Effects of the peroxisome proliferator ciprofibrate and prostaglandin F2α combination treatment on second messengers in cultured rat hepatocytes

Peroxisome proliferators induce hepatic peroxisome proliferation and hepatic tumors in rodents. These chemicals increase the expression of the peroxisomal β-oxidation pathway and the cytochrome P-450 4A family, which metabolizes lipids, including eicosanoids. Peroxisome proliferators transiently induce increased cell proliferationin vivo. However, peroxisome proliferators are weakly mitogenic and are not co-mitogenic with epidermal growth factor (EGF) in cultured hepatocytes. Earlier study found that the peroxisome proliferator ciprofibrate is comitogenic with eicosanoids. In order to study possible mechanisms of the comitogenicity of peroxisome proliferator ciprofibrate and eicosanoids, we hypothesized that the co-mitogenicity may result from synergistic or additive increases of second messengers in mitogenic signal pathways. We therefore examined the effect of the peroxisome proliferator ciprofibrate, prostaglandin F_2α (PGF_2α) and the combination of ciprofibrate and PGF_2α with or without growth factors on the protein kinase C (PKC) activity, and inositol-1, 4, 5-triphosphate (IP₃) and intracellular calcium ([Ca²⁺]_i) concentrations in cultured rat hepatocytes. The combination of ciprofibrate and PGF_2α significantly increased particulate PKC activity. The combination of ciprofibrate and PGF_2α also significantly increased EGF, transforming growth factor-α (TGF-α) and hepatic growth factor (HGF)-induced particulate PKC activity. The combination of ciprofibrate and PGF_2α greatly increased [Ca²⁺]_i. However, the increases of PKC activity and [Ca²⁺]_i by ciprofibrate and PGF_2α alone were much smaller. Neither ciprofibrate or PGF_2α alone nor the combination of ciprofibrate and PGF_2α significantly increased the formation of IP₃. The combination of ciprofibrate and PGF_2α, however, blocked the inhibitory effect of TGF-β on particulate PKC activity and formation of IP₃ induced by EGF. These results show that co-mitogenicity of the peroxisome proliferator ciprofibrate and eicosanoids may result from the increase in particulate PKC activity and intracellular calcium concentration but not from the formation of IP₃.     

Effect of Dexamethasone on Ciprofibrate-Induced Cell Proliferation and Peroxisome Proliferation

Peroxisome proliferators cause liver cell proliferation in additionto other pleiotropic effects such as peroxisome proliferationand induction of certain peroxisomal and cytosolic enzymes inliver. Since dexamethasone has been shown to inhibit mitogen-inducedliver cell hyperplasia, we examined whether dexamethasone inhibitsonly cell proliferation without affecting peroxisome proliferationinduced by peroxisome proliferators such as ciprofibrate. Liversof rats fed a diet containing ciprofibrate (0.025%) with orwithout added dexamethasone (0.5 mg or 1 mg/kg diet) for 1 weekwere evaluated for hepatocyte proliferation and peroxisome proliferation.Dexamethasone administration resulted in abrogation of ciprofibrate-inducedcell proliferation as shown by bromodeoxyuridine (BrdU) labelingand mitoses counts. The hepatocyte proliferative index measuredafter administration of a single dose of BrdU was 18.31.1 and2.30.7% (p< 0.01) in ciprofibrate and ciprofibrate + dexamethasonetreated rats, respectively. With multiple injections of BrdU(daily injections for 7 days) the proliferative index was 225 10 and 183 2% (p<0.02), respectively, in these two groups.Interestingly, whereas the levels of peroxisome proliferator-inducedM, 80,000 polypeptide and catalase and peroxisomal bifunctionalenzyme, and the corresponding mRNAs and peroxisome volume densitywere unaffected. These results show that dexamethasone selectivelyinhibits only cell proliferation without inhibiting the peroxisomeproliferation caused by ciprofibrate. This model should be usefulfor examining the role of cell proliferation versus oxidativestress in peroxisome proliferatorinduced hepatocarcinogenesis.     

Effects of the peroxisome proliferators ciprofibrate and perfluorodecanoic acid on hepatic cellular antioxidants and lipid peroxidation in rats.

The purpose of this study was to determine if hepatic cellular antioxidants and indices of oxidative damage are altered by administration of the peroxisome proliferators ciprofibrate and perfluorodecanoic acid (PFDA). Rats were fed 0.01% ciprofibrate in the diet or were injected with PFDA (0.5 or 5.0 mg/kg, i.p.) every 4 weeks for 6, 14, 30, 54, and 78 weeks. Peroxisomal fatty acyl-CoA oxidase and catalase activities were increased by both ciprofibrate and PFDA throughout the study. Neither ciprofibrate nor PFDA increased the levels of malonaldehyde or conjugated dienes, but ciprofibrate decreased these indices at early time points. Ciprofibrate decreased the following cellular antioxidants or antioxidant enzymes: vitamin C, vitamin D, DT-diaphorase, glutathione peroxidase, glutathione-S-transferase, and glutathione reductase; superoxide dismutase and glutathione were not affected. PFDA decreased DT-diaphorase and increased superoxide dismutase, but did not affect other cellular antioxidants. This study shows that administration of the peroxisome proliferators ciprofibrate and PFDA did not increase indices of lipid peroxidation, but that cellular antioxidant defenses were inhibited for a prolonged period of time by the peroxisome proliferator ciprofibrate.     

Long-term effects of peroxisome proliferators on the balance between hydrogen peroxide-generating and scavenging capacities in the liver of Fischer-344 rats

In order to clarify whether peroxisomal hydrogen peroxide (H2O2) plays an important role in peroxisome proliferator-induced hepatocarcinogenesis, we examined the change in metabolism of peroxisomal H2O2 in vivo and in vitro using male Fischer-344 rats fed clofibrate, bezafibrate and di(2-ethylhexyl)phthalate (DEHP) for up to 78 weeks. Hepatic peroxisomal fatty acyl-CoA oxidase activity increased 12-20-fold after 2 or 4 weeks treatment; later this level gradually decreased toward controls, and at 78 weeks activity was 3-10-times of control. Although hepatic H2O2 levels were increased slightly by clofibrate, bezafibrate and DEHP, the changes did not correlate with the changes in peroxisomal fatty acyl-CoA oxidase activity. In isolated hepatocytes, the rate of leakage of peroxisomal H2O2 from peroxisomes into the cytosol and the hepatocellular H2O2 content was measured. The rate of leakage of peroxisomal H2O2 into cytosol increased 2.5-4-fold when peroxisomal beta-oxidation activity was induced by peroxisome proliferators, and the increases in this rate corresponded with changes in the peroxisomal beta-oxidation activity. In contrast, the hepatocellular H2O2 contents were not affected by induced peroxisomal beta-oxidation. These data show that H2O2 leaking from peroxisome into cytosol would be quickly decomposed, and thus peroxisomal H2O2 does not appear to play an important role in hepatocarcinogenesis by such an oxidative stress mechanism after the long-term treatment with peroxisome proliferators.     

Effect of Rodent Hepatocarcinogenic Peroxisome Proliferators on Fatty Acyl-CoA Oxidase, DNA Synthesis, and Apoptosis in Cultured Human and Rat Hepatocytes

The effects of the rodent hepatocarcinogens clofibric acid and ciprofibrate on the activity of the peroxisomal fatty acyl-CoA oxidase, DNA synthesis, and apoptosis were compared in cultured rat and human hepatocytes. Rat hepatocytes expressed a 10-fold greater level of the peroxisomal fatty acyl-CoA oxidase compared to human hepatocytes. At the highest concentration (1.0 mM), both drugs induced a two- to threefold increase in this enzyme activity in both rat and human hepatocytes. Ciprofibrate (0.1 and 0.2 mM) caused a twofold increase in DNA synthesis in rat hepatocytes, whereas clofibric acid had no effect on DNA synthesis in these cells. In contrast, increasing concentrations of both clofibric acid and ciprofibrate produced inhibition of DNA synthesis in human hepatocytes. By using the terminal transferase dUTP–biotin nick end labeling technique, it was observed that 0.1 and 0.2 mM clofibric acid and ciprofibrate suppressed transforming growth factor-β (TGFβ)-induced apoptosis by 50% in rat hepatocytes, but they had no effect on TGFβ-induced apoptosis in human hepatocytes. Although clofibric acid and ciprofibrate diminished TGFβ-induced apoptosis, they had no effect on the basal apoptotic levels in the rat hepatocyte cultures. However, both drugs significantly increased the percent of apoptotic cells in the human hepatocyte cultures. It is concluded that primary rat and human hepatocyte cultures respond differently to peroxisome proliferators. The differences in effects on DNA synthesis and apoptosis support the hypothesis that human liver cells are refractory to peroxisome proliferator-induced hepatocarcinogenesis.     

Endotoxin induces the expression of prostaglandin H synthase-2 and eicosanoid formation in cells of the human monocytic cell line Mono Mac 6

In controlling inflammation, monocytes and macrophages not only secrete various cytokines but also eicosanoids. Since the human monocytic cell line, Mono Mac 6, represents cells at the stage of mature blood monocytes, cells of this line were investigated to find out whether they also resemble mature monocytes with respect to inflammation. In response to lipopolysaccharide (LPS), Mono Mac 6 cells secreted large amounts of eicosanoids, 6-keto-prostaglandin F_1α (6k-PGF_1α), thromboxane B₂ (TXB₂) and prostaglandin E₂ (PGE₂). The considerable increase in eicosanoid secretion is attributed to the expression of the inducible isoform of the prostaglandin H synthase (PGHS-2) since the PGHS-2-specific steady-state mRNA level, PGHS-2 protein, and the enzyme activity of PGHS were strikingly increased in the absence of detectable PGHS-1 protein. PGHS-2 expression was inhibited by either dexamethasone or cycloheximide. The non-steroidal anti-inflammatory drug (NSAID) diclofenac inhibited the increased PGHS activity. The expression of the PGHS-2 gene was accompanied by the formation of TNFα, IL-6 and IL-8, but not by IL-1β.     

Sodium naproxen: concentration and effect on inflammatory response mediators in human rheumatoid synovial fluid

Twelve patients suffering from rheumatoid arthritis and having swollen knees were treated with 1.1 g/day of sodium naproxen administered in one dose, daily for 5 days.The 72-h wash-out period was verified by the absence of any nonsteroidal anti-inflammatory drug using a HPLC screening. Blood and synovial fluid samples were drawn just before treatment and 24 h after the last dose.Eicosanoids (PGE₂, 6-keto-PGF₁, TXB₂, LTB₄, LTC₄) in synovial fluid were determined by immunoenzy-matic assays. In plasma and synovial fluid, hyaluronic acid was assayed by radiometric assay and sodium naproxen by HPLC. Free drug was determined by equilibrium dialysis. Statistical analysis used nonparametric tests.Pain relief (evaluated on a visual scale), morning stiffness, and scores on the Lee and Ritchie indices all decreased significantly, as did PGE₂ and LTB₄ concentrations. The decrease in 6-keto-PGF₁ and TXB₂ was not significant. No significant change was found for LTC₄ and hyaluronic acid.Total concentrations of sodium naproxen were equivalent in plasma (16.1 g·ml^–1) and synovial fluid (18.9 g·ml^–1). Free fractions were significantly higher in synovial fluid (0.14%) than in plasma (0.11 %), as shown by binding of the drug to human serum albumin, at various protein concentrations.Interestingly, the clinical efficacy, as shown by decreases in morning stiffness and in the Lee index score, correlated with the free concentration of naproxen in synovial fluid.     

Effect of phenobarbital on hepatic eicosanoid concentrations in rats

Phenobarbital is an efficacious tumor-promoting agent in the liver. Studies using inhibitors of eicosanoid synthesis have suggested that eicosanoids are important in the promotion of hepatocarcinogenesis by phenobarbital. We therefore hypothesized that hepatic eicosanoid concentrations are altered following phenobarbital administration. Male Sprague-Dawley rats were fed one of four levels of phenobarbital (0, 0.02, 0.05, and 0.1%). Eight rats from each of the four groups were killed after 10, 24, and 44 days for determination of liver weight and for preparation of microsomes. No significant difference was found among rat weights; however, liver weights were significantly higher in rats fed phenobarbital. Assay of benzyloxyresorufin-O-dealkylase activity showed that cytochromes P-450 2B1 and 2B2 were induced in response to phenobarbital administration. Prostaglandin E₂ concentrations were found to be significantly decreased by phenobarbital treatment after 10 and 24 days, but not after 44 days. Prostaglandin F_2α levels were decreased only by the lowest dietary phenobarbital concentration. Hepatic concentrations of leukotriene C₄ were decreased significantly at 10 days and at 44 days (only for the group administered the highest percentage concentration of phenobarbital), but not at 24 days. These results show that the investigated eicosanoids are generally slightly decreased by phenobarbital administration. Elevated eicosanoid levels therefore do not appear to be necessary for the promoting activity of phenobarbital. Received: 20 February 1997 / Accepted: 21 April 1997     

The synovial prostaglandin system in chronic inflammatory arthritis: differential effects of steroidal and nonsteroidal anti-inflammatory drugs

1 The present study was undertaken to characterize the spectrum of arachidonic acid metabolites present in synovial effusions of patients with rheumatoid or psoriatic arthritis, and to compare changes in their concentration following a short-term treatment with 6α-methyl-prednisolone (6-MeP: 4-8 mg/day) or indoprofen (1.2 g/day), a nonsteroidal anti-inflammatory agent with proven synovial prostaglandin inhibitory effect.

2 Measurements of prostaglandin E₂ (PGE₂), thromboxane (TX) B₂, 6-keto-PGF_1α and PGF_2α were performed by radioimmunoassay techniques in synovial effusions obtained from 23 patients, and validated by thin-layer chromatographic analysis of the extracted immunoreactivity.

3 PGE₂ and TXB₂ accounted for more than 60% of the total immunoreactivity in untreated patients. The absence of any constant ratio between the different arachidonic acid metabolites detected in synovial fluid is consistent with a heterogeneous cellular origin of these compounds.

4 Indoprofen treatment was associated with a consistent reduction of synovial prostaglandin and thromboxane concentrations, ranging from 36% in the case of 6-keto-PGF_1α to 90% in the case of PGE₂.

5 In contrast, 6-MeP caused opposite changes on different metabolites originating via the cyclo-oxygenase pathway. Thus, 6-keto-PGF_1α concentrations were reduced by 35%, PGF_2α concentrations were increased by 30%, while PGE₂ and TXB₂ were unchanged following 6-MeP.

6 Although the mechanism(s) underlying the failure of 6-MeP to reduce synovial PGE₂ and TXB₂ levels are uncertain, the results of the present study clearly indicate that therapeutic doses of steroidal and nonsteroidal anti-inflammatory drugs cause quite distinct changes in arachidonic acid metabolism, which might be relevant to their specific therapeutic actions and side-effects.

     

Mediation of central prostaglandin effects by serotoninergic neurons

Ten days after administration of 5,6-dihydroxytryptamine, which causes degeneration of central serotoninergic neurons, the depressive behavioral effects of PGF₂ and PGE₂ were evidently inhibited. Central chemical serotoninectomy abolished the hyperthermic and hypertensive effects of PGF₂, but only slightly affected those of PGE₂. It is concluded that serotoninergic neurons mediate the depressive behavioral action of both PGF₂ and PGE₂. They also mediate the hyperthermic and hypertensive action of PGF₂ but not of PGE₂. This suggests that these prostaglandins have different central modes of action.     

Effects of exogenous prostaglandins on the release of leukotriene C4-like immunoreactivity and on coronary flow in indomethacin-treated anaphylactic guinea-pig hearts

Summary It is known that both vasoconstrictor cyclooxygenase products and sulfidopeptide-containing leukotrienes (LT) contribute to the biphasic coronary constriction observed in isolated perfused anaphylactic guineapig hearts. We have now investigated the effects of the cyclooxygenase inhibitor indomethacin and of several exogenous prostaglandins (PG) on the release of LTC₄-like immunoreactivity and on various symptoms of cardiac anaphylaxis. Indomethacin decreased basal coronary flow and delayed the onset of coronary vasoconstriction after antigenic challenge. Furthermore, indomethacin inhibited cardiac release of 6-keto-PGF₁ and thromboxane (TX) B₂ and simultaneously enhanced the antigen-induced release of LTC₄-like immunoreactivity significantly. Neither the vasodilators PGE₂ and PGI₂ nor the vasoconstrictors PGF₂, PGD₂ and 11,9-epoxymethano-PGH₂, a compound with biological properties similar to TXA₂, affected the anaphylactic release of immunoreactive LTC₄ in the presence of indomethacin. These results suggest that the indomethacin-induced increase in LT release is not due to inhibition of synthesis of a cyclooxygenase product, which normally curbs anaphylactic release of immunoreactive LTC₄. The indomethacin effect may rather be explained by diversion of arachidonic acid metabolism away from fatty acid cyclooxygenase towards the synthesis of lipoxygenase products.Although the various PG did not significantly affect cardiac release of LTC₄-like immunoreactivity, they antagonized the anaphylactic coronary contriction. This antagonism may be due to direct effects of the PG on vascular smooth muscle tone as well as to indirect effects on the release of anaphylactic mediators not related to LT like histamine and platelet-activating factor. Antigen-induced arrhythmias were completely suppressed by PGF₂, while PGE₂ and PGI₂ tended to decrease the incidence of arrhythmias and the other PG had no consistent effect. It is concluded that the pharmacological effects of the PG used on coronary flow and arrhythmias during cardiac anaphylaxis are not mediated by inhibition of release of LTC₄-like immunoreactivity.     

A comparison of the central actions of prostaglandins A1, E1, E2, F1α, and F2α in the rat

Prostaglandins (PGs) injected into the right lateral brain ventricle (i.v.c.) of the rat increased the sleeping time induced by hexobarbital, chloral hydrate, and ethanol. PGE₁ and PGE₂ intensified chlorpromazine-induced catalepsy, inhibited amphetamine hyperactivity, and significantly depressed the amphetamine-induced stereotypy. NA concentrations were decreased by PGE₁ and PGE₂ and were increased by PGF₂. PGF₂ increased both 5-HT and 5-HIAA levels in rat brain. Total ACh concentrations were increased by PGF₁ and PGF₂. PGE₁, PGE₂, and PGF₂ enhanced the turnover of NA, DA, and 5-HT. PGE₂ counteracted the decreased activity induced by -MT and abolished the hypothermic action of -MT. PGF₂ had little effect on the activity of PCPA pretreated rats, whereas the higher doses of PGF₂ increased body temperature in these animals.     

Molecular modelling of the rat peroxisome proliferator-activated receptor -α (rPPARα) by homology with the human retinoic acid X receptor α (hRXRα) and investigation of ligand binding interactions I: QSARs

The construction of a homology model of the ligand binding domain of the rat peroxisome proliferator-activated receptor-α (rPPARα) based on the crystal structure of the human retinoic acid X receptor-α (hRXRα) is reported. It is demonstrated that many known peroxisome proliferators are able to occupy the putative ligand binding site of the rPPARα, including clofibric acid, ciprofibrate, nafenopin and related compounds. The log. relative potency of several peroxisome proliferators can be quantitatively related (R=0.99) to their binding affinity and lipophilicity as measured by their distribution coefficients (logD_7.4 values) and other QSARs are discussed in the light of receptor–ligand interactions. The molecular modelling of a representative number of peroxisome proliferators within the putative ligand binding site is consistent with experimental information on relative potency and enantioselectivity.     

The effects of celecoxib, a COX-2 selective inhibitor, on acute inflammation induced in irradiated rats

The potential value of selective and non-selective COX-2 inhibitors in preventing some of the biochemical changes induced by ionizing radiation was studied in rats exposed to carrageenan-induced paw edema and 6-day-old air pouch models. The animals were exposed to different exposure levels of γ-radiation, namely either to single doses of 2 and 7.5 Gy or a fractionated dose level of 7.5 Gy delivered as 0.5 Gy twice weekly for 7.5 weeks. The inflammatory response produced by carrageenan in irradiated rats was markedly higher than that induced in non-irradiated animals, and depended on the extent of irradiation. Celecoxib, a selective COX-2 inhibitor, in doses of 3, 5, 10, and 15 mg/kg was effective in reducing paw edema in irradiated and non-irradiated rats in a dose-dependent manner as well as diclofenac (3 mg/kg), a non-selective COX inhibitor. Irradiation of animals before the induction of the air pouch by an acute dose of 2 Gy led to a significant increase in leukocytic count, as well as in the level of interleukin-6 (IL-6), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), LTB₄, PGE₂ (as an index of COX-2 activity), TXB₂ (as an index of COX-1 activity), and the plasma level of MDA. This increase in level of these parameters was more marked than that observed in the non-irradiated animals subjected to the inflammagen. The blood GSH level was not affected by the dose of irradiation used, whereas superoxide dismutase (SOD) activity was suppressed. In many respects, celecoxib (5 mg/kg) was as potent as diclofenac in decreasing the elevated levels of IL-6, IL-1β, TNF-α, LTB₄, PGE₂, but lacked any significant effect on TXB₂ level. Since it is mostly selective for COX-2 with a rare effect on COX-1 enzyme, both drugs at the selected dose levels showed no effect on level of MDA, GSH, and SOD activity.     

Effect of the peroxisome proliferator perfluorodecanoic acid on growth and lipid metabolism in Sprague Dawley rats fed three dietary levels of selenium

The possible interrelationships between the effects of dietary selenium and perfluorodecanoic acid (PFDA) on growth and lipid metabolism were studied in the male Sprague Dawley rat. Rats were divided into groups and placed on diets containing three levels of selenium (0.04, 0.2, and 1.0 ppm as sodium selenite). Two weeks later, half the rats in each group received a single 35 mg/kg IP injection of PFDA in corn oil, while their pair-fed companion received only vehicle. Rats injected with PFDA stopped gaining weight, and weighed less than pair-fed controls, despite equal food intakes. Two weeks following PFDA administration the rats were killed and plasma cholesterol and triglycerides, and liver peroxisomal enzyme activities were quantified. In contrast to other peroxisome proliferators, PFDA increased plasma triglycerides while decreasing plasma cholesterol. The rate of peroxisomal fatty acid -oxidation was decreased, even though the activity of fatty acyl-CoA oxidase, the first enzyme in the peroxisomal fatty acid -oxidation pathway, was increased. Dietary selenium, other than increasing the liver to body weight ratio, did not alter growth or lipid metabolism. This study demonstrates, for the first time, the existence of a non-hypotriglyceridemic peroxisome proliferator-PFDA.     

Modulation of noradrenaline release from the sympathetic nerves of human right atrial appendages by presynaptic EP3- and DP-receptors

Strips of human right atrial appendages were preincubated with [³H]noradrenaline and then superfused with physiological salt solution containing inhibitors of uptake₁ and uptake₂. Tritium overflow was evoked by transmural electrical stimulation (standard frequency: 2 Hz). Prostaglandin E₂ (PGE₂) inhibited the electrically evoked tritium overflow; at the highest concentration investigated, tritium overflow was reduced by about 80% and the pIC_50% value was 7.14. The effect of PGE₂ was not affected by rauwolscine, which, by itself, increased the evoked overflow. Naproxen failed to affect the evoked tritium overflow and its inhibition by PGE₂. The inhibitory effect of PGE₂ on the electrically evoked tritium overflow was mimicked by prostaglandin E₁, the EP₁/EP₃-receptor agonist sulprostone and the EP₂/EP₃-receptor agonist misoprostol with the rank order of potency (pEC_50%): sulprostone (7.68) > misoprostol (7.10) > PGE₁ (6.39). In contrast, PGF_2α, the IP/EP₁-receptor agonist iloprost and the stable thromboxane A₂ analogue U46619 (9,11-dideoxy-11α,9α-epoxy-methanoprostaglandin F_2α) did not change evoked tritium overflow. PGD₂ caused facilitation. These results suggest that the sympathetic nerve fibres innervating human atrial appendages are endowed with presynaptic inhibitory EP₃ and facilitatory DP-receptors. The EP₃-receptors appear not to be tonically activated and do not interact with the α₂-autoreceptors. Received: 11 May 1998 / Accepted: 29 July 1998     

Lack of induction of hepatic DNA damage on long-term administration of peroxisome proliferators in male F-344 rats

In order to evaluate the relationship between hydrogen peroxide (H2O2) generation and subsequent DNA damage caused by peroxisome proliferation, we examined DNA damage and changes in peroxisomal beta-oxidation activity in rat liver. Male F-344 rats were given orally clofibrate, bezafibrate or di(2-ethylhexyl)phthalate (DEHP) for up to 78 weeks. In rats fed DEHP for 52 or 78 weeks hepatocarcinomas or neoplastic nodules were found. In rats treated for 2 weeks with peroxisome proliferators, peroxisomal beta-oxidation activity was increased 10-17 times over control levels. After long-term treatment (20-78 weeks), the level of peroxisomal beta-oxidation activity remained 3-13-times higher in each group. When single strand DNA breaks were measured by a DNA-alkaline elution technique, no increase in DNA damage was observed in livers from rats fed peroxisome proliferators for 2, 40 or 78 weeks. In rats bearing hepatocarcinomas induced by DEHP, the hepatic DNA showed significant breaks; the rate of DNA-alkaline elution was found to increase approximately 5-fold. No significant increase in hepatic lipid peroxide level was observed in each group. These results show that although prolonged treatment with peroxisome proliferators induces markedly peroxisomal beta-oxidation activity, the active oxygen species from peroxisomal beta-oxidation are not enough to give rise to significant DNA damage. Moreover, the change in the activity of peroxisomal beta-oxidation may not relate to hepatocarcinogenesis induced by peroxisome proliferators.     

Lowering of the convulsive threshold by non-steroidal anti-inflammatory drugs

Administration of convulsant doses of pentetrazole induced an increase of PGF_2α, PGE₂, and TXB₂ immunoreactive material in mouse brain in vivo. The non-steroidal anti-inflammatory drugs indomethacin, flurbiprofen and diclofenac in equipotent doses inhibited the convulsion-induced increase of prostaglandins and thrombaxane B₂ (P < 0.01). The same drugs also lowered the LD₅₀ of pemtetrazole (P < 0.05) and accelerated the onset of tonic seizures evoked by pentetrazole (P < 0.05). Ibuprofen in a submaximal centrally effective dose acted in the same way on cerebral PG and TXB₂ synthesis and on then LD₅₀ of pentetrazole but failed to influence significanlty the latency time to the onset of pentetrazole-induced tonic seizures. Aspirin (100 mg/kg) and paracetamol (30 mg/kg), which were without effect on cerebral PG and TXB₂ concentrations following pentetrazole-induced seizures, were also ineffective in lowering the convulsive threshold and the LD₅₀ of pentetrazole. It is concluded that non-steroidal anti-inflammatory drugs act on the convulsive threshold by inhibition of cerebral prostaglandin and/or thromboxane synthesis.     

Effects of joint exposures to selected peroxisome proliferators on hepatic acyl-CoA oxidase activity in male B6C3F1 mice.

The interaction potential of peroxisome proliferators of similar and dissimilar structure was examined in B6C3F1 mice. Mice were fed diets containing varying concentrations of ciprofibrate (Cipro), clofibrate (Clof) or di(2-ethylhexyl)phthalate (DEHP), or combinations of Cipro and Clof or Cipro and DEHP for 4 d. Induction of peroxisomal beta-oxidation, measured by increased acyl-CoA oxidase activity, was used as the endpoint for analysis. An additive response occurred following joint exposure to the structurally related compounds Cipro and Clof, whereas a possible synergistic response occurred at low dose combinations of the structurally dissimilar Cipro and DEHP. These findings represent the first report assessing the in-vivo interaction potential of structurally similar and dissimilar peroxisome proliferators and provides insight into the dose-response nature of joint exposures to certain non-genotoxic carcinogens.     

Effects of prostaglandins F2α and E1 on the longitudinal and circular smooth muscle of the guinea pig caecum in relation to the concentration of extracellular calcium

Summary Responses of the longitudinal and circular smooth muscle of the guinea-pig caecum were studied in media with different concentrations of extracellular calcium. A sucrose-gap method was used to record the electrical and mechanical parameters of smooth muscle activity.In the longitudinal smooth muscle no difference in the action of PGF₂ and PGE₁ was seen at low (0.8 mM), normal (2.5 mM) or high (7.5 mM) concentrations of Ca²⁺. Both PG's stimulated the longitudinal strip and the action of acetylcholine at normal Ca²⁺ concentration was slightly inhibited immediately after the superfusion with the PG's and augmented thereafter. In the circular strip no significant difference between the stimulatory effect of PGF₂ and PGE₁ in low and high Ca²⁺ solution was found. However, at 2.5 mM Ca²⁺ PGF₂ evoked much greater stimulation of the circular strip than did PGE₁. After the end of the superfusion with either PG the effect of acetylcholine was inhibited by either PG at low Ca²⁺ level and potentiated at high Ca²⁺ level; at normal Ca²⁺ level the effect of acetylcholine was inhibited by PGE₁ but not by PGF₂.Thus, the effects of PGF₂ and PGE₁ on the circular smooth muscle differed from each other in their dependence on the concentration of extracellular calcium.