Styrene Inhalation Toxicity Studies in Mice: III. Strain Differences in Susceptibility

Inhalation toxicity studies were conducted to evaluate mousestrain differences in the susceptibility to styrene vapors.Male and female B6C3F1, C57BL/6, Swiss, and DBA/2 mice (8 weeksold) were exposed to 0, 125, 250, or 500 ppm styrene 6 hr/day,for 4 days (20/sex/dose). Histopathological changes and changesin liver weights were evaluated as a measure of hepatotoxicity.Styrene uptake and styrene-7,8-oxide (SO) formation were estimatedby measuring levels of styrene and SO in blood. An estimateof SO detoxification by conjugation with GSH was obtained bymeasuring hepatic GSH depletion. In general, mortality, increasedliver weights, and hepatocellular necrosis were observed inthe 250 and 500 ppm dose groups for all strains and both sexes.Considerable sex and strain differences were observed. Mortality,increased liver weights, and hepatocellular necrosis were greatestin B6C3F1 and C57BL/6 mice in the 250 ppm dose group and inmales; hepatotoxicity was similar in both strains. Swiss miceexhibited dose-dependent increases in mortality, liver weights,and in hepatocellular necrosis, with only slight sex differencesat early time points. Hepatotoxicity in DBA/2, B6C3F1, and C57BL/6strains was greater at 250 than 500 ppm; however, toxicity wasless severe in DBA/2 than in other strains based on absenceof mortality in either sex and less extensive liver necrosisat both 250 and 500 ppm. Blood styrene and SO levels did notcorrelate well with strain differences in toxicity. The relativetoxicity (mortality and hepatotoxicity) was B6C3FI>Swiss>DBA/2;however, relative blood styrene and SO levels were B6C3F1DBA/2>Swiss.Hepatic GSH depletion, (B6C3F1DBA/2>Swiss) correlated withblood SO levels as expected. These results demonstrate significantbiologic variability in the susceptibility of mouse strainsto styrene toxicity. These differences are presumably due tostrain and sex differences in metabolism; however, toxicitydid not correlate well with blood SO levels, suggesting thatother reactive metabolites may contribute to styrene toxicityin mice.     

Styrene Inhalation Toxicity Studies in Mice: II. Sex Differences in Susceptibility of B6C3F1 Mice

Styrene is a commercially important chemical used in the productionof plastics and resins. In initial short-term styrene inhalationstudies, toxicity was significantly greater in male B6C3F1 micethan in females, suggesting that males may metabolize styrenemore extensively and/or may be less able to detoxify reactivemetabolites. In addition, a nonlinear dose-response was observedwhere toxicity and mortality were greater in mice exposed to250 ppm than in those exposed to 500 ppm. These studies wereconducted to investigate potential mechanism(s) for sex differencesand the nonlinear dose-response in styrene toxicity by evaluatingthe effects of repeated styrene exposure on styrene oxide production,hepatic GSH availability, and hepatotoxicity in male and femaleB6C3F1 mice. Mice (36/sex/dose) were exposed to 0, 125, 250,or 500 ppm styrene 6 hr/day for up to 3 days. Styrene exposurecaused increased mortality and hepatotoxicity (centrilobularnecrosis, increased serum liver enzymes) in males and femalesafter one or two exposures to 250 and 500 ppm. Hepatic GSH levelswere decreased in a dose-dependent manner in males and females.After one exposure, GSH levels in males rebounded above controlsin all dose groups. After three exposures to 125 or 250 ppmmales appeared to maintain GSH levels; GSH was still decreasedin the 500 ppm group. GSH levels in females were decreased aftereach exposure in all dose groups to lower levels than in males,and did not rebound above controls. Male mice had significantlygreater blood styrene levels than females after one exposureto 500 ppm; however, there were no significant sex differencesin blood styrene after subsequent exposures. Levels of SO inblood were not significantly greater in male mice than femaleswithin a dose group, and did not change significantly with repeatedstyrene exposures for 3 days. Blood styrene and SO levels weresignificantly higher at 500 ppm than at 250 ppm indicating thatstyrene uptake and metabolism are greater at 500 ppm than at250 ppm. The higher incidence of mortality in male mice andthe nonlinear dose-response to styrene cannot be explained bygender- or dose-related differences in hepatotoxicity, GSH depletion,or blood styrene or SO levels.     

Toxicity of Divinylbenzene-55 for B6C3F1 Mice in a Two-Week Inhalation Study

Divinylbenzene (DVB) is a crosslinking monomer used primarilyfor copolymerization with styrene to produce ion-exchange resins.The toxicity of inhaled DVB was investigated because of thepotential for worker exposure and the structural similarityof DVB to styrene, a potential carcinogen. Male and female B6C3F1mice were exposed to 0, 25, 50, or 75 ppm DVB for 6 hr/day,5 days/week for up to 2 weeks. Six mice/sex/dose group werekilled after 3, 5, and 10 exposures and six mice/sex in the75 ppm group were killed 7 days after 10 exposures. The mostsevere effects occurred in the nasal cavity and liver, withless severe effects occurring in the kidneys. In the nasal cavityolfactory epithelium acute necrosis and inflammation were presentat early time points followed by regeneration, architecturalreorganization, and focal respiratory metaplasia by 7 days afterthe last exposure. Olfactory epithelial changes were concentration-dependentwith extensive involvement at 75 ppm and peripheral sparingat 25 ppm. There was also necrosis and regeneration of olfactory-associatedBowman's glands as well as the lateral nasal (Steno's) glands.Hepatocellular centrilobular (CL) necrosis was observed onlyin the 75 ppm dose group and was similar to that caused by styrene.A time-dependent progression was observed, characterized byCL degeneration after 1 exposure, necrosis after 3 and 5 exposures,and chronic inflammation with CL karyomegaly after 10 exposuresand 7 days after the 10th exposure. Hepatic GSH levels weredecreased in a dose-dependent manner. In the kidneys, transienttubular damage was observed in some male mice exposed to 75ppm, and appeared to be a response to DVB-induced tubular epithelialinjury.     

Characterization of inhaled alpha-methylstyrene vapor toxicity for B6C3F1 mice and F344 rats.

alpha-Methylstyrene (AMS) is a chemical intermediate used in the synthesis of specialty polymers and copolymers. Inhalation studies of AMS were conducted because of the lack of toxicity data and the structural similarity of AMS to styrene, a toxic and potentially carcinogenic chemical. Male and female B6C3F1 mice were exposed to 0, 600, 800, or 1000 ppm AMS 6 h/day, 5 days/week, for 12 days. After 1 exposure, 21% (5/24) of female mice were found dead in the 1000-ppm group, 56% (10/18) in the 800-ppm group, and 6% (1/18) in the 600-ppm concentration group. After 12 exposures, relative liver weights were significantly increased and relative spleen weights were significantly decreased in both male and female mice at all concentrations. No microscopic treatment-related lesions were observed. A decrease in hepatic glutathione (GSH) was associated with AMS exposure for 1 and 5 days. Male and female F344 rats were exposed to 0, 600 or 1000 ppm AMS for 12 days. No mortality or sedation occurred in AMS-exposed rats. Relative liver weights were significantly increased in both males and females after 12 exposures to 600 or 1000 ppm. An increased hyaline droplet accumulation was detected in male rats in both concentration groups; no significant microscopic lesions were observed in other tissues examined. Exposure of male and female F344 rats and male NBR rats to 0, 125, 250 or 500 ppm AMS, 6 h/day for 9 days resulted in increased accumulation of hyaline droplets in the renal tubules of male F344 rats in the 250 and 500 ppm concentration groups. Although AMS and styrene are structurally very similar, AMS was considerably less toxic for mice and more toxic for male rats than styrene.     

Glutathione depletion by methyl chloride and association with lipid peroxidation in mice and rats

Inhalation of methyl chloride (CH3Cl) by male B6C3F1 mice resulted in a concentration-dependent depletion of glutathione (GSH) in liver, kidney, and brain. Exposure for 6 hr to 100 ppm CH3Cl decreased the concentration of GSH in mouse liver by 45%, while exposure to 2500 ppm for 6 hr lowered liver GSH to approximately 2% of control levels. For those exposures which decreased liver GSH to less than 20% of control levels, the extent of liver GSH depletion was closely correlated with the capacity of a 9000g supernatant fraction from the liver to undergo lipid peroxidation in vitro. GSH was depleted to a lesser extent in mouse brain and kidney, compared to liver, and no relationship to peroxidation was observed for single exposures to CH3Cl. A dose-dependent decrease in liver GSH was also produced by diethyl maleate, although a nearly lethal amount (2 ml/kg) was required to lower liver GSH to less than 10% of control levels. Under these conditions the amount of lipid peroxidation was 3.5-fold less than in mice exposed to 2000 ppm CH3Cl. Exposure of rats to 2000 ppm CH3Cl reduced liver GSH to 20% of control levels, compared to 4.5% in mice similarly exposed, and under these exposure conditions the amount of lipid peroxidation measured in vitro was 40-fold greater in mouse liver than in rat liver. During exposure of mice to 2500 ppm CH3Cl, ethane expiration increased to an extent comparable to that produced by administration of 2 ml/kg of CCl4. These findings suggest that GSH depletion in liver may be an important component of CH3Cl-induced hepatotoxicity.     

Inhalation pharmacokinetics of ethylbenzene in B6C3F1 mice

The objective of the present study was to characterize the inhalation pharmacokinetics of ethylbenzene (EB) in male and female B6C3F1 mice following single and repeated exposures. Initially, groups of 28 male and female mice were exposed for 4 h to 75, 200, 500, or 1000 ppm in order to determine potential non-linearity in the kinetics of EB. Then, groups of male and female mice were exposed for 6 h to 75 ppm and 750 ppm (corresponding to the NTP exposures) for 1 or 7 consecutive days, to evaluate whether EB kinetics was altered during repeated exposures, The maximal blood concentration (Cmax; mean+/-SD, n=4) observed in female mice at the end of a 4-h exposure to 75, 200, 500, and 1000 ppm was 0.53+/-0.18, 2.26+/-0.38, 19.17+/-2.74, and 82.36+/-16.66 mg/L, respectively. The areas under the concentration vs. time curve (AUCs) following 4-h exposure to 75, 200, 500, and 1000 ppm were 88.5, 414.0, 3612.2, and 19,104.1 mg/L/min, respectively, in female mice, and 116.7, 425.7, 3148.3, and 16,039.1 mg/L/min in male mice. The comparison of Cmax and the kinetic profile of EB in mice exposed to 75 ppm suggests that they are similar between 1-day and 7-day exposures. However, at 750 ppm, the rate of EB elimination would appear to be greater after repeated exposures than single exposure, the pattern being evident in both male and female mice. Overall, the single and repeated exposure pharmacokinetic data collected in the present study suggest that EB kinetics is saturable at exposure concentrations exceeding 500 ppm (and therefore at 750 ppm used in the NTP mouse cancer bioassay) but is in the linear range at the lower concentration used in the bioassay (75 ppm). These data suggest that consideration of the nature and magnitude of non-linear kinetics and induction of metabolism during repeated exposures is essential for the conduct of a scientifically sound analysis of EB cancer dose-response data collected in B6C3F1 mice.     

Styrene Inhalation Toxicity Studies in Mice: I. Hepatotoxicity in B6C3F1 Mice

Studies were conducted to evaluate the toxic effects of short-termrepeated styrene inhalation in B6C3F1 mice. Male and femalemice were exposed to 0, 125, 250, or 500 ppm styrene, 6 hr/day,for up to 14 days. Styrene toxicity was characterized by severecentrilobular hepatic necrosis and deaths after one exposureto 500 ppm or two exposures to 250 ppm. Mortality and hepatotoxicitywere not increased by additional exposures, and in survivingmice, regeneration and repair of initial hepatic injury occurredin spite of continued exposure for 14 days. A marked sex differencewas observed, with male mice significantly more susceptibleto styrene toxicity than females. A nonlinear dose responsewas observed where mortality in male and female mice was greaterin the 250 ppm dose group than that in the 500 ppm dose group.Severe congestion and necrosis of the liver was present in moribundmice; hepatic congestion and serum alanine aminotransferaseand sorbitol dehydrogenase were significantly greater in moribundanimals.     

Subchronic Inhalation Studies of Styrene in CD Rats and CD-1 Mice

Groups of 10 male and 10 female Charles River (CRL) CD (Sprague-Dawley-derived)rats were exposed to styrene vapor at 0, 200, 500, 1000, or1500 ppm 6 hr per day 5 days per week for 13 weeks. Styrenehad no effect on survival, hematology, or clinical chemistry.Males at 1500 ppm weighed 10% less after 13 weeks and malesand females at 1000 and 1500 ppm consumed more water than controls.Histopathologic changes were confined to the olfactory epitheliumof the nasal mucosa. Groups of 20 male and 20 female CRL CD-1and B6C3F1 mice were exposed to styrene vapor at 0, 15, 60,250, or 500 ppm 6 hr per day 5 days per week for 2 weeks. Mortalitywas observed in both CD-1 and B6C3F1 mice exposed to 250 or500 ppm; more female mice, but not males, died from exposureto 250 ppm than from 500 ppm. Groups of 10 male and 10 femaleCRL CD-1 mice were exposed to styrene vapors at 0, 50, 100,150, or 200 ppm 6 hr per day 5 days per week for 13 weeks. Twofemales exposed to 200 ppm died during the first week. Livertoxicity was evident in the decedents and in some female survivorsat 200 ppm. Changes were observed in the lungs of mice exposedto 100, 150, or 200 ppm and in the nasal passages of all treatmentgroups, those exposed to 50 ppm being less affected. Satellitegroups of 15 male rats and 30 male mice were exposed as describedabove for 2, 5, or 13 weeks for measurement of cell proliferation(BrdU labeling). No increase in cell proliferation was foundin liver of rats or mice or in cells of the bronchiolar or alveolarregion of the lung of rats. No increase in labeling index oftype II pneumocytes was seen in mouse lungs, while at 150 and200 ppm, an increased labeling index of Clara cells was seenafter 2 weeks and in occasional mice after 5 weeks. Large variationsin the labeling index among animals emphasize the need for largegroup sizes. For nasal tract effects, a NOAEL was not foundin CD-1 mice, but in CD rats, the NOAEL was 200 ppm. For othereffects, the NOAEL was 500 ppm in rats and 50 ppm in mice.     

Inhibition of the acute toxicity of methyl chloride in male B6C3F1 mice by glutathione depletion

Previous data have demonstrated that methyl chloride (MeCl) is toxic to B6C3F1 mice under both acute and chronic exposure conditions, and that conjugation of MeCl with glutathione (GSH) is a key step in the metabolism of MeCl. This study examined the role of GSH in mediating the acute toxicity of MeCl to liver, kidney, and brain of male B6C3F1 mice. The lethal effects of a single 6-hr inhalation exposure of B6C3F1 males to 2500 ppm MeCl were completely prevented by pretreatment with the GSH synthesis inhibitor, L-buthionine-S,R-sulfoximine (4 mmol L-BSO/kg, ip 1.5 hr prior to MeCl exposure). GSH levels (measured as nonprotein sulfhydryl) in liver and kidney were depleted to 19 and 25% of control values, respectively, at the start of the exposure; the ratio of dead/exposed mice during the 18-hr postexposure declined from 14/15 mice to 0/10. Also, the LC50 for MeCl increased from 2200 to 3200 ppm in male mice pretreated with BSO. The hepatic toxicity of MeCl was detected by increased alanine aminotransferase (ALT) activities in serum 18 hr after a 6-hr exposure to 1500 ppm MeCl (2147 +/- 1327 IU/liter vs 46 +/- 6 in controls). Liver toxicity was inhibited when B6C3F1 males were depleted of GSH prior to MeCl exposure by BSO pretreatment (43 +/- 2), fasting (100 +/- 47), or injection of diethyl maleate (42 +/- 16). The effects of GSH depletion on MeCl toxicity to brain and kidney were determined in B6C3F1 males exposed to 1500 ppm MeCl 6 hr/day, 5 days/week for 2 weeks, with and without daily pretreatment with 2 mmol L-BSO/kg. This dose of BSO depleted hepatic and renal GSH by 28 and 60%, respectively, at the start of MeCl exposure. BSO-pretreated mice were protected from the central nervous system toxicity of MeCl, as assessed by microscopic examination of the granule cell layer of the cerebellum. BSO pretreatment also inhibited the renal toxicity of MeCl as measured by incorporation of [3H]thymidine ([3H]TdR) into renal DNA, an indicator of cell regeneration after cortical necrosis. [3H]TdR incorporation was 105 +/- 10,337 +/- 40, and 60 +/- 15 dpm/microgram DNA in nonexposed controls, MeCl, and MeCl + BSO treatment groups, respectively. These results indicate that GSH is an important component in the toxicity of MeCl to multiple organ systems in B6C3F1 mice. Reaction of MeCl with GSH appears to constitute a mechanism of toxication, contrary to the role usually proposed for GSH in detoxifying xenobiotics.     

Carcinogenicity and Toxicity of Inhaled Nitrobenzene in B6C3F1 Mice and F344 and CD Rats

The potential carcinogenicity and toxicity of inhaled nitrobenzenewere evaluated following chronic (2-year) exposure in mice andrats. Male and female B6C3F1 mice were exposed to 0, 5, 25,or 50 ppm nitrobenzene, while male and female F344 rats andmale CD rats were exposed to 0, 1, 5, or 25 ppm nitrobenzene.All exposures were for 6 hr/day, 5 days/week excluding holidays,for a total of 505 days over 2 years. Survival was not adverselyaffected by nitrobenzene exposure, and only mild exposure-relateddecreases in body weights (<10% of control) were occasionallynoted. Nitrobenaene exposure resulted in increased incidenceof neoplasia in male B6C3F1 mice (pulmonary alveolar/bronchiolarand thyroid follicular cell neoplasms), female B6C3FI mice (mammarygland neoplasms), male F344 rats (hepatocellular and renal neoplasms),female F344 rats (endometrial stromal neoplasms), and male CDrats (hepatocellu lar neoplasms). In addition, there were marginalincreases in the incidence of hepatocellular neoplasia in femaleB6C3F1 mice and thyroid follicular neoplasia in male F344 rats.Groups of nitrobenzene-exposed mice and rats with increasedincidence of renal and thyroid neoplasia also had increasedincidences of hyperplasia in these tissues. Toxicity resultingfrom chronic inhalation of nitrobenzene was manifested by methemoglobinemia,anemia, and adaptive or degenerative changes in the nose, liver,and testis. The results indicate that inhaled nitrobenzene iscarcinogenic and toxic in mice and rats, and that the spectrumof these responses in animals is dependent on species, sex,and genetic background.     

Inactivity of styrene in the mouse sperm morphology test

Male F1-mice (C3H/He X C57B1/6J) were exposed to styrene by inhalation (150 and 300 ppm; 6 h per day; 5 days) or intraperitoneally (175, 350 and 700 mg/kg per day, 5 days). No statistically significant increase was detected in the frequency of abnormal sperm heads 3 weeks (spermatids exposed) or 5 weeks (late spermatogonia/early spermatocytes exposed) after the beginning of the exposures.     

Disposition of inhaled isoprene in B6C3F1 mice

Isoprene (2-methyl-1,3-butadiene) is the monomeric unit of widely occurring natural products called terpenes. Isoprene is widely used in industry with nearly 1.1 million pounds produced in the United States in 1987. The purpose of this investigation was to determine the toxicokinetics of inhaled isoprene in B6C3F1 mice and to compare the data to previously published toxicokinetic data in F344 rats (A. R. Dahl, L. S. Birnbaum, J. A. Bond, P. G. Gervasi, and R. F. Henderson, 1987. Toxicol. Appl. Pharmacol. 89, 237-248). The comparative toxicokinetics in the two species will be useful for extrapolation of rodent toxicity data to humans. Male B6C3F1 mice were exposed to nominal concentrations of 20, 200, and 2000 ppm isoprene or [14C]isoprene for up to 6 hr. For all exposures, steady-state levels of isoprene were reached rapidly (i.e., within 15 to 30 min) after the onset of exposure. The mean (+/- SE) steady-state blood levels of isoprene (identified by headspace analysis) for the 20, 200, and 2000 ppm exposures were 24.8 +/- 3.3, 830 +/- 51, and 6800 +/- 400 ng isoprene/ml blood, respectively. At the two higher exposure concentrations, the increases in blood levels of isoprene were proportional to the increases in air concentrations of isoprene. There was approximately a 2.3-fold decrease in the retained 14C/inhaled 14C ratio with increasing exposure concentration. Depending on the exposure concentration, from 52% (20 ppm isoprene) to 73% (2000 ppm isoprene) of the metabolite-associated (nonisoprene) radioactivity was excreted in the urine over a 64-hr postexposure period. 14CO2 exhalation after the end of the 6-hr exposure was minimal (2%) at the 20 ppm exposure and increased up to 18% at the higher isoprene exposure concentrations. These data suggest that metabolism of isoprene in mice is nonlinear within the range of exposure concentrations used in this study. Hemoglobin adduct formation reached near-maximum between 200 and 2000 ppm isoprene exposure concentration, consistent with our conclusion that pathways for metabolism of isoprene were saturated. Isoprene metabolites were present in blood after inhalation of isoprene at all concentrations studied. There were substantial differences in the toxicokinetics of inhaled isoprene in mice compared to rats. In mice, fractional retention of inhaled isoprene, which reflects, in part, metabolism of isoprene, was linearly related to exposure concentrations up to 200 ppm but decreased at 2000 ppm; in rats, fractional retention of inhaled isoprene decreased with increasing exposure concentration over a range of exposures from 8 to 1500 ppm.(ABSTRACT TRUNCATED AT 400 WORDS)     

Toxicity of formaldehyde vapor in B6C3F1 mice exposed for 13 weeks

Groups of 10 male and 10 female B6C3F1 mice were exposed to 0, 2, 4, 10, 20, or 40 ppm of formaldehyde vapor 6 h/day, 5 days/week for 13 weeks. Clinical abnormalities (dyspnea, listlessness, and hunched posture), significant mortality, and body weight loss were observed in the 40 ppm groups. Pathologic changes were observed in the nose, larynx, trachea, and bronchi of treated males and females and in the uterus and ovaries of treated females. Squamous metaplasia and inflammation were present in the nasal tissues of male and female mice in the 10, 20, and 40 ppm groups and in the larynx of males and females in the 20 and 40 ppm groups. The trachea had squamous metaplasia and hyperplasia of the epithelium in addition to submucosal fibrosis and inflammation in the 20 and 40 ppm groups. In some mice, epithelial-lined, irregular connective tissue bands spanned the tracheal lumen. Metaplasia of the bronchial epithelium was confined to the 40 ppm exposure groups. These effects on the respiratory system were more prevalent in male than in female mice. Hypoplasia of the uterus and ovaries, probably secondary to body weight loss, was confined to the 40 ppm exposure group. In conclusion, 13-week inhalation exposures of B6C3F1 mice to 10, 20, and 40 ppm of formaldehyde vapor induced histologic lesions in the upper respiratory system and concentrations of 40 ppm were lethal to those mice.     

Susceptibility to 1,3-butadiene-induced leukemogenesis correlates with endogenous ecotropic retroviral background in the mouse

Previous studies have revealed marked differences in the pattern of carcinogenesis between rats and mice exposed to 1,3-butadiene (BD) that do not appear to be readily explained on the basis of pharmacokinetics or metabolism. Chronic exposure of B6C3F1 mice to BD produces a high incidence of thymic lymphoma (TL) that is not observed in rats. The potential of the endogenous ecotropic retroviral background to influence susceptibility to BD leukemogenesis was examined by comparing the incidence of TL between B6C3F1 and NIH swiss mice. Proviral ecotropic sequences are truncated in the NIH Swiss mouse, and the virus is not expressed. Chronic exposure to BD (1250 ppm) for up to 1 year resulted in a fourfold difference in the incidence of TL between B6C3F1 (57%) and NIH Swiss (14%) mice. These results provide presumptive evidence for retrovirus involvement since NIH Swiss mice lack ecotropic viruses and appear to be relatively resistant to induction of lymphoma by BD.     

Effect of dose on the absorption and excretion of [14C]benzene administered orally or by inhalation in rats and mice

The effect of dose on the absorption and excretion of [14C]benzene was studied using 13-week old male F344/N rats, Sprague-Dawley rats, and B6C3F1 mice. Gastrointestinal absorption of benzene administered by gavage was greater than 97% in these species for doses between 0.5 and 150 mg benzene/kg body wt. At oral doses below 15 mg/kg, greater than 90% of the 14C excreted was in the urine as nonethylacetate extractable material. Above 15 mg/kg, in both rats and mice, an increasing percentage of the administered benzene was exhaled unmetabolized, suggesting saturation of metabolic pathways. Above 50 mg/kg, total metabolites (as determined by 14C in the urine, feces, and carcass after 2 days) were not linearly related to administered dose. Total metabolites per unit body weight was equal in F344/N rats and B6C3F1 mice at gavage doses up to 50 mg/kg; however, total metabolites in mice did not increase at higher doses. For inhalation exposures, the percentage of inhaled benzene that was absorbed and retained during a 6-hr exposure decreased from 33 +/- 6% (mean +/- standard deviation) to 15 +/- 9% in rats, and from 50 +/- 15 to 10 +/- 2% in mice as the exposure concentration was increased from approximately 26 to 2600 micrograms/liter (10 to 1000 ppm at 615 Torr, 23 degrees C). Total metabolite formation was exponentially related to the benzene exposure concentration with one-half the maximal amount of metabolite formation occurring at 220 micrograms/liter (84 ppm) for B6C3F1 mice and 650 micrograms/liter (250 ppm) for F344/N rats. Total metabolites were higher in mice than in rats at any of the vapor concentrations used due mainly to the higher amount inhaled by mice. Saturation of overall metabolism in mice but not in rats at high doses by both routes of administration indicates species differences in metabolism of benzene.     

Inhalation Toxicity of Butylene Oxide

Inhalation Toxicity of Butylene Oxide. Miller, R.R., Quast,J.R., Ayres, J.A. and McKenna, M.J. (1981). Fundam. Appl. Toxicol.1:319–324. Exposure of male and female Fischer 344 ratsand B6C3F1 mice to 0,400,800 or 1600 ppm butylene oxide vapors6 hours per day, 5 days per week, for a total of 9 days duringa 2-week interval revealed a definite species difference insensitivity to these high concentrations of the test material.All mice in the 1600 ppm group were dead prior to the 3rd dayof exposure while all rats exposed to 1600 ppm survived untilscheduled sacrifice with no obvious signs of distress exceptfor a pronounced retardation of growth. Inflammatory and degenerativechanges in the nasal mucosa were detected histopath-ologicallyin rats in the 1600 ppm group. Myeloid hyperpla-sia in the bonemarrow, and elevated mean white blood cell counts for male andfemale rats in the 1600 ppm group may possibly have been relatedto the inflammatory nasal lesions or to generalized stress.A subchronic inhalation toxicity study in which Fischer 344rats and B6C3F1 mice were exposed to 0,75,150 or 600 ppm for13-weeks resulted in no treatment-related mortalities. Slightgrowth retardation, particularly for female rats and mice, wasapparent for animals in the 600 ppm group. Histopathologic examinationsrevealed treatment-related lesions of the nasal mucosa in bothrats and mice in the 600 ppm group. There were no histopathologicobservations in rats or mice in the 75 or 150 ppm groups whichwere considered to be related to exposure to the test material.     

The effects of styrene on lung cells in female mice and rats.

Styrene has been shown to cause an increase in the incidence of lung tumors in CD-1 mice following chronic exposure at 40 and 160 ppm, whereas no treatment-related increase in tumors in any organ was seen in rats chronically exposed to up to 1000 ppm styrene. So far most of the mechanistic studies have been performed with male animals. The aim of the present study was to further elucidate the target cell population in mouse lungs exposed to styrene, and to investigate possible differential in vivo effects (e.g., glutathione depletion, increased lipid peroxidation, and oxidative DNA damage). Groups of female CD-1 mice were exposed to styrene at concentrations of 0, 172 or 688 mg/m3 (0, 40 or 160 ppm) for 6 h per day on 1 day, 5 consecutive days or for 20 days during a 4 week period. Groups of female Crl:CD rats were exposed to styrene at concentrations of 0, 688 or 2150 mg/m3 (0, 160 or 500 ppm) for a single 6 h period or for 6 h per day on 5 consecutive days. No signs of lung toxicity were observed in rats. The cytology of cells in lung lavage fluid provided no signs of an inflammatory response in either rats or mice. In mice, both exposure levels caused decreased CC16 protein concentrations in lung lavage fluid after 1 and 5 exposures and in mouse blood serum throughout the study, suggesting that styrene may cause destruction of Clara cells in mice. Degenerative lesions in mouse Clara cells (vacuolar cell degeneration, cell necrosis) were revealed by electronmicroscopy. After 5 and 20 exposures of mice at 160 ppm, cellular crowding, expressed as an irregular epithelial lining and indicative of a very early hyperplasia was noted. Although a depletion of glutathione was noted in mouse lung homogenates after 20 exposures, there was no evidence of oxidative stress as indicated by unchanged concentrations of 8-OH-deoxyguanosine. Malondialdehyde, an indicator of lipid peroxidation, was slightly increased in mice after 1 exposure at 160 ppm only.     

Comparative Carcinogenicity of Polybrominated Biphenyls with or without Perinatal Exposure in Rats and Mice

Comparative Carcinogenicity of Polybrominated Biphenyls withor without Perinatal Exposure in Rats and Mice. CHHABRA, R.S., BUCHER, J. R., HASEMAN, J. K., ELWELL, M. R., KURTZ, P.J., AND CARLTON, B. D. (1993). Fundam. Appl. Toxicol. 21, 451–460. Chronic toxicity and carcinogenicity studies of a polybrominatedbiphenyl mixture (PBB) were conducted in F344/N rats and B6C3F1mice of each sex. The major objective of the study was to determineif exposure to PBB during the perinatal period, in additionto conventional exposure of animals for 2 years, enhances thesensitivity of the bioassay to identify the carcinogenic potentialof this chemical. The studies were designed to determine thetoxic and carcinogenic effects of dietary PBB in rats and micereceiving (i) perinatal exposure up to 8 weeks of age followedby control diet for 2 years, (ii) exposure for 2 years beginningat the age of 8 weeks, and (iii) combined perinatal/adult exposureto PBB (perinatal exposure to 8 weeks of age followed by adultexposure for 2 years). During the perinatal period, rats wereexposed to PBB at dose levels ranging from 1 to 10 ppm and adultexposure concentrations ranged from 3 to 30 ppm in the diet.In the mice, the dose levels ranged from 3 to 30 ppm in bothperinatal and adult exposure portions of the chronic studies.A total of eight dose groups (including controls) were usedwith 60 animals in each group. Liver was the major target organof PBB toxicity. Perinatal exposure alone (through dietary administrationof 10 ppm PBB to the dams) had no effect on the incidences ofneoplasms in female F344/N rats, but in male rats, perinatalexposure was associated with a marginally increased incidenceof hepatocellular adenomas that may have been related to chemicaladministration. In male and female B6C3F₁ mice, perinatal exposureto 30 ppm PBB resulted in significantly increased incidencesof hepatocellular neoplasms. In adult-only dietary exposurestudies, PBB was carcinogenic in male and female F344/N ratsand male and female B6C3F₁ mice based on increased incidencesof hepatocellular neoplasms. Combined perinatal and adult dietaryexposure to PBB confirmed the findings of the adult-only exposuresfor the increased incidences of hepatocellular neoplasms inrats and mice. In male rats, there were no enhancing effectsof combined perinatal and adult exposure. However, perinatalexposure enhanced the susceptibility of female rats receivingadult exposure of 10 or 30 ppm to the induction of liver neoplasms.For male and female rats, a combined analysis of the incidencesof leukemia in the adult-only, perinatal-only, and combinedperinatal and adult exposure groups revealed an apparent associationbetween increasing incidences of mononuclear cell leukemia andexposure to PBB. In male and female mice, it was not possibleto adequately assess the enhancing effects of combined perinataland adult exposure on hepatocellular tumors, because adult-onlyexposure to 10 or 30 ppm PBB resulted in high incidences (84–98%)of liver neoplasms. However, with increased perinatal exposure,there were increases in the numbers of mice with hepatocellularcarcinomas and in the numbers of mice with multiple hepatocellularadenomas, suggesting that combined perinatal and adult exposureincreases PBB-related hepatocellular carcinogenicity relativeto adult-only exposure.     

Prechronic Inhalation Toxicity Studies of Isobutyl Nitrite

Isobutyl nitrite (IBN) is a volatile liquid that has becomeincreasingly popular as an inhaled recreational drug. To investigateshort-term toxic effects and establish exposure parameters forchronic inhalation studies, F344/N rats and B6C3F1 mice wereexposed to IBN vapors on a 6 hr/day + t90, 5 days/week schedule.Twelve exposures were administered at concentrations of 0, 100,200, 400, 600, and 800 ppm IBN. This exposure series resultedmortality in rats exposed to 600 ppm and mice exposed to 800ppm. Animals exposed at the lower concentrations developed hyperplasiaof the bronchiolar and nasal turbinate epithelium (rats andmice) and lymphocytic atrophy in the spleen and thymus (mice).Longer term, 13-week, subchronic exposures were conducted atconcentrations of 0, 10, 25, 75, 150, and 300 ppm IBN. Exposureto 300 ppm IBN reduced the body weight gains in both sexes ofrats and in female mice. IBN-related clinical pathology changesincluded reduced RBC counts accompanied by moderate increasesin mean corpuscular volume and reticulocyte counts, increasedWBC counts, and mildly increased methemoglobin concentration.Bone marrow hyperplasia was observed in all groups of IBN-exposedrats, while in mice only females at l50 ppm IBN displayed thischange. Excessive splenic pulp hematopoiesis was noted in miceat all IBN exposure levels. Respiratory system changes includedincreased lung weights in rats and female mice at 300 ppm, hyperplasiaof the nasal mucosa (male rats at 75 ppm and female rats at150 ppm), and hyperplasia of the lung epithelium (male miceat 150 ppm and female mice at 75 ppm). The results suggestedthat a concentration of 150 ppm could be used as the highestexposure level for subsequent chronic inhalation tests.     

Enhanced suppression of humoral immunity in DBA/2 mice following subchronic exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD).

Previous studies have indicated that mice which differ in their acute susceptibility to responses mediated by the Ah receptor have a pattern of suppression of the antibody response which is consistent with a role by the putative dioxin receptor. The objective of the present investigation was to compare the TCDD-induced suppression of the antibody response following acute and subchronic exposures in B6C3F1 mice, an Ah-high-responder strain, and DBA/2 mice, an Ah-low-responder strain. Results of our initial studies demonstrate that suppression of humoral immunity can be enhanced in DBA/2 mice approximately 10-fold following subchronic versus acute exposures to the same cumulative doses of TCDD. This change in suppression of the antibody response in DBA/2 mice was not accompanied by significant changes in liver weight (hepatomegaly), as was observed in the B6C3F1 strain when exposed under comparable conditions. In contrast, effects on thymus weight (involution) were enhanced in the DBA/2 mice following subchronic exposure and demonstrated a higher degree of atrophy than was seen in the B6C3F1 strain (68 versus 56% decrease in thymic weight at the 42 micrograms/kg cumulative dose). These findings suggest that multiple mechanisms may be operating to suppress humoral immunity in vivo and that the conditions of exposure can alter the toxic effects of TCDD in the DBA/2, Ah-low responsive, mouse strain.