bluuebunny/crossref_metadata_2025 huggingface dataset:
Influence of Bipyridylium Compounds on Microsomal Mixed‐Function Oxidation Activities
Authors: Zychlinski Lech , Raska‐Emery Patricia , Montgomery Mark R.
Published: October 2006

The two principal bipyridyl herbicides, paraquat and diquat, were investigated for their influence on microsomal mixed‐function oxidation (MFO) activities and on NADPH oxidation rates in lung, liver, and kidney preparations. In lung microsomal preparations, benzphetamine N‐demethylation was found to be inhibited by paraquat and diquat in a concentration‐dependent manner, but ethylmorphine N‐demethylation was unaffected by these bipyridyls. In liver microsomal fractions, both benzphetamine and ethylmorphine N‐demethylases were inhibited by paraquat and diquat. Neither bipyridyl affected MFO activity in kidney preparations. A kinetic investigation of the enzyme inhibition showed that only Vmax was affected by paraquat and diquat, providing the first evidence for noncompetitive inhibition by the bipyridyls. In all microsomal preparations, NADPH oxidation was stimulated significantly by paraquat and to an even greater extent by diquat in the absence or presence of benzphetamine or ethylmorphine. The influence of MFO substrates on the stimulation varied widely among the three organ systems. In lung, paraquat‐ or diquat‐mediated stimulation of NADPH oxidation was equal in the absence of MFO substrates and in the presence of ethylmorphine, but the stimulation was increased in the presence of benzphetamine. Stimulation of NADPH oxidation by the bipyridyls, in liver as well as in kidney preparations, was equal in all situations in the absence of MFO substrates and in the presence of benzphetamine or ethylmorphine, although the quantity of this stimulation was greater in liver than in kidney fractions. It is apparent that the bipyridyls are potent stimulators of in vitro NADPH oxidation in microsomal preparations from several organs. The quantity of the NADPH oxidation stimulation seems to be a decisive factor in the inhibition of xenobiotic metabolism. Whether the stimulation of NADPH oxidation and the noncompetitive inhibition of xenobiotic metabolism play a significant role in bipyridyl toxicity are under further investigation.
Liver Cell Calcium Homeostasis in Carbon Tetrachloride Liver Cell Injury: New Data With Fura21
Authors: Dolak James A. , Waller Robert L. , Glende Eric A. , Recknagel Richard O.
Published: October 2006

The calcium fluorescent probe fura2 was used to measure concentration of free calcium in the cytosol of isolated rat hepatocytes in suspension. The resting level in untreated hepatocytes was 121 nM. On addition of CCl4 at a concentration of 0.5 mM, cytosolic free calcium rose sharply and reached a statistically significant (P<0.05) steady plateau level of about 190 nM within five minutes. With a concentration of 1.0 mM CCl4, cytosolic free calcium rose within ten minutes to a plateau level of about 200 nM. Use of fura2, along with the capacity of Mn2+ ions to effectively quench fura2 fluorescence, provided the basis for a simple and decisive method to determine whether the added CCl4 was permeabilizing the hepatocyte plasma membrane by direct solvent action. It was found that up to a concentration of 1.0 mM, CCl4 did not permeabilize the plasma membrane, but direct attack on the plasma membrane was unequivocally demonstrated for concentrations of 2 mM CCl4 and above. Finally, an hypothesis is presented for resolution of the puzzling dilemma that emerged from the observation, reported from two laboratories, that CCl4 can rapidly mobilize liver mitochondrial calcium despite the well‐known relative resistance of these organelles to the damaging effects of this toxic agent.
Chromatographic and catalytic properties of multiple forms of rabbit pulmonary cytochromes P‐450
Authors: Ueng Tzuu‐Huei , Peiperl Martha D. , Alvares Alvito P.
Published: October 2006

Rabbit pulmonary cytochrome P‐450 forms 2,5, and 6 were resolved using anion‐exchange high‐performance liquid chromatography (HPLC) and their properties compared with rabbit liver cytochrome P‐450 isozymes LM2 and LM6. Although rabbit pulmonary form 2 and liver LM2 had similar electrophoretic mobilities and similar substrate specificities in reconstitution experiments, they differed in their HPLC elution profiles. High‐performance liquid chromatography of pulmonary microsomes from rabbits treated with 3‐methylcholanthrene (3‐MC) revealed the induction of form 6 isozyme, which had a retention time, electro‐phoretic mobility, and substrate specificity similar to those of rabbit liver LM6. In reconstitution experiments, forms 2 and 6 showed the highest substrate specificities toward benzphetamine and 7‐ethoxyresorufin, respectively. Rabbit lung cytochrome P‐450 form 5 was relatively inactive toward all substrates tested.
Lack of inducibility of brain monooxygenase activities including parathion desulfuration
Authors: Chambers Janice E. , Forsyth Carol S.
Published: October 2006

The ability of phenobarbital and β‐naphthoflavone to induce parathion desulfuration, aminopyrine N‐demethylation, and NADPH–cyto‐chrome‐c reductase activity in the brain and liver of male and female rats was investigated. Activities of all three enzymes were found in similar levels in both the mitochondrial and microsomal fractions of brain. There were no sex differences in brain activities. Liver activities were from 10‐ to 30‐fold higher than brain activities when computed on a tissue‐wet‐weight‐equivalent basis. Although exposure to both inducers increased all three enzyme activities and cytochrome P‐450 in liver, neither inducer increased the enzyme activities in mitochondrial or microsomal brain fractions of either sex. Thus, these brain monooxygenase activities appear to be refractory to induction by two classical types of cytochrome P‐450 inducers. This lack of inducibility could serve to protect the animal against environmentally enhanced increases in the activation of xenobiotics to neurotoxic metabolites, such as parathion desulfuration to paraoxon.
Drug‐induced protoporphyria in the olfactory mucosa of the hamster
Authors: Reed Celia J. , van den Broeke Leon T. , De Matteis Francesco
Published: October 2006

Administration of 3,5‐diethoxycarbonyl‐4‐ethyl‐1,4‐dihydro‐2,6‐dimethylpyridine (4‐ethyl‐DDC) to hamsters resulted in a marked loss of cytochrome P‐450‐dependent reactions (peroxidase, 7‐ethoxycoumarin O‐deethylase, and 7‐ethoxyresorufin O‐deethylase) in both liver and olfactory epithelium within 2 hr. This inactivation of cytochrome P‐450 was accompanied by inhibition of ferrochelatase (FK), stimulation of 5‐aminolevulinate synthase (ALA‐S), and accumulation of protoporphyrin both in the liver and to a lesser degree, in the olfactory epithelium. These results suggest that the mechanism of induction of protoporphyria in nasal tissues is similar to that occurring in the liver, namely, suicidal metabolism of 4‐ethyl DDC by cytochrome P‐450 resulting in formation of N‐ethylprotoporphyrin, a potent inhibitor of FK. The consequent depletion of heme leads to stimulation of ALA‐S and, thus, porphyrin accumulation. Investigation of the dose‐response to 4‐ethyl DDC demonstrated that, in liver, maximal inhibition of FK and accumulation of protoporphyrin occurred at a dose of 50 mg/kg while ALA‐S activity continued to increase up to a dose of 100 mg/kg. This is compatible with an additional effect of the drug on ALA‐S involving induction of cytochrome P‐450 and, thus, further depletion of heme. In the olfactory epithelium, stimulation of ALA‐S was significantly less marked, suggesting that this secondary effect does not operate in nasal tissue. This is consistent with reports that olfactory cytochrome P‐450s are noninducible.
Induction of phosphatidylcholine biosynthesis via cdpcholine pathway in lung and liver of rats following intratracheal administration of ddt and endosulfan
Authors: Narayan Satya , Dani H. M. , Misra U. K.
Published: October 2006

The induction of phosphatidylcholine (PC) biosynthesis via the CDPcholine pathway in lung and liver of rats has been shown following the intratracheal administration of 1,1,1‐trichloro‐2m2‐bis(p‐chlorophenyl) ethane (DDT) (5 mg/100 g body weight) and endosulfan (1 mg/100 g body weight) for 3 days. Controls received only the vehicle solution (groundnut oil, 0.1 ml/100 g body weight). The treatment of DDT and endosulfan significantly increased the PC contents and the incorporation of radioactive [methyl‐3H]choline into PC of lung and liver microsomes. The incorporation of radioactive [methyl‐14C]methionine into microsomal PC of lung and liver was not affected significantly by treatment with either of the insecticides. 1,4,5,6,7‐hexachloro‐5‐norbornene‐2,3‐dimethano cyclic sulfite (endosulfan) administration significantly increased the activity of choline kinase and phosphocholine cytidylyl‐transferase (both cytosolic and microsomal) of lung, whereas DDT increased the activity of only the latter. In liver, both DDT and endosulfan administration significantly increased the activity of choline kinase and phosphocholine cytidylyltransferase (both cytosolic and microsomal). However, the activity of phosphocholinetransferase was not affected in both lung and liver microsomes of rats treated with these insecticides. The PC precursor pool sizes, choline and phosphorylcholine, of lung and liver tissues were not altered by DDT and endosulfan treatments. The present results suggest that the increased level of PC and incorporation of radioactive [methyl‐3H]choline into microsomal PC could be the result of increased activity of choline kinase and phosphocholine cytidylyltransferase of lung and liver of rats following intratracheal administration of DDT and endosulfan.
Acetaminophen metabolism in vivo by pregnant, fetal, and neonatal guinea pigs
Authors: Ecobichon D. J. , Hidvegi S. , Comeau A. M. , Varma D. R.
Published: October 2006

Acetaminophen (50 mg/kg body weight) was administered by iv injection to pregnant guinea pigs (60–65 days of gestation) and by ip injection to cesarian‐derived term (67 days of gestation) pups. At suitable time intervals after treatment, the concentrations of drug, glucuronide (GLU), and sulfate (SO4) in blood plasma, urine, and bile were measured by high‐performance liquid chromatography (HPLC). At 60–65 days of gestation, guinea pig fetuses formed both GLU and SO4, an approximate ratio of 2:1 being observed with mean concentrations of the order of 43 and 27 μg/mL being measured for GLU and SO4, respectively at 180 min post‐treatment. At the same time interval, the major detoxification product found in the blood plasma of the pregnant dams was GLU (104 μg/mL) with only minute amounts (4.2 μg/mL) of SO4 being detected. In cesarian‐derived and acetaminophen‐treated pups, euthanized at 2 or 4 hr post‐treatment, plasma levels of GLU were approximately twofold higher relative to the concentration of SO4 at both time intervals. Significant differences were not observed in either bile or urine at 2 hr post‐treatment but by 4 hr after treatment the levels of GLU found in the bile and urine were two‐ or threefold higher than those of SO4. In contrast to the adult guinea pig where GLU forms some 90% of the urinary excretory product and SO4 accounts for only 7%, the SO4 pathway of detoxification appears to be of significant importance to the fetal and neonatal animal.
The sulfur‐cyanolysis sites of serum albumin: Metabolite competition studies
Authors: Jarabak Rebecca , Westley John
Published: October 2006

In the presence of a source of sulfane sulfur, a cyanolysis reaction catalyzed by serum albumin may contribute to cyanide detoxication. The active site for this catalysis by serum albumin has been investigated in competition studies with ligands that have known albumin binding sites. Despite complications caused by the occurrence of multiple primary and secondary sites for many ligands, the results show that the primary sites for bilirubin, steroids, indoles, aspirin, and palmitate are distinct from that for sulfur. Laurate is a tight‐binding partial inhibitor of the cyanolysis reaction, competitive with cyanide rather than with sulfur. In view of the formal mechanism previously established for the catalyzed reaction, this result indicates that the sulfur‐cyanolysis site is probably near the site occupied by laurate.
The role of enteric bacteria in the anerobic metabolism of 5‐aminosalicylate
Authors: Tucker Mark A. , Bisesi Michael S. , Smith Timothy J.
Published: October 2006

The primary (and inactive) enteric metabolite of 5‐aminosalicylate is N‐acetyl‐5‐amino‐salicylate. Previous studies have demonstrated acetylation of this anti‐inflammatory agent by intestinal and bacterial homogenates. To assess the contribution of anerobic bacteria to the N‐acetylation in vivo, we have measured the production of N‐acetyl‐5‐aminosalicylate in anerobic microculture. Our results indicate that enteric bacteria play a minor role in N‐acetylation, but may contribute to the production of other metabolites of pharmacologic and toxicological interest.
Multiple Pathways for the Oxidative Metabolism of Estrogens in Syrian Hamster and Rabbit Kidney
Authors: Degen G. H. , Blaich G. , Metzler M.
Published: October 2006

Microsomal preparations from hamster kidney, a target tissue for the carcinogenic action of stilbene‐type and steroidal estrogens, catalyze the oxidative metabolism of diethylstilbestrol (DES). The formation of the major metabolite Z,Z‐dienestrol and of reactive intermediates capable of protein binding were mediated by enzyme activities requiring nicotinamide‐adenine dinucleotide phosphate (reduced form‐NADPH), cumene hydroperoxide, or arachidonic acid (ARA). In addition, hydroxylated DES metabolites were detected in NADPH‐supplemented incubations. The NADPH‐dependent oxidation of DES was inhibited by SKF 525A and metyrapone. Monooxygenase‐catalyzed metabolism was apparently responsible for the majority of DES oxidation in microsomes from whole hamster kidneys in vitro and this activity is preferentially localized in the kidney cortex. However, ARA‐dependent, i.e., prostaglandin H synthase (PHS) mediated oxidation of DES and of the catechol estrogen 2‐hydroxyestrone was demonstrated as well in the medulla of both rabbit and hamster kidney. It is proposed that monooxygenase and PHS activities act in concert in the metabolic activation of carcinogenic estrogens. This appears to apply in particular to steroidal estrogens, since catechol estrogens formed by monooxygenases are further oxidized to reactive intermediates by PHS and other peroxidatic enzymes.
Strain and sex differences in the response of mice to drugs that induce protoporphyria: Role of porphyrin biosynthesis and removal
Authors: Holley Anne , King L. J. , Gibbs A. H. , de Matteis F.
Published: October 2006

A hepatic green pigment, inhibitory toward ferrochelatase, has been isolated from the liver of mice treated with griseofulvin, isogriseofulvin, or 3,5‐diethoxycarbonyl‐1,4‐dihydrocollidine and has been shown to exhibit identical chromatographic characteristics to authentic N‐methyl protoporphyrin. All four possible structural isomers have been demonstrated, and each drug produced primarily the same isomer. N‐Methyl protoporphyrin has also been found in very small amounts in the liver of untreated mice, but the isomeric composition appeared to differ from that of the drug‐induced N‐methyl protoporphyrin.

Intraperitoneal administration of 3,5‐diethoxycarbonyl‐1,4‐dihydrocollidine to female C3H / He / Ola and NIH / Ola inbred mice produced a marked dose‐related loss of hepatic ferrochelatase activity, which was identical in magnitude in the two strains. Induction of hepatic 5‐aminolevulinate synthase (ALA‐S), and accumulation of liver protoporphyrin, however, were greater in C3H / He / Ola mice. The strain difference in ALA‐S response was most marked when inhibition of ferrochelatase (the “specific” effect of the drug) was maximal, and this suggests that a genetic variation exists in the sensitivity of ALA‐S to a second drug action, the so‐called nonspecific action, which is shared by many lipid‐soluble compounds.

Male mice of three strains accumulated greater amounts of hepatic protoporphyrin than females after treatment with griseofulvin, yet no significant difference was found between the two sexes in the extent of ferrochelatase inhibition. Stimulation of ALA‐S activity was slightly greater in males, but when porphyria was very marked, ALA‐S activities were significantly lower in this sex. Our results indicate a biphasic relationship between ALA‐S activity and porphyrin accumulation, with a depression of the activity of the enzyme occurring at high liver protoporphyrin concentrations.
Oxidant injury increases cell surface receptor binding of angiotensin ii to pulmonary artery endothelial cells
Authors: Patel J. M. , Madhavi Sekharam K. , Block Edward R.
Published: October 2006

Nitrogen dioxide (NO2), an environmental oxidant, is known to activate phospholipase A1 and modulate the plasma membrane structure of porcine pulmonary artery endothelial cells. We evaluated the effects of exposure to NO2, purified phospholipase B (which acts as phospholipase A1 and A2), or phospholipase A2 on 125I‐angiotensin II (Ang II) receptor binding, internalization, or both in pulmonary endothelial cells. Exposure to 5 ppm NO2 for 48 hr at 37°C or 0.075 U each of phospholipase B or A2 in phosphate‐buffered saline (PBS) for 30 min at 24°C resulted in an increase in total Ang II binding (i.e., cell surface bound and internalized) by 45% (p <0.05), 50% (p <0.05), and 85% (p <0.001), respectively, compared to controls. An Ang II receptor antagonist, [Sar1 IIe8] Ang II, competitively displaced Ang II binding to control, NO2‐, phospholipase B‐, and phospholipase A2‐exposed cells. Dissociation of bound Ang II in the presence of PBS was <1% of total bound Ang II in control, NO2‐, and phospholipase B‐exposed cells and was 50% of total bound Ang II in phospholipase A2‐exposed cells. In the presence of isotonic acetic acid/NaCl, in excess of 90% of cell surface‐bound Ang II was dissociated from control, NO2‐, and phospholipase B‐exposed cells, and there was <2% of Ang II detectable when acid‐treated cells were subjected to NaOH solubilization. In cells exposed to phospholipase A2, acetic acid treatment did not release cell‐bound Ang II, and the remaining Ang II was recovered in the NaOH solubilized fraction. These results demonstrate that NO2 exposure results in increased cell surface receptor binding and not internalization of Ang II and that phospholipase B‐but not phospholipase A2‐induced modulation of the membrane structure mimicked the effect of NO2 on Ang II binding in pulmonary endothelial cells.
Role of hypergastrinemia in the antiatrophy effect of 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin on oxyntic gland mucosa of the rat stomach
Authors: Michael Theobald H. , Mably Thomas A. , Ingall Glynnis B. , Peterson Richard E.
Published: October 2006

Atrophy of the gastrointestinal mucosa that occurs in pair‐fed control rats is not observed in 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD)‐treated rats (1). Our objective was to determine if the gastrointestinal trophic hormone, gastrin, is involved in the antiatrophy effect of TCDD on the gut mucosa. Adult male Sprague‐Dawley rats treated with 100 μg/kg of TCDD were slightly hypergastrinemic 7 days after dosing and markedly hypergastrinemic 14 days after treatment whereas pair‐fed control rats were normogastrinemic. After 14 days of feed restriction, atrophy of the oxyntic gland and ileum mucosa occurred in pair‐fed control rats but only atrophy of the ileum mucosa developed in TCDD‐treated animals. The oxyntic gland mucosa of TCDD‐treated rats was protected from mucosa atrophy as well as from mucosa erosions. The protection against feed restriction‐induced atrophy was demonstrated by measurements of oxyntic gland mucosal height and DNA and protein content. Since hypergastrinemia stimulates growth of oxyntic gland mucosa, but not ileum mucosa, the antiatrophy effect of TCDD on mucosa of the oxyntic gland might in part be due to hypergastrinemia. In support of this interpretation, TCDD treatment exerted an antiatrophy effect on the oxyntic gland mucosa only when TCDD‐treated animals were hypergastrinemic. For example, hypergas‐trinemia does not develop within the first 48 hr after TCDD administration, and TCDD treatment affords no protection against fasting‐induced atrophy of the oxyntic gland mucosa during this time. On the other hand, the ability of TCDD treatment to protect against feed restriction‐induced erosions of the oxyntic gland mucosa might be mediated by hypergastrinemia since these events occur at a later time. It is unlikely, however, that increased gastric mucosa production of cytoprotective prostaglandins is involved in the antierosion action of TCDD, because TCDD treatment does not protect the oxyntic gland mucosa against ethanol‐ or indomethacin‐induced erosions. The significance of these findings is that this is one of the first reports of a gastrointestinal trophic hormone, gastrin, being involved in the mechanism of action of TCDD.
Periportal hepatotoxicity due to allyl alcohol: A myriad of proposed mechanisms
Authors: Badr Mostafa Z.
Published: October 2006

It has long been recognized that periportal and centrilobular regions of the liver lobule differ in their sensitivity to many hepatotoxins. For example, carbon tetrachloride damages centrilobular regions while allyl alcohol damages periportal areas of the liver lobule (1). One determinant of the selective susceptibility of different areas of the liver lobule to chemically induced injury is believed to be the sublobular distribution of enzymes involved in detoxification, or activation to more toxic metabolites (2–4). In addition to sublobular enzyme activity, substrate delivery and the availability of necessary cofactors can influence the zone‐specific toxicity of some chemicals. Furthermore, it has been proposed that oxygen gradient across the liver lobule is an important determinant of zonal hepatotoxicity. Thus, studies regarding the zone‐specific nature of hepatotoxicants require that a variety of complex interrelated metabolic processes be considered, and so far exact mechanisms responsible for zone‐selective hepatotoxicity are not completely understood.
Interaction of benzanthrone with cytochrome p450: Altered patterns of hepatic xenobiotic metabolism in rats
Authors: Das Mukul , Garg Kalpana , Joshi Anjulika , Singh Giriraj B. , Khanna Subhash K.
Published: October 2006

Benzanthrone, an anthraquinone dye intermediate, is commonly used for the synthesis of a number of polycyclic vat and disperse dyes. Our prior studies have shown that benzanthrone can be metabolized by rat hepatic microsomal cytochrome P450 (P450) (Biochem. Int., 18, 1989, 1237). In this study, the interaction of benzanthrone with rat hepatic microsomal P‐450 and its effect on xenobiotic metabolism have been investigated. Parenteral administration of benzanthrone (40 mg/kg body weight) for 3, 7, or 21 days caused no change in the relative body weight or organ weight of rats. The levels of P450 were found to be reduced (33%‐50%) in all the benzanthrone‐exposed animals at all the time periods. In vitro addition of benzanthrone caused a spectral change with oxidized P450 and concentration‐dependent reduction in the carbon monoxide spectrum of dithionite‐reduced P450. The addition of benzanthrone to hepatic microsomes prepared from phenobarbital‐treated rats resulted in spectral changes characterized by an absorbance maximum at 397 nm indicative of type I binding. In vitro addition of benzanthrone showed a concentration‐dependent inhibition of hepatic aminopyrine N‐demethylase (APD) and ethoxyresorufin‐O‐deethylase (ERD) activities with respective I50 values of 9.5 × 10−4 and 8.0 × 10−5 M. However, the inhibition of aryl hydrocarbon hydroxylase (AHH) even at the highest concentration of benzanthrone (10−2 M), was of the order of only 29%. In vivo administration of benzanthrone also led to the inhibition of APD, AHH, and ERD activities at all treatment times although the magnitude of inhibition was of a lower order. Benzanthrone treatments caused significant depletion of glutathione content with a concomitant enhancement of lipid peroxidation. Also, benzanthrone resulted in a significant inhibition of the cytosolic enzyme, GSH‐transferase (23%–34%), although the other enzyme, quinone reductase, which helps in the removal of toxic quinones, was found to be elevated (184%–199%). These results suggest several mechanisms by which benzanthrone may inhibit P450. This impairment of xenobiotic metabolism as well as the enhancement of lipid peroxidation may be relevant to the observed symptoms of benzanthrone‐induced hepatotoxicity.
Effects of diethyldithiocarbamate (ddtc) on the plasma biotransformations of tetrachloro(d,i‐trans)‐1,2‐diaminocyclohexaneplatinum(iv) (tetraplatin) in fischer 344 rats
Authors: Carfagna Patrick F. , Wyrick Steven D. , Holbrook David J. , Chaney Stephen G.
Published: October 2006

We have studied the effects of diethyldithiocarbamate (DDTC) on the biotransformations of toxic doses of tetrachloro (d,l‐trans)1,2‐diaminocyclohexaneplatinum(IV) (tetraplatin) in Fischer 344 rats. In animals not treated with DDTC, tetraplatin was rapidly converted to dichloro(d,I‐trans)1,2‐diaminocyclohexaneplatinum(II) [PtCl2(dach]. Subsequent biotransformations included the transient formation of the (d,I‐trans)1,2‐diaminocyclohexane‐aquachloroplatinum(II) [Pt(H2O)(Cl)(dach)]+ complex, followed by formation of the platinum (Pt)‐methionine and either Pt‐cysteine or Pt‐ornithine complexes. Significant amounts of free (d,I‐trans) 1,2‐diaminocyclohexane (dach) were observed in plasma as a result of intracellular trans‐labilization reactions. DDTC caused a marked decrease in both total and protein‐bound platinum in the circulation. A significant increase in the plasma concentration of free dach was also observed as a result of formation of the Pt(DDTC)2 complex. Some of the free dach could have arisen from intracellular reactions with DDTC, but the displacement of platinum from plasma proteins was more than sufficient to account for the increase in free dach in the circulation. DDTC treatment also decreased plasma concentrations of tetraplatin, PtCl2(dach), [Pt(H2O)(Cl)(dach)]+, the Pt‐methionine complex, and one unidentified biotransformation product, but had no effect on the Pt‐cysteine (or Pt‐ornithine) complex. These effects of DDTC on protein‐bound platinum and low‐molecular‐weight biotransformation products in plasma may contribute to the decrease in tetraplatin toxicity seen in DDTC‐treated rats.
Evidence for stimulated glutathione synthesis by phenobarbital pretreatment during an oxidative challenge in isolated hepatocytes
Authors: Utley William S. , Mehendale Harihara M.
Published: October 2006

Hepatocytes isolated from phenobarbital‐pretreated and naive male Sprague‐Dawley rats were preincubated with 80 μM N, N‐bis (2‐chloroethyl)‐N‐nitrosourea and subsequently exposed to varying concentrations of menadione. We observed that the reduced glutathione levels of the hepatocytes isolated from the sodium phenobarbital(PB)‐pretreated, but not the naive rats, recovered to near‐control levels after exposure to 200 μM menadione. Since this recovery occurred in the presence of N, N‐bis (2‐chloroethyl)‐N‐nitrosourea (an inhibitor of glutathione reductase), we hypothesized that this represented a PB‐mediated increase in de novo synthesis of glutathione. To test this hypothesis and to further assess the possible contribution of glutathione reductase in the recovery of the glutathione levels, we preincubated hepatocytes isolated from PB‐pretreated and naive rats with 2 mM buthionine sulfoximine, with or without N, N‐bis (2‐chloroethyl)‐N‐nitrosourea. Following exposure to menadione, samples were periodically removed for glutathione assessment. Consistent with our hypothesis, the addition of buthionine sulfoximine abrogated the ability of the PB‐pretreated hepatocytes to restore glutathione levels following a menadione challenge. Buthionine sulfoximine in combination with N, N‐bis (2‐chloroethyl)‐N‐nitrosourea completely abolished hepatocellular glutathione homeostasis for all of the concentrations of menadione employed. The findings from this investigation underscore the importance of phenobarbital‐mediated increases in glutathione synthesis, as well as the enhanced levels of glutathione reductase, in maintaining the pool of reduced glutathione and ultimately mitigating the consequences of oxidative stress. In addition, these findings suggest that PB pretreatment increases the reserve capacity of the hepatocyte for glutathione synthesis via a hitherto undescribed hormetic mechanism, a reserve expressed fully only on an oxidative stress of sufficient magnitude.
Glutathione S‐transferase‐catalyzed conjugation of 9,10‐epoxystearic acid with glutathione
Authors: Sharma Rajendra , Gupta Sanjiv , Singhal Sharad S. , Ansari G. A. S. , Awasthi Yogesh C.
Published: October 2006

The possible role of glutathione S‐transferases (GST) in detoxification of fatty acid epoxides generated during lipid peroxidation has been evaluated. Present studies showed that cytosolic human glutathione S‐transferases belonging to α, H, and μ classes isolated from human liver and lung catalyzed the conjugation of glutathione and 9,10‐epoxystearic acid. The product of enzymatic reaction, i.e., conjugate of GSH and epoxystearic acid, was isolated and characterized. The Michaelis constant (Km) values of the α, μ, and π classes of GSTs for 9,10‐epoxystearic acid were found to be 0.47, 0.32 and 0.80 mM, respectively, whereas the maximal velocity (V max) values for the α, μ, and π classes of GSTs were found to be 142, 256, and 52 mol/min/mol, respectively. These results indicate that even though 9,10‐epoxystearic acid is a substrate for all the three classes of GSTs, the μ class isozymes have maximum activity toward this substrate and may preferentially metabolize fatty acid epoxides more effectively as compared to the other classes of GSTs.
Purified NAD(P)H‐quinone oxidoreductase enhances the mutagenicity of dinitropyrenes in vitro
Authors: Hajos Antal K. D. , Winston Gary W.
Published: October 2006

The effect of highly purified rat liver cytosolic NAD(P)H‐quinone oxidoreductase [EC 1.6.99.2] on the mutagenicity of 1,3‐ 1,6‐ and 1,8‐dinitropyrene (DNP) was studied in the Ames Salmonella typhimurium mutagenicity assay. NAD(P)H‐quinone oxidoreductase over the range of 0.02–0.8 μ g/plate (38–1500) units increased up to threefold the mutagenicity of all three DNPs in S. typhimurium TA 98. In TA98NR, a strain deficient in “classical” nitroreductase, the mutagenicity of 1,6‐ and 1,8‐DNP was essentially unchanged, whereas that of 1,3‐DNP was markedly reduced. NAD(P)H‐quinone oxidoreductase enhanced the mutagenicity of 1,6‐ and 1,8‐DNP to approximately equivalent extents in TA98NR and TA98. The mutagenicity of 1,3‐DNP in TA98NR was potently enhanced by the addition of NAD(P)H‐quinone oxidoreductase in a dose‐responsive manner. In the presence of 0.8 μg NAD(P)H‐quinone oxidoreductase, 1,3‐DNP displayed a mutagenic response in TA98NR that was comparable to that obtained in TA98. NAD(P)H‐quinone oxidoreductase was found to increase the mutagenicity of 1,6‐ but not 1,3‐ or 1,8‐DNP to mutagenic intermediates in TA98/1,8‐DNP6, a strain deficient in O‐acetyltransferase activity. The results suggest that NAD(P)H‐quinone oxidoreductase not only catalyzes reduction of the parent DNP but also that of partially reduced metabolites generated from that DNP. Such reductive metabolism may lead to increased formation of the penultimate mutagenic species.
Ovarian toxicity of hexachlorobenzene (HCB) in the superovulated female rat
Authors: Foster Warren G. , Pentick Julie A. , McMahon Avril , Lecavalier Pierre R.
Published: October 2006

Hexachlorobenzene (HCB) is a persistent environmental contaminant which has been measured in human serum, fat, semen, and follicular fluid. In animal testing HCB has been shown to be a reproductive toxin. Discrepant results were obtained from prior studies concerning the effect of HCB treatment on ovarian steroidogenesis. The current study was designed to assess the impact of HCB on the ovary and gonadal steroid levels in the superovulated rat. Female Sprague‐Dawley rats (n = 24) were dosed with HCB (0.0, 1.0, 10.0, or 100.0 mg/kg BW/day) for 21 days. All rats received 10 IU pregnant mare serum gonadotropin (PMSG) s.c. on day 18 of treatment and 15 IU of human chorionic gonadotropin (hCG) on day 20. A terminal blood sample was collected and circulating levels of estradiol (Emr2) and progesterone. (P4) were determined. Serum concentrations of P4 were significantly (p < 0.0034) elevated by HCB treatment at all dose levels. Ovarian weights were significantly increased (p < 0.05) in the lowest dose group only compared to the control group. Serum concentrations of Ev uterine weight, weight gain, and general animal health were not affected by HCB treatment. We conclude that during HCB treatment the rat ovary remains responsive to gonadotropin stimulation. Moreover, it is suggested that HCB effects on ovarian steroidogenesis are indirect.
Oxidant and angiotensin II‐induced subcellular translocation of protein kinase C in pulmonary artery endothelial cells
Authors: Patel Jawaharalal M. , Sekharam K. Madhavi , Block Edward R.
Published: October 2006

We recently reported that nitrogen dioxide (NO2), an environmental oxidant, alters the dynamics of the plasma membrane lipid bilayer structure, resulting in increased phosphatidylserine content and angiotensin II (Ang II) receptor binding. Angiotensin II is known to elicit receptor‐mediated stimulation of diacylglycerol (DAG) production in pulmonary artery endothelial cells. Because protein kinase C (PKC) is a phosphatidylserine‐dependent enzyme and is activated by DAG, we examined whether NO2 resulted in activation and/or translocation of PKC from predominantly cytosolic to membrane fractions of these cells. We also evaluated whether NO2 exposure resulted in increased production of DAG in pulmonary artery endothelial cells. Exposure to 5 ppm NO2 for 1–24 hr resulted in significant increases in PKC activity in the cytosolic and membrane fractions (p < 0.05 for both fractions) compared to activities in control fractions. Exposure to Ang II resulted in translocation of PKC activity from cytosol to membrane fractions of both control and NO2‐exposed cells. This translocation of PKC from cytosolic to membrane fraction was prevented by the specific receptor antagonist [Sar1 Ile8] Ang II. Exposure of 5 ppm NO2 for 1–24 hr provoked rapid increases in [3H]glycerol labeling of DAG in pulmonary artery endothelial cells. These results demonstrate that exposure to NO2 increases the production of second messenger DAG and activates PKC in both the cytosolic and membrane fractions, whereas Ang II stimulates the redistribution of PKC from cytosolic to membrane fractions of pulmonary artery endothelial cells.
Action of some retinol derivatives and their provitamins on microsome‐catalyzed formation of benzo[a]pyrene‐DNA adduct
Authors: Shah G. M. , Goswami U. C. , Bhattacharya R. K.
Published: October 2006

Several vitamin A compounds have been tested for their ability to suppress formation of DNA adduct by the carcinogen benzo[a]pyrene (B[a]P) in an in vitro reaction catalyzed by rat liver microsomes. Retinol, retinal, 3‐dehydroretinol and 3‐hydroxyretinol were found to be effective inhibitors of adduct formation. Certain carotenoids that are precursors of these retinoids also displayed considerable inhibitory capacity. Carotenoids and the 3‐substituted retinoids appeared to modulate the DNA adduct formation exclusively through their action on microsomal enzymes, since an effective inhibition in each case was observed on the formation of B[a]P‐7,8‐diol, a proximate carcinogenic metabolite of B[a]P. Unsubstituted retinoids, on the other hand, had marginal effect on enzymes but were found effective in accelerating inactivation of B[a]P‐7,8‐diol‐9,10‐epoxide, the ultimate carcinogenic metabolite that binds to DNA.
Effects of niacin on bleomycin‐induced increases in myeloperoxidase, prolyl hydroxylase, and superoxide dismutase activities and collagen accumulation in the lungs of hamsters
Authors: O'Neill Charles A. , Giri Shri N.
Published: October 2006

It has been shown that lung nicotinamide adenine dinucleotide (NAD) depletion accompanies bleomycin (BL)‐induced lung fibrosis in the hamster and that treatment with niacin (NA), a precursor of NAD, was found to attenuate lung fibrosis caused by this agent. Niacin was used in the present study to investigate changes in some biochemical parameters and enzymes involved in the development of BL‐induced lung fibrosis in the hamster. Niacin (500 mg/kg, IP), or an equivalent volume of saline (SA, IP), was given daily 2 days prior to intratracheal instillation of BL (7.5 U/5 mL/ kg) or SA and everyday thereafter throughout the study. Hamsters were killed at 1, 4, 7, 10, and 14 days after the BL or SA instillation and their lungs processed for various biochemical assays. Hydroxyproline content and superoxide dismutase (SOD) activity in SABL treated animals were significantly (P ± 0.05) elevated at 7 and 10 days, peaking at 14 days to 161 ± 11% and 159 ± 11% of the SASA treated animals, respectively. Although the hydroxyproline level of NABL treated animals was significantly elevated at 7 and 10 days and peaked at 14 days to 123 ± 8% of the NASA control, these values were significantly lower than the SABL treated animals at the corresponding times. The lung SOD activity of NABL groups was significantly higher at 4 days but significantly lower at 10 and 14 days than the SABL groups at the corresponding times. Prolyl hydroxylase (PH) activity and total lung calcium in SABL treated groups were significantly elevated compared to SASA treated groups starting at 4 days, with PH peaking at 10 days to 163 ± 13% and calcium peaking at 7 days to 148 ± 8% of SASA treated groups. The NABL treated animals displayed a significant elevation in PH activity at 4 days only (132 ± 15%), while the calcium content in this group was significantly increased at 4 and 14 days compared to NASA treated animals. However, the activity of PH in the NABL treated animals was significantly lower than the SABL treated animals at 7, 10, and 14 days. The calcium content of the NABL group was significantly lower than the SABL group at 7 and 10 days. The thiobarbituric acid reactive substance equivalents (TBARS) content and myeloperoxidase (MPO) activity were significantly elevated at all time points in SABL groups as compared to SASA groups, with peak elevation of TBARS to 160 ± 9% at 4 days and MPO to 268 ± 40% at 1 day. The TBARS content of NABL groups was significantly elevated above NASA groups at 1, 4, 7, and 10 days. The MPO activity in NABL groups was significantly elevated above NASA groups at 1, 4, 7, and 14 days, but the activity was significantly lower than SABL groups at 4 and 14 days. The data suggest that NA inhibits the extent of oxidative damage or enhances processes involved in the repair of BL‐induced alveolar epithelial damage.
Bioactivation of 7‐hydroxymethyl‐12‐ methyibenz[a]anthracene by rat liver bile acid sulfotransferase I
Authors: Falany Charles N. , Wheeler James , Coward Lori , Keehan Dawn , Falany Josie L. , Barnes Stephen
Published: October 2006

The bioactivation of 7‐hydroxy‐methyl‐12‐methylbenz[a]anthracene (HMBA) to an electrophilic sulfuric acid ester metabolite has been shown to be catalyzed by rat liver bile acid sulfotransferase I (BAST I). The sulfation and activation of HMBA by BAST I was determined by the ability of sulfated HMBA to form DNA ad‐ducts. The BAST I was also shown to react with rabbit anti‐human dehydroepiandrosterone sulfotransferase antisera and to represent a major form of hydroxysteroid/bile acid sulfotransferase in female rat liver cytosol. Higher levels of BAST I activity and immunoreactivity as well as HMBA‐DNA adduct formation were detected in female rat liver cytosol than in male rat liver cytosol. The bioactivation of HMBA by pure BAST I was dependent on the presence of 3′‐phosphoadenosine 5′‐phos‐phosulfate (PAPS) in the reaction and was inhibited by dehydroepiandrosterone, a physiological substrate for BAST I. Glutathione, a cellular nucleophile with important protective properties, decreased DNA adduct formation in the HMBA sulfation reaction in the absence of glutathione S‐transferase activity. These results indicate the usefulness of BAST I to investigate the sulfation and activation of HMBA and probably other hydroxy‐methylated polyaromatic hydrocarbons to electrophilic and mutagenic metabolites under defined reaction conditions.
Heterogeneity of rat hepatic Ah Receptor: Identification of two receptor forms which differ in their biochemical properties
Authors: Denison Michael S.
Published: October 2006

In cytosol, the rat hepatic Ah receptor (AhR) appears to exist in two distinct forms (AhRα, AhRβ) in similar concentration. The binding of ligand (2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD)) to AhRα requires the receptor be in its oligomeric 8–10 to S conformation (bound to other protein subunits), while ligand binding to AhRβ can occur with the dissociated 5–6 S form. Occupancy of AhRβ by ligand (TCDD) protects it from salt‐dependent inactivation; AhRβ is not inactivated by high salt conditions. The addition of molybdate to cytosol during tissue homogenization stabilized AhRα against salt‐dependent inactivation and subunit dissociation but did not prevent dissociation of AhRβ by high salt. Although the presence of molybdate appears to stabilize AhRα in its oligomeric 8–10 S, it had no significant effect on the overall amount of TCDD:AhR complex which bound to its specific DNA recognition site, the dioxin responsive element (DRE). These results suggest that AhRα, unlike AhRβ, is either unable to transform or bind to the DRE with high affinity.
The differential hepatotoxicity and cytochrome P450 responses of fischer‐344 rats to the three isomers of dichlorobenzene
Authors: Allis John W. , Simmons Jane Ellen , House Dennis E. , Robinson Barbara L. , Berman Ezra
Published: October 2006

The acute hepatotoxicity and response of hepatic cytochrome P450 to treatment with the three isomers of dichlorobenzene (DCB) have been investigated. The objectives were to estimate the onset of toxicity and to further elucidate the role of cytochrome P450 in the metabolism and toxicity of these compounds. In a study design employing one animal per dose level, Fischer‐344 rats were gavaged with up to 25 different dosages, then evaluated 24 h later. Hepatic necrosis, serum alanine aminotransferase, and serum aspartate aminotransferase exhibited similar patterns demonstrating that ortho‐DCB (O‐DCB) was the most toxic in terms of both earliest onset and degree of response at higher dosages. For these three end‐points, meta‐DCB (m‐DCB) exhibited a lesser toxicity. Para‐DCB (p‐DCB) did not cause changes in these three endpoints, but hepatic degenerative changes were found. Total hepatic cytochrome P450 responses were also different after treatment with each isomer. The o‐DCB produced a dose‐dependent decrease in P450 beginning at dosages lower than the onset of necrosis and appeared to be a suicide substrate for P450. The m‐DCB treatment increased P450 at dosages below the onset of necrosis and decreased P450 at higher dosages, with the decline preceding the onset of hepatocyte death. Treatment with p‐DCB increased P450 beginning at 380 mg/kg. The combination of toxicity and P450 profiles has provided a framework for interpreting literature data on the metabolism and toxicity of the DCBs in rats. It is also noteworthy that o‐DCB and p‐DCB were administered at dosages several times the oral rat LD‐50 (RTECS) without any lethality.
Interactions in vitro of some organophosphoramidates with neuropathy target esterase and acetylcholinesterase of hen brain
Authors: Jokanovic M. , Johnson M. K.
Published: October 2006

For organophosphates or phosphorates to initiate delayed neuropathy two steps are necessary: (1) progressive covalent reaction with neuropathy target esterase (NTE) to produce a form of inhibited NTE which can be reactivated by incubation with aqueous potassium fluoride (KF) and (2) progressive “aging” of inhibited NTE to a form which can no longer be reactivated by KF. However, it has been shown recently that certain N‐unsubstituted organophosphoro‐monoamidates (analogues of methamidophos) cause delayed neuropathy even though the inhibited NTE appeared not to have aged (Johnson et al. (1991). Arch. Tox‐icol., 65, 618–624). In order to study the generality of this phenomenon, we have examined some N‐substituted compounds. We report In vitro studies of inhibition and reactivation and aging of both NTE and acetylcholinesterase (AChE) prior to tox‐icological tests. All the compounds studied were less inhibitory to both NTE and AChE in concentrated rather than in dilute suspensions of EDTA‐washed brain particles without added cofactors. There was an apparent disposal of up to 100 üumoles of test compound by particles from 95 mg hen brain, which is far greater than can be explained by covalent binding. The activity is distinct from calcium‐dependent “A” esterase. Several N‐alkyl phosphoromonoamidates were found to be potent and selective inhibitors of NTE: second‐order rate constant for O‐n‐pentyl N‐benzylphosphoramido‐fluoridate (Cmpd 6) = 5.6×107 M−1 min−1 at 37°, which is about l00% higher than for acetylcholinesterase (AChE). Inhibited NTE and AChE from several chiral phosphoromono‐amidates did not reactivate spontaneously (21 hours at 37°). Virtually 100% reactivation by KF of AChE inhibited by phosphoromonoamidates was achieved at all times tested. Acetylcholinesterase inhibited by 2,5‐dichlorophenyl N,N′‐di‐n‐butylphosphorodiamidate was 42–56% reactivated by incubation with KF (192 mM in pH 5.2 buffer for 30 minutes at 37°rpar;. We believe this is the first report of reactivation of any enzyme after inhibition by a phosphorodi'amidate. For NTE inhibited by tabun (O‐ethyl N‐dimethylphosphoroamidocyanidate), virtually complete and rapid aging (t1/2 = 5.5–8.4 minutes) was observed. Consistent but only partial reactivation by KF was achieved 2 or more hours after inhibition of NTE by Cmpd 6 or by its 2,6‐difluoro‐an‐alogue (Cmpd 7). However, a small but significant aging (approximately 15–20% loss of reactivatability) was measured soon after a 1 minute inhibition by Cmpd 7, but no further change occurred in 21 hours. We conclude that, of the two chiral forms of NTE obtainable from chiral N‐substituted phosphoromonoamidates, one form can and does age rapidly while the enantiomeric form does not. The toxicological significance of these results is discussed.
Immobilized rhodanese: Some aspects of anion inhibition kinetics and modulation by cations
Authors: Nok Andrew J. , Nasir Shuaibu M. , Sa'Adatu Yusuf
Published: October 2006

The rat liver rhodanese (thiosulphate: cyanide sulfurtransferase EC 2.6.1.1) has been immobilized on polyacrylamide gels. The immobilized enzyme had a pH optimum of 7.4 and Km values of 3.25 mM and 1.12 mM for S2O2−3 and KCN, respectively. The enzyme was competitively inhibited by NaNO2 and CH3COONa and noncompetitively by amyl‐nitrite. A modulation of activity was observed in the presence of Ca2+, Zn2+, and Cu2+. The results are discussed in line with the detoxicating function of liver rhodanese.