of 3 and 9 induction applications. The results of this study demonstrated that, although most of these sensitisers were detected by both protocols, the modified method (9 induction applications) was no more sensitive than the standard method (3 induction applications). As the modified protocol is also potentially more stressful to the animals, it is concluded that the use of additional induction applications in the Buehler test cannot be justified from either a scientific or an animal welfare perspective.

Morphine pharmacokinetics and pain assessment after intracerebroventricular administration in patients with terminal cancer.

Morphine pharmacokinetics and pain relief were evaluated after intracerebroventricular administration of morphine (0.4 +/- 0.11 mg) in seven patients with cancer suffering from intractable pain. Ventricular cerebrospinal fluid (CSF), lumbar CSF, and plasma morphine concentrations were analyzed by a specific morphine radioimmunoassay. A two-compartment model was sufficient to describe the kinetics of morphine in ventricular CSF. Morphine diffuses to the lumbar level, and the mean maximum concentration was 192 +/- 105 ng/ml at 4.5 +/- 1.3 hours. Ventricular and lumbar CSF morphine kinetics showed a similar decline during the elimination phase, with terminal half-lives of 3.8 +/- 0.6 hours and 4.2 +/- 1.6 hours, respectively. Pain relief was evaluated by a visual analog scale: the test showed a rapid onset of analgesia (less than 10 minutes). Analgesic effectiveness reached a maximum between 6 and 10 hours. The relationship between pharmacologic effect and morphine concentrations in ventricular CSF resulted in an anticlockwise hysteresis curve. The presence of morphine in lumbar CSF suggested an additive spinal action of morphine, which probably plays a role in the duration of analgesia.

Influence of goat colchicine specific antibodies on murine colchicine disposition.

The potential use of colchicine-specific antibodies (IgG(C)) to overcome colchicine intoxication in mice is of interest in human poisoning. Pharmacokinetics in mice are similar to those in humans. A short distribution half-life (t 1/2 a = 34 min) is associated with a long elimination half-life (t1/2 beta = 48 h) together with a large volume of distribution at steady-state (Vss = 2.5 l/kg) and a low total body clearance (ClT = 1 ml/min/kg). This extensive and rapid distribution to tissues impairs the success of conventional therapies. Despite the administration of a relatively low amount of IgG (C) (15% binding sites vs, colchicine molecules), the beneficial effect of IgG(C) is demonstrated by the alteration in colchicine pharmacokinetics which occurs rapidly following IgG(C) administration as demonstrated by rise in blood toxin concentrations (4-fold relative to IgG(N)-treated controls). This sequestration in the blood is associated with a colchicine redistribution from peripheral to the blood compartment. This extraction effect is revealed by lower colchicine tissue levels in IgG(C)-treated mice than in controls. As a consequence, Vss decreased in the IgG(C) group. Moreover, ClT is diminished in the IgG(C)-treated group because the relatively large immunoglobulin can not be excreted renally. In addition to this toxin displacement, study of free and bound colchicine plasma levels shows a lower percentage of free toxin in the IgG(C)-treated group (33 to 0%) compared to 70% in the control group. This pharmacokinetic study provides evidence that the administration of IgG(C) alters the colchicine disposition by sequestrating and extracting colchicine in blood compartment.

The effect of nortriptyline-specific active immunization on amitriptyline toxicity and disposition in the rabbit.

Rabbits were actively immunized by a conjugate of nortriptyline (NT) to study the effect of specific anti-NT antibodies on toxicity and disposition of amitriptyline (AT). Control and immunized rabbits received 115 mg/kg AT intraperitoneally (i.p.). The lethality dose (LD) profile exhibited a gentle slope; LD100 and LD0 were separated by 100 mg/kg. Mortality was significantly reduced from LD67 to LD43 (P less than 0.05). Total plasma concentrations of the toxin were increased in the immunized group compared to the control group. AUC0.5-24 h value was 5-fold higher in the immunized group than in the control group. Moreover, a smaller fraction of unbound toxin in plasma was observed in the immunized group than in the control group. These observations indicate that AT was actively sequestered by antibodies. The intensity of this phenomenon was a function of both the antibody affinity constant (10(9) M-1) and the neutralizing capacity (varying from 0.005 to 0.2 mg/kg) of the circulating antibodies in each immunized rabbit. Results clearly show that anti-NT antibodies are able to effectively sequestrate AT.

The effect of colchicine-specific active immunization on colchicine toxicity and disposition in the rabbit.

Anti-colchicine antibodies raised in rabbits are effective at protecting rabbits from acute colchicine intoxication. The positive effect depends on the ratio between the binding site capacity of the specific antibodies and the colchicine dose. Immunized rabbits receiving 6 mg/kg colchicine intravenously (LD100) died within 8 h as rapidly as those of the non-immunized control group. In contrast, if the colchicine dose was reduced to 3 mg/kg (LD83), rabbits were protected and mortality decreased to 17%. Study of plasma colchicine pharmacokinetics indicated that colchicine was totally sequestrated by antibodies in the 3 mg/kg group and only 55-80% sequestrated in 6 mg/kg group. This sequestration contributed to reducing colchicine diffusion into tissues (the volume of distribution decreased 7-fold) and to increasing the terminal half-life and the total body clearance of the drug. Moreover, as the slope of the dose-lethality curve was steep, a small binding capacity was sufficient to neutralize colchicine toxicity at 3 mg/kg. Results clearly indicate that anti-colchicine antibodies are able to effectively sequestrate colchicine. Moreover, the amount of circulating antibodies is a crucial limiting factor for the effectiveness of immunotoxicotherapy.

Dose-dependent reversal of acute murine colchicine poisoning by goat colchicine-specific Fab fragments.

The use of colchicine-specific Fab fragments is of interest in human poisoning. In the present study, we show the efficacy of Fab fragments in reversing colchicine toxicity in mice. High affinity antibodies (Ka = 2 x 10(10) M-1) against colchicine were raised in goats; Fab fragments were purified by DEAE chromatography after papain hydrolysis of IgG. Mice were intoxicated with a 100% lethal colchicine dose (3.8 mg/kg). When a half molar dose (M/2) of Fab fragments in relation to the colchicine dose was intravenously and intraperitoneally administered 90 min after colchicine infusion using a multiple dosage schedule, 80% of the Fab-treated mice survived compared to the control group which did not receive Fab fragments (P less than 0.01). Using a M/4 and M/8 dose of Fab fragments, the mortality was respectively 50% and 80%. The dose-effect relationship was linear (r = 0.99). Delayed administration of a M/2 dose of Fab fragments 6 h after colchicine administration resulted in 50% survival (P less than 0.01). Body temperature and body weight were selective markers of the severity of the intoxication. In the control group, a marked decrease of body temperature was observed following the first few hours after the intoxication (-21% compared to basal value 48 h after colchicine). In the Fab-treated group, the decrease was inversely related to the Fab fragment dose. Body temperature returned to the basal values 7 days after intoxication. A progressive decrease in body weight was concomitantly observed in intoxicated mice until death, while values returned to baseline 9 days after colchicine in surviving Fab-treated mice.

Comparison of colchicine toxicity on different dysmyelinating mutant models.

The administration of colchicine to dysmyelinating mutant mice may serve as an in vivo pharmacological tool for the study of the mechanisms involved in the formation of the myelin sheath. The study of the acute toxicity of colchicine in these mutants demonstrated that male animals were much more sensitive than female animals. All of the mutants and their controls were also more resistant to colchicine than the Swiss strain usually used in toxicity studies.

Reversal of murine colchicine toxicity by colchicine-specific Fab fragments.

High-affinity Fab fragments (2 x 10(10) M-1) specific to colchicine were produced to evaluate their potency in reversing murine colchicine intoxication. Intraperitoneal injection of a 4.46 mg/kg colchicine dose was lethal for 100% of mice. 1.5 h after colchicine administration, a group of 10 mice was treated with colchicine-specific Fab fragments at a half-stoichiometrical dose compared to the colchicine dose by intravenous and intraperitoneal routes. 70% of the Fab-infused mice survived (P less than 0.01). This high efficiency of colchicine-specific Fab fragments in reversing acute murine colchicine toxicity suggests that Fab fragments would be an efficient antidote for the treatment of human colchicine poisoning.

Induction of supernumerary ribs with sodium salicylate.

Although many chemical agents induce supernumerary ribs (SNR), few efforts have been published examining the induction of SNR in the presence or absence of maternal toxicity and the effect of dose on SNR length. A single administration of sodium salicylate on Day 9 of pregnancy at different dose levels (120, 180, 240, and 300 mg/kg) was used to induce SNR in the thoracolumbar region. At 180, 240, and 300 mg/kg, body weight loss was observed for dams following the administration, associated with reduced food consumption. The mean litter incidence of SNR in the control groups ranged between 0 and 17.1% and in the treated groups (from 180 to 300 mg/kg) between 50.5 and 88.6%. At 120 mg/kg, no adverse effects were noted in dams and the incidence of SNR was in the range of the control groups. Furthermore, in the three highest dose groups, increased incidences of 27 presacral vertebrae (PSV) were noted. In most instances, fetuses with 27 PSV had extra ribs whereas fetuses with 26 PSV tended to have rudimentary ribs. The distinction of SNR between rudimentary and extra ribs is usually based on a ratio of the length of 14th to 13th rib of 0.50. However, this value does not reflect the separation of the apparent bimodal distribution of SNR induced by sodium salicylate. A ratio of 0.35, which corresponds to the superior limit of the SNR of control fetuses, seemed to better define the two populations of SNR.

Toxicokinetic-toxicodynamic models describing the relation of plasma and red blood cell potassium with plasma digitalis in acute human digitalis poisoning.

Toxicity of cardiac glycosides involves the inhibition of the Na(+)-K(+) ATPase pump. As a consequence, extracellular K(+) concentration rises and intracellular K(+) concentration strongly decreases. Red blood cell (RBC) K(+) is a practical marker of ATPase inhibition. In a group of 15 patients intoxicated by digitoxin and lanatoside C, correlations between the calculated digitoxin ingested dose or plasma digitoxin levels and the kinetics of plasma K(+) and RBC K(+) have been assessed using kinetic-effect modelling. A correlation between the calculated ingested dose of digitoxin with RBC K(+) was found (r = 0.64). A direct relation based on the linear model fitted the relation between extracellular K(+) and digitalis concentration. An indirect relation based on the Emax sigmoid model fitted the relation between RBC K(+) and digitoxin concentrations. Specific parameters were obtained from the linear model with a = 0.0196 +/- 0.0272 and b = 0.455 +/- 0.035. Specific parameters were derived from the Emax sigmoid model with k(eo) = 0.0139 +/- 0.0052/hr and EC(50) = 91.95 +/- 20.55 ng/ml, where k(eo) = first-order rate constant of the disappearance of the toxic effect and EC(50) = digitoxin concentration decreasing the RBC K(+) concentration by 50%. These data showed that the in vitro assays of plasma K(+) and RBC K(+) are convenient and predictive assays for evaluating the severity of human digitoxin poisoning.

Fab-bound colchicine appears to adopt Fab fragment disposition in rats.

The disposition of colchicine-specific Fab fragments and the effect of Fab fragment administration on the disposition of colchicine were studied in anaesthetized bile duct-cannulated rats. One group of rats (n = 6) received a 125I-Fab dose of 38 mg kg-1 i.v. The plasma disposition was characterized by a volume of distribution of 179 +/- 48 mL kg-1, total body clearance of 1.02 +/- 0.07 mL min-1 kg-1, t1/2 alpha of 0.17 +/- 0.03 h and t1/2 beta of 1.3 +/- 0.3 h. Fab fragments were in part excreted by the renal route (15.6 +/- 6% of the Fab dose), while biliary excretion was a minor route (< 2% of the Fab dose). Two other groups of rats received 15 micrograms kg-1 colchicine (n = 6) or 15 micrograms kg-1 colchicine plus 38 mg kg-1 colchicine-specific Fab fragments (n = 6) by intravenous infusion. Pharmacokinetics of colchicine was markedly altered in the Fab-colchicine-treated rats. In this group, distribution volume and total body clearance of colchicine were decreased by factors of 22 and 10, respectively, compared with the values in the colchicine-treated group and were very similar to those of Fab fragments. An 80% reduction of cumulative biliary excretion of colchicine was observed in Fab-colchicine-treated rats (P < 0.01). The fraction of colchicine dose excreted by the urinary route was 38 +/- 6.9 and 9 +/- 0.7% respectively in Fab-colchicine- and colchicine-treated groups (P < 0.01). These data show that during Fab treatment, colchicine followed the elimination kinetics of Fab fragments.(ABSTRACT TRUNCATED AT 250 WORDS)

Specific anti-digoxin Fab fragments: an available antidote for proscillaridin and scilliroside poisoning?

The purpose of this study was to evaluate the capacity of specific anti-digoxin Fab fragments to bind to and neutralize scilliroside and proscillaridin in acute poisoning. Apparent affinity constants were determined with values of 2.6 10(8)M-1 for scilliroside and 3.8 10(7)M-1 for proscillaridin. These results are in accordance with a possible in-vivo neutralization of these toxins.

Reversal of advanced colchicine toxicity in mice with goat colchicine-specific antibodies.

Colchicine-specific antibody (IgG(C] was tested in mice for reversal of colchicine toxicity. The mouse model was chosen because it reflects human pathophysiology in colchicine poisoning. IgG(C) was administered when at least 85% of colchicine was distributed in tissues. It resulted in a dramatic decrease in lethality from 85% (control group) to 10% (treated group). The decrease in toxic effects was confirmed by evaluating physiological parameters. The recovery of thermoregulation was very rapid in mice treated with IgG(C), while recovery in body weight was less marked. IgG(C) administration, therefore, decreases the intensity but may extend the duration of colchicine toxicity (reversible binding). The total neutralizing binding capacity of IgG(C) used was such that administered IgG(C) neutralizing binding sites were either 7 or 15% of the injected colchicine dose. In spite of this low neutralizing capacity the treatment was successful because of the ability of IgG(C) to buffer the amount of colchicine molecules on the critical slope of the dose-lethality curve.

Relationship between red blood cell potassium and plasma digitoxin concentrations in intoxicated patients.

After severe acute self-poisoning by cardiac glycosides, significant and persistent depletion of red blood cell K+ due to inhibition of Na+K+ ATPase is seen. Because of a delay between the time course of plasma digitalis concentrations and that of red blood cell K+ depletion, no direct relation exists between the two, and RBC K+ has hitherto not been considered useful as prognostic indicators of clinical outcome. In an effort to solve this problem, red blood cell K+ was measured by atomic absorption spectrophotometry and plasma digitoxin concentration assayed in six patients admitted to an intensive care unit after digitoxin self-poisoning. Using the effect compartment model of Sheiner, a relationship based on a sigmoid Emax model was able to relate the digitoxin concentration at the action site to red blood cell K+ depletion. Thus the duration of red blood cell K+ depletion could be predicted from two relative simple in vitro assays. Since RBC K+ is a marker of the inhibition of Na+K+ ATPase by digitoxin, this method could be of use for the management of patients self-poisoned with digitalis.

Postnatal evolution of supernumerary ribs in rats after a single administration of sodium salicylate.

Radiographs were used to follow the postnatal evolution of 14th ribs in rat pups. Initially, 30 pregnant female rats were randomly distributed into two groups receiving 0 or 300 mg kg(-1) sodium salicylate on day 9 of pregnancy. In the treated group, adverse effects were noted on body weight changes and food consumption during the 2 days following dosing. At birth, a high majority of pups had extra ribs at the 300 mg kg(-1) dose. Radiographs done on postnatal days 1, 6, 14, 28 and 54 showed a reduction in the incidence of rudimentary ribs only, whereas extra ribs, often associated with 27 presacral vertebrae, had the same incidence from birth to adult stage. Furthermore, extra ribs seemed to exhibit similar growth evolution to the other thoracic ribs. This work helps to clarify the postnatal evolution of supernumerary ribs because it was performed on the same animals from birth to adult stage, showing that the reversibility was related to rib length and, in consequence, concerned the rudimentary ribs only. The coexistence of additional presacral vertebrae primarily with extra ribs suggests that both kinds of supernumerary ribs (rudimentary and extra) might be different phenomena and could be considered separately in developmental toxicology studies.

Pharmacokinetics of Vipera aspis venom after experimental envenomation in rabbits.

Toxicokinetic studies of Vipera aspis venom were performed in rabbits after experimental envenomation. Venom proteins with a molecular weight greater than 6 kDa (high-molecular weight proteins) and which reacted in enzyme-linked immunosorbent assay with specific antiviper venom Fab'2, were also responsible for the lethal potency and the capillary permeability increasing activity of the venom. Conversely, low-molecular weight proteins were not detected by enzyme-linked immunosorbent assay and were pharmacologically inactive. The toxicokinetics of both classes of venom components were studied, using high-molecular weight and low-molecular weight radiolabeled proteins as well as enzyme-linked immunosorbent assay. After intravenous injection, Vipera aspis venom in plasma followed a biexponential decline with a distribution half-life of 0.7 hr and an elimination half-life of 12 hr. The distribution volume was 1.2 l.kg-1 and the systemic clearance was 84 ml.hr-1.kg-1. Venom levels in plasma after intramuscular injection of three doses (300, 500 and 700 micrograms/kg) of venom increased within the few hours after the venom administration to reach maximal values proportional to the injected doses. They subsequently followed a monoexponential decline, with an apparent terminal half-life of 32.5 hr. Absorption was a kinetically complex process, rapid during the first 24 hr and continued at a slower rate over the subsequent 72 hr. Bioavailability of venom was about 65%, regardless of the administered dose, and less than 5% of venom injected was excreted by the renal route.

[Immunotoxicotherapy: protective effect of anti-colchicine antibodies in acute colchicine poisoning in rabbits and mice]. / Immunotoxicothérapie: effet protecteur d'anticorps anticolchicine lors de l'intoxication aiguë à la colchicine chez le lapin et la souris.

Anti-colchicine antibodies are able to neutralize toxic effects of colchicine after acute intoxication in rabbits and mice. The protecting effect is demonstrated by active immunization (rabbits) or passive immunization (mice). These data suggest that the immunotoxicotherapy may be useful for compounds (colchicine) with intracellular action.

Pharmacokinetics of colchicine: a review of experimental and clinical data.

Thanks to the development of a sensitive and specific radioimmunoassay for colchicine, the pharmacokinetics of colchicine is now well-established after single oral doses. Absorption is characterized by a zero-order rate constant while disposition appears biexponential with a rapid distribution phase (t1/2 = 1.8 h) and a long elimination phase (t1/2 = 20 h). All studies confirm the large total body clearance (35 l/h) predominantly by the extrarenal route and the large distribution volume (700 l). Further studies need to be performed to investigate colchicine absorption and to describe the metabolic pathway of the drug. To date, relationships between colchicine plasma levels and pharmacological effects have not been defined. Monitoring of plasma levels in patients with familial Mediterranean fever should improve treatment with colchicine. However, the therapeutic range has not been precisely determined. The use of colchicine in the treatment of liver cirrhosis and primary biliary cirrhosis is a recent development; so, assuming that a large part of total body clearance depends on hepatic function, the influence of hepatic diseases on colchicine disposition needs to be investigated in order to define the most appropriate therapeutic dosing.