The role of oxidative stress in the ochratoxin A-mediated toxicity in proximal tubular cells.

Balkan endemic nephropathy (BEN), a disease characterized by progressive renal fibrosis in human patients, has been associated with exposure to ochratoxin A (OTA). This mycotoxin is a frequent contaminant of human and animal food products, and is toxic to all animal species tested. OTA predominantly affects the kidney and is known to accumulate in the proximal tubule (PT). The induction of oxidative stress is implicated in the toxicity of this mycotoxin. In the present study, primary rat PT cells and LLC-PK(1) cells, which express characteristics of the PT, were used to investigate the OTA-mediated oxidative stress response. OTA exposure of these cells resulted in a concentration-dependent elevation of reactive oxygen species (ROS) levels, depletion of cellular glutathione (GSH) levels and an increase in the formation of 8-oxoguanine. The OTA-induced ROS response was significantly reduced following treatment with alpha-tocopherol (TOCO). However, this chain-braking anti-oxidant did not reduce the cytotoxicity of OTA and was unable to prevent the depletion of total GSH levels in OTA-exposed cells. In contrast, pre-incubation of the cell with N-acetyl-L-cysteine (NAC) completely prevented the OTA-induced increase in ROS levels as well as the formation of 8-oxoguanine and completely protected against the cytotoxicity of OTA. In addition, NAC treatment also limited the GSH depletion in OTA-exposed PT- and LLC-PK(1) cells. From these data, we conclude that oxidative stress contributes to the tubular toxicity of OTA. Subsequently, cellular GSH levels play a pivotal role in limiting the short-term toxicity of this mycotoxin in renal tubular cells.

The regulation of oxidative drug metabolism by growth hormone in the dwarf goat: differences from and similarities to the mechanisms in rats.

The effects of bovine GH (BST), administered in different dose patterns, on in-vivo oxidative drug metabolism, were studied in female dwarf goats. Animals received recombinantly derived methionyl BST at a dose of 500 micrograms/kg body weight per 24 h for 6 days. It was administered to one group of goats as one s.c. injection per day, another group received a similar 24-h dose divided into three s.c. injections given at 8-h intervals, and the third group received 50 micrograms BST/kg body weight every 2.5 h by a pulsative i.v. infusion. Oxidative metabolic capacity was assessed by determining plasma sulphadimidine (SDD) elimination and urinary metabolite excretion. SDD shows a marked sex-dependent plasma elimination in dwarf goats, with male goats having a lower plasma clearance than female goats. When BST was given by daily injection, no clear effects on SDD plasma clearance or urinary metabolite excretion were observed. However, when the total dose was divided into three injections given at 8-h intervals, the plasma SDD elimination rate decreased. This was associated with a decrease in urinary excretion of the two main hydroxy SDD metabolites. When BST was given by discontinuous i.v. infusion, simulating the male endogenous plasma GH pattern, a marked decrease in SDD plasma clearance was observed. In addition, the excretion of the two urinary hydroxy metabolites was considerably reduced. These results suggest that GH can affect drug oxidation in dwarf goats via mechanisms similar to those suggested for rats. However, in the dwarf goat, the sex differences in drug metabolism are opposite to those in rats.

Tiamulin inhibits human CYP3A4 activity in an NIH/3T3 cell line stably expressing CYP3A4 cDNA.

Tiamulin is an antibiotic frequently used in veterinary medicine. The drug has been shown to produce clinically important interactions with other compounds that are administered simultaneously. An NIH/3T3 cell line, stably expressing human cytochrome P450 (EC 1.14.14.1) cDNA (CYP3A4), was used to study the effect of tiamulin on CYP3A4 activity. The 6 beta-hydroxylation activity of testosterone, which is increased in CYP3A4-expressing cells compared to vector-transfected cells, showed reduced activity after incubation with 1 microM tiamulin and was completely reduced to background level after incubation with 2, 5 and 10 microM tiamulin. The CYP3A4-expressing cell line was used in combination with a shuttle vector containing the bacterial lacZ' gene to study the effect of tiamulin on CYP3A4-mediated mutagenicity of aflatoxin B1. The mutation frequency of aflatoxin B1 could be completely inhibited by tiamulin in CYP3A4-expressing cells, but no effect was observed on the mutation frequency of the direct mutagen ethylmethanesulphonate. Western blotting of homogenates of the CYP3A4-expressing cell line showed stabilization of CYP3A4 protein after incubation with tiamulin, supporting the hypothesis that the mechanism of inhibition is by binding of tiamulin to the cytochrome.

Differential effects of pentoxifylline on the hepatic inflammatory response in porcine liver cell cultures. Increase in inducible nitric oxide synthase expression.

Pentoxifylline (PTX) has been shown to exert hepatoprotective effects in various liver injury models. However, little information is available about the effect of PTX on the hepatic acute phase response. In the present study, the effect of PTX on a lipopolysaccharide (LPS)-induced acute phase response in primary porcine liver cell cultures was examined. During 72 hr of incubation with or without LPS, the ability of PTX to influence the secretion of tumour necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), acute phase proteins, and nitric oxide (NO) was assessed. PTX completely inhibited LPS-induced TNF-alpha production and attenuated IL-6 only after 48 hr of incubation. In contrast, PTX potentiated NO production and the expression of inducible nitric oxide synthase (iNOS) in hepatocytes after stimulation with LPS. The increased expression of iNOS and concurrent production of NO was also observed when liver cell cultures were incubated with dibutyryl cyclic adenosine monophosphate. No effect of PTX on acute phase protein secretion was observed during 72 hr of incubation. The present results show that PTX differentially affects the endotoxin-induced inflammatory response in primary porcine liver cell cultures by suppressing TNF-alpha and IL-6 while potentiating NO production.

Characterization of cytochrome P450 isoenzymes in primary cultures of pig hepatocytes.

Despite the fact that pigs are increasingly used in pharmacological and toxicological studies, knowledge on the enzymes which metabolize xenobiotics, in particular cytochrome P450 (CYP) enzymes, in pigs is still very limited. Primary cultures of pig hepatocytes were used to characterize CYP enzymes. The characterization was performed at the level of enzymatic activities, apoprotein and mRNA analyses. Enzyme inducers investigated were beta-naphthoflavone (BNF), phenobarbital (PB), dexamethasone (DEX) and rifampicin (RIF). After 48hr of BNF treatment, CYP1A protein and mRNA levels were increased, and ethoxyresorufin O-deethylation and caffeine 3-demethylation were strongly induced. PB and RIF increased the levels of CYP3A apoprotein and mRNA, whereas BNF down-regulated CYP3A and related activities. PB and RIF treatment resulted in increased ethylmorphine N-demethylation and testosterone hydroxylation, which appears to be the result of CYP3A induction. Hybridization of pig RNA with a human CYP2C9 cDNA probe showed a PB and RIF inducible CYP, which was down-regulated by BNF. Similar inducing effects were observed for tolbutamide, a marker substrate for CYP2C. DEX was not a potent inducer, although some induction of CYP3A mRNA was observed. The present results indicate the absence of CYP2B and probably CYP2D enzymes and activities in pig liver. Despite some dissimilarities, the results indicate that pigs, apart from their very human-like physiology, might represent a more appropriate model species for oxidative drug metabolism in humans than rats.

Selective changes in oxidative xenobiotic metabolism in vivo and in vitro after parenteral administration of recombinant bovine somatotrophin to rats.

Effects of recombinant bovine somatotrophin (rBST) on in vivo and in vitro oxidative drug metabolism were studied in male rats. rBST was given subcutaneously at a dose of 250 or 500 micrograms 100 g-1 bodyweight 24 h-1 in different dosage patterns. Sulphadimidine (SDD) plasma clearance, urinary excretion of 6-hydroxy-SDD and the in vitro microsomal SDD-hydroxylations were only inhibited when rBST was given in three injections per 24 hours. The hepatic microsomal ethylmorphine N-demethylation rate and the testosterone hydroxylation rate at the 6 beta position were significantly reduced after one rBST injection per 24 hours. Microsomal testosterone hydroxylation rates at the 16 alpha and 2 alpha-positions were reduced depending on the frequency of rBST administration. It is concluded that the inhibition of in vivo and in vitro drug oxidation in rats by rBST is associated with selective changes in activity of cytochrome P450 enzymes in the liver.

Effects of triiodothyronine treatment on pharmacokinetic properties and metabolite formation of antipyrine in dwarf goats.

The influence of triiodothyronine (5 micrograms/kg of body weight, sc, q 12 h for 7 days) on antipyrine (AP, 25 mg/kg, IV) plasma elimination and urinary metabolite excretion was studied in castrated male dwarf goats. After triiodothyronine treatment, a significant increase in AP elimination was found. However, the observed changes in clearances for production of AP metabolites (nor-AP, 3-hydroxymethyl-AP; 4-hydroxy-AP, and 4,4'-dihydroxy-AP) do not suggest a clear selectivity of triiodothyronine toward any of the metabolic pathways of AP.

Effects of Ehrlichia phagocytophila infection on serum thyroid hormone concentrations and on antipyrine clearance and metabolite formation in dwarf goats.

The influence of infection with Ehrlichia phagocytophila (EP) on serum thyroid hormone concentrations and on antipyrine (25 mg/kg of body weight, IV) plasma elimination and urinary metabolite excretion was studied in castrated male dwarf goats. Mean thyroid hormone concentrations moderately decreased in EP-infected goats, with maximal decrease in total and free triiodothyronine and thyroxine serum concentrations of 56, 64, 23, and 19%, respectively. The estimated pharmacokinetic values of antipyrine (AP) in EP-infected goats were similar to those in the goats when healthy. However, glucuronidation of the AP-metabolites 3-hydroxymethyl-AP, 4,4'-dihydroxy-AP, and 4-hydroxy-AP was reduced during the febrile episode of the acute-phase response to EP infection.

Oral bioavailability of sulphamethoxydiazine, sulphathiazole and sulphamoxole in dwarf goats.

To get a better insight into the oral bioavailability of sulphonamides in ruminants, sulphamethoxydiazine (pKa 7.0), sulphathiazole (pKa 7.2), and sulphamoxole (pKa 7.4) were administered to dwarf goats (n = 5). The drugs were given at 2-week intervals by the intravenous or intraruminal route at a dose of 100 mg per kg body weight. After IV injection, the mean half-life (t1/2 beta in h +/- SEM) was 0.80 +/- 0.10 h, 2.35 +/- 0.38 h, and 3.36 +/- 1.25 h, for sulphathiazole, sulphamoxole, and sulphamethoxydiazine, respectively and the mean distribution volume (Vd beta) was 0.23 +/- 0.05 l/kg, 0.23 +/- 0.04 l/kg, and 0.33 +/- 0.02 l/kg. After intraruminal administration, the mean bioavailability varied from 86.0 +/- 11.8% for sulphamethoxydiazine to 46.6 +/- 4.3% for sulphamoxole, and 52.6 +/- 7.2% for sulphathiazole. The elimination half-life was significantly prolonged, probably due to a low rate of drug absorption from the gastrointestinal tract. In contrast to chloramphenicol, the sulphonamides studied were stable when incubated in rumen fluid at 39 degrees C.

Genistein induces breast cancer-associated aromatase and stimulates estrogen-dependent tumor cell growth in in vitro breast cancer model.

In breast cancer, the interaction between estrogen-producing breast adipose fibroblasts (BAFs) and estrogen-dependent epithelial tumor cells is pivotal. Local estrogen production is catalyzed by aromatase, which is differentially regulated in disease-free and tumorigenic breast tissue. The use of aromatase inhibitors to block local estrogen production has proven effective in treatment of estrogen-dependent breast cancer. However, a major problem during breast cancer treatment is the sudden onset of menopause and many women seek for alternative medicines, such as the soy isoflavone genistein. In this study, we show that genistein can induce estrogen-dependent MCF-7 tumor cell growth and increase breast cancer-associated aromatase expression and activity in vitro. We have previously developed an in vitro breast cancer model where the positive feedback loop between primary BAFs and estrogen-dependent MCF-7 tumor cells is operational, thereby representing a more natural in vitro model for breast cancer. In this model, genistein could negate the growth inhibitory action of the aromatase inhibitor fadrozole at physiologically relevant concentrations. These data suggest that soy-based supplements might affect the efficacy of breast cancer treatment with aromatase inhibitors. Considering the high number of breast cancer patients using soy supplements to treat menopausal symptoms, the increasing risk for adverse interactions with breast cancer treatment is of major concern and should be considered with care.

Sulfamethazine as a model compound to assess sex hormone-dependent cytochrome P-450 activity in rats.

Plasma disposition and urinary recovery of sulfamethazine (SMZ), its N4-acetylated metabolite (N4AcSMZ), and two of its hydroxylated metabolites [5-hydroxysulfamethazine (5OHSMZ) and 6-hydroxymethylsulfamethazine (6CH2OHSMZ)] were determined in male and female rats, in castrated males, and in rats pretreated with various steroid hormones. Male rats had a 2-fold higher SMZ plasma clearance than females, castrates, and males treated with flutamide (a testosterone antagonist). When castrated male rats were treated with testosterone or trenbolone, SMZ plasma clearance returned to normal values. Higher SMZ plasma clearance rates in the presence of androgens went together with higher urinary recoveries of the 6CH2OHSMZ metabolite. In hepatic microsomes of male rats lower apparent KM values, and higher Vmax values for 6CH2OHSMZ and 5OHSMZ formation were found than in microsomes of female rats. Castration or treatment of male rats with flutamide markedly reduced microsomal SMZ hydroxylation rates. Pretreatment of male or female rats with phenobarbital or triacetyl-oleandomycin had no effect on microsomal SMZ hydroxylation, whereas a continuous infusion with bovine somatotropin in male rats caused a marked decrease in SMZ hydroxylation rate. Finally, SMZ hydroxylation to 6CH2OHSMZ and 5OHSMZ in hepatic microsomes from male rats was strongly inhibited by monoclonal antibodies against P-4502C11. These results suggest that the male-specific P-4502C11 enzyme plays an important role in the hydroxylation of SMZ to 6CH2OHSMZ and 5OHSMZ in rats. SMZ seems a useful model compound to assess hormone effects on oxidative biotransformation (in vivo and in vitro) in rats.

Tiamulin selectively inhibits oxidative hepatic steroid and drug metabolism in vitro in the pig.

The simultaneous use of the antibiotic tiamulin with certain ionophoric antibiotics (monensin, salinomycin) may give rise to a toxic interaction in pigs and poultry. In the present study, effects of tiamulin on hepatic cytochrome P450 activities in vitro were studied using pig liver microsomes. When tiamulin was added to the incubation medium the N-demethylation rate of ethylmorphine and the hydroxylation of testosterone at the 6 beta- and 11 alpha-positions was strongly inhibited. Tiamulin inhibited these activities more than SKF525A or cimetidine, but less than ketoconazole. The microsomal N-demethylation rate of erythromycin and the hydroxylation of testosterone at the 2 beta-position were inhibited to a lesser degree, whereas the ethoxyresorufin-O-deethylation, aniline hydroxylation and testosterone hydroxylations at the 15 alpha- and 15 beta-positions were not affected by tiamulin. No in vitro complexation by tiamulin of cytochrome P450 resulting in a loss of CO-binding capacity could be demonstrated. Results from the present study suggest a selective inhibition of cytochrome P450 enzymes in pigs, probably belonging to the P4503A subfamily. The mechanism of this interaction is still unclear. However, interactions between tiamulin and those veterinary drugs or endogenous compounds which undergo oxidative metabolism by P450 enzymes must be considered. More research is needed to reveal which of the P450 enzymes are affected by tiamulin in order to improve the understanding and probably the predictability of this interaction.

Cytochrome P-450 complex formation in rat liver by the antibiotic tiamulin.

Tiamulin is a semisynthetic diterpene antibiotic frequently used in farm animals. The drug has been shown to produce clinically important--often lethal--interactions with other compounds. It has been suggested that this is caused by a selective inhibition of oxidative drug metabolism via the formation of a cytochrome P-450 metabolic intermediate complex. In the present study, rats were treated orally for 6 days with tiamulin at two different doses: 40 and 226 mg/kg of body weight. For comparison, another group received 300 mg of triacetyloleandomycin (TAO) per kg, which is equivalent to the 226-mg/kg tiamulin group. Subsequently, microsomal P-450 contents, P-450 enzyme activities, metabolic intermediate complex spectra, and P-450 apoprotein concentrations were assessed. In addition, effects on individual microsomal P-450 activities were studied in control microsomes at different tiamulin and substrate concentrations. In the rats treated with tiamulin, a dose-dependent complex formation as evidenced by its absorption spectrum and an increase in cytochrome P-4503A1/2 contents as assessed by Western blotting (immunoblotting) were found. The effects were comparable to those of TAO. Tiamulin induced microsomal P-450 content, testosterone 6 beta-hydroxylation rate, erythromycin N-demethylation rate, and the ethoxyresorufin O-deethylation activity. Other activities were not affected or decreased. When tiamulin was added to microsomes of control rats, the testosterone 6 beta-hydroxylation rate and the erythromycin N-demethylation were strongly inhibited. It is concluded that tiamulin is a potent and selective inducer-inhibitor of cytochrome P-450. Though not belonging to the macrolides, the compound produces an effect on P-450 similar to those of TAO and related compounds.

The antibiotic tiamulin is a potent inducer and inhibitor of cytochrome P4503A via the formation of a stable metabolic intermediate complex. Studies in primary hepatocyte cultures and liver microsomes of the pig.

Tiamulin is a semisynthetic antibiotic frequently used in agricultural animals. The drug has been shown to produce clinically important--often lethal--interactions with other compounds that are simultaneously administered. To explain this, it has been suggested that tiamulin selectively inhibits oxidative drug metabolism via the formation of a cytochrome P450 metabolic intermediate complex. The aim of the present study was to provide further support for this hypothesis. When hepatic microsomes and cultured primary pig hepatocytes were incubated with tiamulin, a maximum in the absorbance spectrum at 455 nm was observed, which disappeared after adding KFe(CN)6. When hepatocytes were incubated with tiamulin for 72 hr, cytochrome P450 content and cytochrome P4503A apoprotein levels were increased. Tiamulin strongly inhibited and concentration dependently inhibited the hydroxylation rate of testosterone at the 6 beta-position in both microsomes and hepatocytes, and the microsomal N-demethylation rate of ethylmorphine. Other testosterone hydroxylations were inhibited to a lesser extent or not affected. The relative inhibition of the hydroxylation of testosterone at the 6 beta-position was more pronounced in microsomes from rifampicin- and triacetyloleandomycin-treated pigs. The results indicate that cytochrome P450 complex formation can at least partly explain the interactions observed with tiamulin. Tiamulin seems to be a strong, probably selective, inhibitor of the cytochrome P4503A subfamily and an interesting tool for further research.

Species- and sex-related differences in the plasma clearance and metabolite formation of antipyrine. A comparative study in four animal species: cattle, goat, rat and rabbit.

1. The plasma disposition of antipyrine, and its urinary metabolite pattern, were studied in both sexes of four animal species: rat, dwarf goat, rabbit and cattle. 2. No sex differences in plasma elimination of antipyrine were found in rabbit and goat; however, in rat and cattle the effect of sex was marked. As expected, male rat showed a higher plasma clearance value than female. In contrast bulls showed a significantly lower clearance value than cows. 3. Metabolite patterns varied widely from one species to another. The major urinary metabolite in rabbit and the two ruminant species was 4-hydroxy-antipyrine (OHA), whereas in rat 3-hydroxymethylantipyrine (HMA) was quantitatively the most important metabolite. 4. HMA was excreted in the 24 h urine in larger amounts by male rats than by females. Metabolism of antipyrine to HMA was also sexually different in the dwarf goat, but in this species females were more active than males. The effect of sex on the metabolite pattern in cattle was marked. 5. It is concluded that in ruminants there may be xenobiotic metabolic pathways which are under hormonal control, just as there are in rats and mice. If hormones influence drug metabolism in food-producing animals, residue levels of xenobiotics or their metabolites in food from animal origin may differ with the sex of the animal, or may be altered after treatment with anabolic hormones.

Effect of gonadal hormones on the plasma clearance and metabolite formation of antipyrine in the dwarf goat.

The effect of gonadal hormones on the plasma elimination and urinary metabolite profile of antipyrine was studied in dwarf goats. Female goats were treated with testosterone and male goats were treated with 17 beta-oestradiol. Castrated males were treated with either testosterone or 17 beta-oestradiol. Antipyrine (25 mg/kg, i.v.) was given both before and after the hormonal treatments. The effects of the hormonal status on the plasma elimination of the parent compound were not consistent. This was possibly due to the fact that formation of the main metabolite of antipyrine in the goat, 4-hydroxy antipyrine (OHA), was not affected by sex or hormonal treatment. On the other hand, there were clear effects of hormonal status on urinary excretion of the three other metabolites. In females and castrated males testosterone suppressed the formation of norantipyrine (NORA), 3-hydroxymethylantipyrine (HMA) and 4,4'-dihydroxyantipyrine (DOHA). Intact males produced smaller amounts of these metabolites than females. It is concluded that distinct xenobiotic metabolizing pathways exist in the dwarf goat, which are influenced in their activity by gonadal hormones. This confirms previous findings in rats and mice. The possibility that sex hormones influence drug metabolism in food-producing animals could have consequences for veterinary therapeutics and public health. This study also demonstrates that, when using the antipyrine test for the assessment of hepatic drug metabolism, it is very important to include the determination of metabolites.

Regioselective O-demethylation of scoparone (6,7-dimethoxycoumarin) to assess cytochrome P450 activities in vitro in rat. Effects of gonadal steroids and the involvement of constitutive P450 enzymes.

1. In the rat, scoparone (6,7-dimethoxycoumarin) is regioselectively O-demethylated in vitro by the hepatic cytochrome P450 (P450) system, yielding isoscopoletin and scopoletin. 2. Scoparone has been proposed as an indicator substrate for the assessment of P450 differentiation in vitro in rat. It has been suggested that isoscopoletin formation mainly reflects activity of enzymes of the P4501A subfamily, whereas scopoletin formation has been associated with P4502B activity. 3. In the present study, rate of formation of scopoletin and isoscopoletin were measured in hepatic microsomes from male, female and castrated male rats, in castrates treated with testosterone, in males treated with oestradiol, and in females treated with testosterone. Furthermore, effects of induction by phenobarbital (PB), beta-naphthoflavone (BNF), isoniazid, triacetyloleandomycin, and dexamethasone were studied in both sexes. 4. Scoparone metabolism was partly sex- and steroid-dependent. Variation of isoscopoletin formation with sex or hormonal status correlated well with ethylmorphine demethylation. 5. Scoparone-O-demethylation was regioselectively induced by PB and BNF. Induction effects were not very large and showed no sex differences. 6. Microsomal metabolism of scoparone was partly inhibited by a monoclonal antibody against P4502C11. Scopoletin and isoscopoletin formation were inhibited when ethylmorphine was added to the incubation mixture. Scoparone competitively inhibited testosterone 6 beta-hydroxylation. 7. It is concluded that a number of P450 enzymes, including those which are constitutively expressed, contribute to the biotransformation of scoparone. This lack of selectivity limits the usefulness of scoparone as a general indicator substrate for P450 differentiation in rats.

Selective effects of a bacterial infection (Actinobacillus pleuropneumoniae) on the hepatic clearances of caffeine, antipyrine, paracetamol, and indocyanine green in the pig.

1. In order to investigate the effect of a bacterial acute phase response model on drug disposition in vivo, plasma clearances of antipyrine, caffeine, paracetamol and indocyanine green were investigated in the healthy and Actinobacillus pleuropneumoniae-infected pig. 2. Indocyanine green plasma and endogenous creatinine clearance were not changed during the infection, which indicates that hepatic blood flow and renal function were not significantly affected. 3. In the A. pleuropneumoniae-infected pig, plasma clearances of antipyrine and caffeine, both marker substrates for hepatic oxidative biotransformation, were decreased by 72 and 68% respectively. The clearance of paracetamol, a drug mainly glucuronidated in the pig, was reduced by 39%. 4. It is concluded that the most important change in drug elimination during an acute phase response induced by A. pleuropneumoniae is a suppression of oxidative hepatic biotransformation.

Some pharmacokinetic data of aditoprim and trimethoprim in healthy and tick-borne fever infected dwarf goats.

Aditoprim (AP) is a new dihydrofolate reductase inhibitor, which is structurally related to trimethoprim (TMP). The pharmacokinetics of AP (10 mg/kg) and TMP (20 mg/kg) were assessed in healthy dwarf goats. Therapeutic efficacy against rickettsial infections was tested in tick-borne fever (TBF) infected goats. The animals were given TMP (n = 5) or AP (n = 5) by i.v. injection, and subsequently the drugs were administered orally (same groups, similar doses). Finally, both groups were infected with TBF and the i.v. experiment was repeated. Plasma concentration-time curves for both drugs followed first-order two-compartment decay. For TMP, mean t1/2 beta +/- SEM (h) was 0.84 +/- 0.06 (i.v. control) and 0.90 +/- 0.06 (i.v. infected), respectively, whereas for AP values of 8.00 +/- 0.31 (i.v. control) and 10.28 +/- 0.67 (i.v. infected) were obtained (P less than 0.05). Mean Vd beta +/- SEM values (l/kg) were 3.84 +/- 0.27 (i.v. control) and 4.07 +/- 0.85 (i.v. infected) for TMP (NS) and 7.02 +/- 0.63 vs 9.29 +/- 0.21 (P less than 0.05) for AP. After i.v. injection, rumen fluid concentrations of AP were significantly (P less than 0.05) higher and more persistent than those of TMP. For AP, the plasma and rumen fluid concentrations at 3 h were 1.20 +/- 0.06 micrograms/ml and 0.85 +/- 0.17 microgram/ml, respectively. After oral administration of TMP, Cmax in plasma was 0.12 +/- 0.01 microgram/ml and the maximum was reached after 1.2 +/- 0.16 h; systemic bioavailability (F) was 10.3% (relative to AUC i.v.). Oral treatment with AP resulted in a Cmax value of 0.21 +/- 0.02 microgram/ml with Tmax of 22.5 +/- 1.65 h and a F value of 71%. Based on WBC, serum ALP and rectal temperature responses, it was concluded that both TMP and AP were inactive against Ehrlichia phagocytophila.

Endotoxin-induced liver damage in rats is minimized by beta 2-adrenoceptor stimulation.

OBJECTIVE AND DESIGN: To investigate the effects of beta(2)-adrenoceptor (beta(2)-AR) stimulation on endotoxin-induced liver damage and systemic cytokine levels in rats. SUBJECTS: Standard male Wistar rats. TREATMENT: A disease-model of lipopolysaccharide (LPS)-induced acute systemic inflammation was used. The beta(2)-selective AR agonist clenbuterol was administered before, during, and after LPS-challenge to investigate its effects on the acute inflammatory response and associated liver-failure. METHODS: The following parameters have been measured in plasma: TNF alpha, IL-1 beta, IL-6, IL-10, AST, ALT, and Bilirubin. Liver histological examination was performed to look for changes in tissue morphology. RESULTS: Administration of clenbuterol (p.o.) one hour before, or intravenous at the same time as LPS-challenge resulted in a marked reduction of plasma levels of TNF alpha, IL-1 beta, and IL-6. A change both in plasma-level and in time-concentration profile of the anti-inflammatory cytokine IL-10 was found. Clenbuterol minimized LPS-induced liver damage, as represented by significantly lowered concentrations of several parameters for liver-failure (AST, ALT, Bilirubin), and improved hepatic tissue morphology. Clenbuterol administration after LPS challenge failed to inhibit TNF alpha-release but reduced liver-damage. Simultaneous use of the beta(2)-AR antagonist propranolol augmented LPS-induced liver failure, suggesting a role of endogenous adrenoceptor-agonists in prevention of organ-failure during systemic inflammation. CONCLUSIONS: The results indicate that a selective beta(2)-AR agonist might be used as an additional therapeutic agent in the clinic for the treatment of (acute) systemic inflammatory disorders in order to reduce or prevent subsequent liver failure.