Effects of oral androstenedione on phospholipid fatty acids, ATP, caspase-3, prostaglandin E(2) and C-reactive protein in serum and livers of pregnant and non-pregnant female rats.

Androstenedione, a steroidal dietary supplement taken to enhance athletic performance, could affect serum and liver lipid metabolism, induce liver toxicity or alter inflammatory response depending on dose and duration of exposure. Pregnancy could further exaggerate these effects. To examine this, mature female rats were gavaged with 0, 5, 30 or 60 mg/kg/day androstenedione beginning two weeks prior to mating and continuing through gestation day 19. Non-pregnant female rats were gavaged over the same time frame with 0 or 60 mg/kg/day androstenedione. Serum was collected and livers were removed from dams on gestation day 20 and from non-pregnant rats after 5 weeks of treatment. Androstenedione had no effect on serum total cholesterol, triglycerides or HDL-cholesterol, but significantly decreased C-reactive protein in pregnant rats and prostaglandin E(2) in serum of both pregnant and non-pregnant rats. There were treatment related decreases in liver ATP and, to a lesser degree, caspase-3 and no change in alkaline phosphatase of pregnant female rats. Androstenedione decreased docosahexaenoic acid in both serum and liver phospholipids of pregnant female rats. In conclusion, oral androstenedione did not result in overt hepatotoxicity in pregnant female rats, but produced modest changes in lipid metabolism and may impair regeneration of injured hepatic cells or tissue.

Effects of oral androstenedione on steroid metabolism in liver of pregnant and non-pregnant female rats.

It is unknown whether androstenedione, a steroidal dietary supplement taken to enhance athletic performance, can affect physiological hormone levels by altering liver enzyme activities that metabolize steroid hormones. Altered hormone levels could be especially devastating during pregnancy. Mature female rats were gavaged with 0, 5, 30 or 60 mg/kg/day androstenedione beginning two weeks prior to mating and continuing through gestation day 19. Non-pregnant female rats were gavaged over the same time frame with 0 or 60 mg/kg/day androstenedione. Livers were removed from dams on gestation day 20 and from non-pregnant rats after five weeks' treatment. Liver microsomes were incubated with 200 microM testosterone, and the reaction products were isolated and analyzed by HPLC. In pregnant rats, formation of 6alpha-, 15beta-, 7alpha-, 16beta-, and 2beta-hydroxytestosterone was increased significantly vs. control at the highest dose level only. Formation of 6beta-hydroxytestosterone increased significantly at both the 30 and 60 mg/kg/day dose levels. In non-pregnant rats, 60 mg/kg/day androstenedione significantly increased formation of 15beta-, 6beta-, 16beta-, and 2beta-hydroxytestosterone. The data suggest that high oral doses of androstenedione can induce some female rat liver cytochromes P450 that metabolize steroid hormones and that the response to androstenedione does not differ between pregnant and non-pregnant female rats.

Flaxseed increased alpha-linolenic and eicosapentaenoic acid and decreased arachidonic acid in serum and tissues of rat dams and offspring.

The effects of dietary flaxseed (FS), and defatted flaxseed meal (FLM) on serum and tissue fatty acid profiles were investigated. Pregnant Sprague-Dawley rats were fed AIN-93 based diets balanced in calories, fat, nitrogen, and fiber. Diets contained 0, 20%, 40% FS or 13% or 26% FLM by weight. The control, FS and FLM diets differed in linoleic acid to alpha-linolenic acid (ALA) fatty acid ratio. These diets were fed continuously during gestation, suckling period and 8 weeks post-weaning (F(1)). FS fatty acids were bioavailable and metabolized by pregnant and F(1) rats. ALA and eicosapentaenoic acid increased; linoleic and arachidonic acid decreased; and docosahexaeonic acid was unchanged in serum, 'gastric milk' and liver of FS and FLM-fed pregnant and F(1) rats. FS more than FLM, changed fatty acids profiles, but FLM and 40% FS significantly reduced serum cholesterol. Dietary 40% FS may have increased oxidative stress as evidenced by a reduction in liver vitamin E.

The Autosow raised miniature swine as a model for assessing the effects of dietary soy trypsin inhibitor.

Toxicological effects of dietary soy trypsin inhibitor (TI) were assessed in male miniature swine, a model chosen for its similarities to human digestive physiology and anatomy. The TI preparation was extracted from defatted raw soy flour. From 1 through 5 weeks of age, piglets were automatically fed either a TI liquid diet [Autosow TI group (ASTI)] or a control liquid diet [Autosow control group (ASC)]. From 6 to 39 weeks of age, these animals received either swine chow and TI or swine chow and control article. The TI diets were formulated to contain a TI activity of approximately 500 mg TI/100 g dry matter. A sow control (SC) group suckled from birth to 6 weeks of age and then fed as the ASC group with swine chow plus control article from 6 to 39 weeks of age. The SC piglets grew faster than ASC piglets during postnatal weeks 1 and 2; however, the ASC piglets were significantly heavier than the SC piglets (P=0.001) at 6 weeks of age. Compared with the ASC group, TI caused a moderate decrease in feed consumption and a moderate but reversible decrease in growth from 2 to 5 weeks of age, but not thereafter. Some control and TI-fed Autosow-reared piglets had loose stools until 6 weeks of age; the effect was significantly greater in the TI-fed group. Otherwise, all swine were active and had normal appearance and behavior.

Pathological evaluation, clinical chemistry and plasma cholecystokinin in neonatal and young miniature swine fed soy trypsin inhibitor from 1 to 39 weeks of age.

The potential toxicity of dietary soy trypsin inhibitor (TI) was evaluated in neonatal miniature swine. From 1 to 6 weeks of age, two groups of male piglets were artificially reared in an Autosow and automatically fed either TI or control liquid diet. From 6 to 39 weeks of age, these two groups were fed either TI or control chow diet. A third group, sow control (SC), suckled from birth to 6 weeks of age, were also weaned to control chow from 6 to 39 weeks of age. Clinical chemistry and plasma cholecystokinin (CCK) determined at 6, 18, 30 and 39 weeks of age, and serum amylase activity with gross and histopathological analyses of major organs at 6 and 39 weeks of age are reported. TI had no effect on plasma CCK, serum amylase activity, or numerous clinical chemistry values. TI-fed piglets had a larger relative liver weight at 6 weeks of age. Relative pancreas weight decreased with age but was not affected by TI. Gross and histopathological analyses of major organs, except the spleen, were within normal limits. Increased incidence of extramedullary hematopoiesis was noted in the spleen of the TI group at 6 but not at 39 weeks of age. There was no consistent pattern in immunohistochemical foci for secretin, gastrin releasing polypeptide or CCK, and no change in DNA, RNA, mitotic index or nuclear density of pancreatic cells. At 6 weeks of age, TI increased pancreatic protein and amylase activity but not trypsin or chymotrypsin activity. None of the effects suggested that this dose of TI was toxic to either the neonatal or sexually mature miniature male swine.

Effect of long-chain fatty acids in the culture medium on fatty acid composition of WEHI-3 and J774A.1 cells.

As a first step in determining the mechanism of action of specific fatty acids on immunological function of macrophages, a comparative study of the effect of long-chain polyunsaturated fatty acids (PUFA) in the medium was conducted in two macrophage cell lines, J774A.1 and WEHI-3. The baseline fatty-acid profiles of the two cell lines differed in the % distribution of saturated (SFA) and unsaturated fatty acids (UFA). J774A.1 cells had a higher % of SFA (primarily palmitic acid) than WEHI-3 cells. Conversely, WEHI-3 cells had a higher % of UFA (primarily oleic acid) than J774A.1 cells. Neither cell line had detectable amounts of alpha-linolenic acid (ALA) or eicosapentaenoic acid (EPA). The most abundant polyunsaturated fatty acid in both cells lines was arachidonic acid (AA). The efficiency of transport of fatty acids from the medium to the macrophages by two delivery vehicles (BSA complexes and ethanolic suspensions) was compared. Overall, fatty acids were transported satisfactorily by both delivery systems. Alpha-linolenic acid and doscosahexenoic acid (DHA) were transported more efficiently by the ethanolic suspension system. Linoleic acid (LA) was taken up more completely than ALA, and DHA was taken up more completely than EPA by both cell cultures and delivery systems. A dose-response effect was demonstrated for LA, ALA, EPA and DHA in both J774A.1 and WEHI-3 cells. Addition of polyunsaturated fatty acids (PUFA) to the cell cultures modified the total lipid fatty acid composition of the cells. The presence of ALA in the culture medium resulted in a significant decrease in AA in both cell lines. The omega-3/omega-6 fatty acid ratio (omega-3/omega-6), polyunsaturated/saturated fatty acid ratio (P/S), and unsaturation index (UI) increased directly with the amount of PUFA and omega-3 fatty acid provided in the medium. The results indicate that the macrophage cell lines have similar, but not identical, fatty acid profiles that may be the result of differences in fatty acid metabolism. These distinctions could in turn produce differences in immunological function. The ethanol fatty-acid delivery system, when compared with the fatty acid-BSA complex system, is preferable for measurement of dose-response effects, because the cellular fatty acid content increased in proportion to the amount of fatty acid provided in the medium. Similar dose-response results were observed in a previous in vivo study using flaxseed, rich in ALA, as a source of PUFA.

Effects of intermittent exposures of aflatoxin B(1) on hepatic and testicular glutathione S-transferase in rats.

Glutathione S-transferase (GST) plays a major role in the detoxification of the potent hepatocarcinogen aflatoxin B(1) (AFB(1)). This study evaluated the effects of intermittent exposures to AFB(1) on hepatic and testicular GST in rats. Male Fischer 344 rats were fed diets containing AFB(1) (0, 0.01, 0.04, 0.4 and 1.6 ppm) intermittently at 4-week intervals up to 20 weeks. The control animals were fed an AFB(1)-free NIH-31 diet. Rats consuming diets with 0.01 ppm AFB(1) did not show the induction of hepatic or testicular GST activity. Intermittent exposures to AFB(1) at concentrations of 0.04-1.6 ppm significantly increased the GST activities. The increase of the enzyme activity was proportional to the dose and length of AFB(1) exposure.

Peroxidation of membrane lipids and oxidative DNA damage by fumonisin B1 in isolated rat liver nuclei.

Fumonisin B1 (FB1), a contaminant of corn, has been reported to be a hepatocarcinogen in rats. In an attempt to understand its mechanisms of action, a model system of isolated rat liver nuclei was used to determine what effects, if any, FB1 might have on nuclear membrane lipids and DNA. The data suggested that FB1 induced lipid peroxidation concurrently with DNA strand breaks in this in vitro system. Iron and copper had no statistically significant stimulatory effects on these reactions. In addition, the active oxygen scavengers catalase, superoxide dismutase (SOD), mannitol and sodium azide had no significant inhibitory effects on the FB1-induced DNA strand breaks. However, a small but significant reduction in lipid peroxidation by catalase and mannitol was observed. These results suggested that hydroxyl radicals may be the initiators of the nuclear membrane lipid peroxidation, which results in production of peroxyl radicals. In turn, the peroxyl radicals may be responsible for the DNA strand breaks. An alternative explanation is that the hydroxyl radicals, produced close to the DNA-bound metal ions, may induce direct site-specific strand breaks, which are insensitive to the scavengers of active oxygen.

Growth patterns in selected organs of the miniature swine as determined by gross macromolecular composition.

As part of a larger study designed to characterize the early developmental stages of the Hormel-Hanford strain miniature pig, the brain, kidney, liver, pancreas, and spleen from male animals were examined for changes in RNA, DNA, and protein contents from 1 to 196 d after birth. Distinct patterns were found for changes with age in macromolecular levels. Protein levels increased from d 1 to 56 in all organs except spleen, in which little change was noted. Gel electrophoresis showed little qualitative change in the liver protein profile during this period. A fat-free, non-nucleic acid, protein-containing fraction, insoluble in hot alkali, appeared in the brain after approximately 1 wk following birth. DNA concentrations decreased markedly from d 1 to d 196 for brain, kidney, and spleen but decreased more gradually for liver and pancreas. RNA levels declined slightly or remained the same in all organs except pancreas, where a large increase occurred from d 1 to weaning (56 d). Growth proceeded in all organs by increases in cell number (hyperplasia), as evidenced by increases in total (level or concentration x organ weight) DNA, or by hypertrophy, as evidenced by increases in the ratio of protein to DNA or by a combination of both processes. Hypertrophic growth was attained by d 56 and continued to sexual maturity in all organs except spleen. Hyperplastic growth continued to sexual maturity in all organs except brain.

Developmental effects of combined exposure to ethanol and vitamin A.

The potential for ethanol (EtOH) to influence the developmental toxicity of vitamin A was investigated. 11 groups of approximately 31 FDA-bred Osborne-Mendel rats received either a control or isocaloric 6.4% EtOH liquid diet (containing 4000 IU vitamin A/litre) ad lib. The vehicle control, EtOH and pair-fed (pair-fed against the EtOH group) groups received corn oil (the vehicle) by gavage. Vitamin A was administered by gavage without EtOH at 40,000, 80,000, 120,000 or 160,000 IU/kg daily. Vitamin A was administered by gavage at 10,000, 20,000, 40,000 or 80,000 IU/kg with EtOH ad lib., daily throughout the study. Combined EtOH and vitamin A resulted in significant reductions in maternal diet consumption and body weight when doses of vitamin A were as low as 10,000 IU/kg. The most severe effects on overall (days 0-20) maternal body weight gain were observed in the groups receiving 120,000 or 160,000 IU vitamin A/kg alone or EtOH in combination with 80,000 IU vitamin A/kg. The overall diet consumption (days 0-20) paralleled the overall weight gain. In general, pups exposed to ethanol and vitamin A had a tendency to weigh less than those exposed to vitamin A alone, but to weigh more than those exposed to EtOH alone. EtOH combined with vitamin A at 80,000 IU/kg resulted in an increased incidence of cleft palate relative to the vehicle control or either treatment alone. The incidence of exencephaly and protruding tongue was significantly greater in the group given vitamin A at 160,000 IU/kg, compared with the vehicle control group. The most consistent statistically significant skeletal finding in the groups receiving combined treatment was a treatment-related increased incidence of supernumerary ribs [14th rib (C7), 14th rib bud (L1) and 15 ribs]. In addition, the incidence of misshapen zygomatic arch was also significantly increased in the group exposed to EtOH and vitamin A at 80,000 IU/kg. The incidence of moderately enlarged renal pelvis and severely enlarged ureter proximal to the kidney was increased in the group exposed to EtOH and vitamin A at 80,000 IU/kg relative to the vehicle control, or either treatment alone. Therefore, for some of the endpoints examined in this investigation, it would appear that ethanol potentiates the developmental effects of vitamin A.

Effect of ethanol and vitamin A excess on vitamin A status in the liver, plasma and foetuses of pregnant rats.

The effect of maternal consumption of dietary ethanol and high doses of vitamin A by gavage was investigated by evaluating plasma, liver and foetal vitamin A in Osborne-Mendel pregnant rats with a view to assessing whether ethanol modulated the potential toxicity of excess vitamin A. All groups received 4000 IU vitamin A/litre in a liquid diet. Ethanol-exposed groups also received 6.4% (v/v) ethanol in the liquid diet. Vitamin A was administered by gavage once per day in corn oil in doses ranging from 10,000 to 160,000 IU/kg body weight. Plasma vitamin A levels in ethanol-exposed groups were similar to levels in a pair-fed group. Plasma vitamin A levels were similar in the group given ethanol plus 40,000 IU vitamin A/kg and the group given 40,000 IU vitamin A/kg only, but were higher in the group receiving ethanol plus 80,000 IU vitamin A/kg than in the group given 80,000 IU vitamin A/kg only. Retinyl esters were present in the plasma of animals receiving 160,000 IU vitamin A/kg only, indicating possible saturation of the liver with vitamin A. Retinyl palmitate levels in female foetuses of the group administered ethanol plus 80,000 IU vitamin A/kg were significantly higher than those of the group administered 80,000 IU vitamin A/kg only; no significant differences in levels of retinyl palmitate in male foetuses were observed between these two groups. This observation suggests a possible sex difference in the modulation of vitamin A toxicity by ethanol in the foetus.

Mineral interactions in rats fed AIN-76A diets with excess calcium.

The effects of moderate increases in dietary calcium on maternal and foetal mineral interactions were studied in Charles River CD/VAF Plus rats. Female rats were given 0.50, 0.75, 1.00 or 1.25% dietary calcium as calcium carbonate in AIN-76A diets for 6 wk before mating, during mating and for 20 days of gestation. Inductively coupled argon plasma-atomic emission spectrometry was used to determine mineral levels in the tissues of non-pregnant rats after 42 days on the diets, in the tissues of pregnant rats on day 20 of gestation and in the whole body of day-20 foetuses. The femurs of the non-pregnant and pregnant rats had a dose-related linear increase in calcium content. In livers of the non-pregnant rats, dose-related linear increases in the phosphorus, zinc and magnesium content were observed, but there was a dose-related decrease in the iron content. There were dose-related linear decreases in the iron and copper contents of the kidneys from the non-pregnant rats. In pregnant rats dose-related linear decreases were observed in the iron content of the liver and in the zinc, iron and magnesium contents of the kidney. The foetuses from rats given a moderate increase in dietary calcium had dose-related decreases in the whole-body contents of phosphorus, iron, copper and magnesium.

Foetal development in rats fed AIN-76A diets supplemented with excess calcium.

This study was designed to evaluate the developmental effects of moderate dietary calcium increases in rats fed nutritionally adequate diets. Female Charles River CD/VAF Plus rats were given 0.50 (control), 0.75, 1.00 or 1.25% dietary calcium as calcium carbonate in AIN-76A diets for 6 wk before mating, during mating and for 20 days of gestation. On gestation day 20, the animals were killed and caesarean sections were performed. Both the non-pregnant and pregnant rats in the 0.75, 1.00 and 1.25% groups ate slightly more than did the control group during most of the intervals measured, but not all the increases were statistically significant. There was no consistent pattern of increase or decrease in weight gain. No dose-related changes were found in maternal clinical findings, the average number of implantations, resorptions and viable foetuses, or foetal length or weight. Under the conditions of the study, there were no statistically significant increases as compared with the control group in the litter incidence regarding specific external, visceral or skeletal variations of the foetuses. Dietary calcium was neither foetotoxic nor teratogenic at the concentrations used.

Teratogenic potential of FD & C red no. 3 when given in drinking water.

FD & C Red No. 3 (erythrosine), a commonly used food additive, was administered to pregnant Osborne-Mendel rats to study its teratogenic potential. Dosing solutions of 0.05, 0.1, 0.2 or 0.4% in distilled water were available at all times and corresponded to daily doses of 64, 121, 248 and 472 mg FD & C Red No. 3/kg body weight. Distilled water served as the control. On gestation day 20, the animals were killed and caesarean sections were performed. The treated animals consumed less fluid than did the control animals, but only random decreases were statistically significant and no dose relationship was seen. Only the 0.2% group consumed significantly more feed than the controls during gestation. Maternal weight gain during days 0-20 was not significantly affected in any group. No dose-related changes were seen in maternal clinical findings, implantations, foetal viability, foetal size (weight and length) or visceral development. No dose-related teratogenesis was seen. Skeletal development was not affected; the few statistically significant increases in skeletal variations were not dose related and were considered to be random. FD & C Red No. 3 was neither foetotoxic nor teratogenic at the dose levels tested in drinking water.

Study of the teratogenic potential of FD & C yellow No. 5 when given in drinking-water.

FD & C Yellow No. 5 was available to pregnant Osborne-Mendel rats throughout gestation at dose levels of 0.05, 0.1, 0.2, 0.4 or 0.7% in solution in distilled drinking-water. Based on fluid consumption, the rats received 67.4, 131.8, 292.4, 567.9 and 1064.3 mg FD & C Yellow No. 5/kg body weight/day. Distilled water served as the control. No dose-related changes were seen in mean daily food consumption or maternal body-weight gain. Starting during the second trimester of gestation, fluid consumption was significantly greater in the rats given 0.7% FD & C Yellow No. 5 than in the controls. The females were killed on gestation day 20. No dose-related changes were seen in maternal clinical findings, implantations, foetal viability or foetal size (weight and length). No dose-related foetal terata were seen. Neither visceral development nor skeletal development (sternebral and other skeletal bones) was affected by the dye. The small numbers of statistically significant increases in skeletal variations in the 0.05 and 0.4% levels are considered random because they are not dose related.

Study of the teratogenic potential of guar gum.

Guar gum in the diet at 0, 1, 2, 4, 7.5 or 15% was available ad lib. to male and female Osborne-Mendel rats for 13 wk before mating, during mating and throughout gestation. During gestation, the females consumed 0, 0.7, 1.4, 2.7, 5.2 or 11.8 g guar gum/kg of body weight/day, respectively. The animals were killed on gestation day 20. No behavioural effects were seen in any of the treated dams, and no females died during the experiment. Pregnant females in the treated groups consumed less food than the controls during gestation days 0-20 but the decrease was significant only in the 4 and 7.5% groups and was not dose related. Ingestion of guar gum before mating did not affect fertility. In the dams fed 1-7.5% guar gum, there was no effect on the number of corpora lutea or implantations. The dams fed 15% guar gum had slightly fewer corpora lutea and implantations than the controls but no effect was seen on implantation efficiency in this group. The number of viable foetuses/litter was also reduced slightly but not significantly in the 15% group, but since the number of resorptions was not affected, this decrease appears to be an effect of the decreased number of corpora lutea. There was no compound-related effect on foetal development or sex distribution. No terata were seen.

Potential reversibility of skeletal effects in rats exposed in utero to caffeine.

Groups of Osborne-Mendel rats, killed at three time intervals following mating, were studied to determine whether prenatal skeletal ossification delays observed following low-level caffeine administration represent transient or persistent ossification problems. Group A litters were killed on gestation day 20; group B neonates were killed on post-natal day 0; and group C pups were killed on post-natal day 6. Within each group, dose levels of 0, 0.018, 0.036 or 0.07% caffeine in distilled water were available ad lib. to groups of 30-60 dams from gestation day 0 to day 20. Average daily caffeine consumption was 24.7-29.0 mg/kg body weight for the 0.018% group, 42.7-48.8 mg/kg body weight for the 0.036% group and 70.6-75.1 mg/kg body weight for the 0.07% group. In group A litters, the mean number of viable foetuses was significantly less in the mid-dose and high-dose animals than in the controls. In the same group, the average number of foetuses per litter with at least one sternebral ossification delay was increased significantly in all treated groups and the average number of foetuses per litter with at least two sternebral variations was significantly increased in the mid- and high-dose groups. The percentages of litters containing foetuses with at least two and at least three sternebral variations and the average number of foetuses per litter with at least three sternebral variations were significantly increased only in the high-dose group. Foetuses from the mid- and high-dose groups also had significant increases in certain skeletal defects, namely missing centra and reduced ossification of the dorsal arch. Foetuses from the high-dose group also had significant increases in bipartite supraoccipital, and reduced ossification of the hyoid, metacarpals and metatarsals. In group B, day 0 neonates from all treated groups showed a significantly increased incidence of delayed sternebral ossification (average number of foetuses per litter with one or more missing, incomplete or bipartite sternebrae). The percentages of litters containing neonates with delayed sternebral ossification were also increased significantly in all treated groups. Neonates from the 0.07% level in group B also exhibited a significant increase in the incidence of supernumerary rib bud, and in reduced ossification of the metacarpals, metatarsals and calcaneus bones. Significant increases were also seen, in group B, in the low- and mid-dose animals, respectively, in supernumery rib bud and in reduced ossification of the metatarsals.(ABSTRACT TRUNCATED AT 400 WORDS)

Teratogenic potential of purified pentachlorophenol and pentachloroanisole in subchronically exposed Sprague-Dawley rats.

Male and female Sprague-Dawley (Spartan) rats were exposed to dietary levels of 0, 60, 200 or 600 ppm purified pentachlorophenol (PCP) or pentachloroanisole (PCA) for 181 days, through mating and pregnancy. The daily intakes of PCP were 0, 4, 13 or 43 mg/kg body weight and of PCA were 0, 4, 12 or 41 mg/kg body weight. Animals exposed to PCP generally consumed more food than control animals during pregnancy. Dams at the high-dose level of both compounds showed evidence of toxicity, weighing less on day 0 of gestation and gaining less throughout pregnancy than did the controls. Dams exposed to the high dose of PCP gained less weight during pregnancy (exclusive of the gravid uterus) than control dams. At the 43 mg/kg/day dose level PCP was embryolethal. Foetuses at the lower dose levels of PCP exhibited dose-related decreases in body weights. A reduction in crown-rump length and an increase in foetal skeletal variations were seen at 13 mg/kg/day in PCP animals only. An intake of 41 mg PCA/kg/day was associated with a decrease in the number of corpora lutea and in embryolethality. PCA exposure also resulted in reductions in foetal body weight and crown-rump lengths of males at 4 and 41 mg/kg/day. Female foetuses were unaffected.

Use of dual control groups to estimate false positive rates in laboratory animal carcinogenicity studies.

Tumor incidence data from 18 recently completed carcinogenicity studies utilizing male and female mice and rats were examined to determine if the frequency of significant (p less than 0.05) pairwise differences between the two concurrent control groups employed in these experiments exceeded chance expectation. Although marked study-to-study variability was observed for some tumors, no evidence of extra-binomial within-study variability between the two concurrent control groups was found. The total number of observed significant (p less than 0.05) paired-control differences was virtually identical to what would be expected from the usual binomial model assumptions; the corresponding overall observed (44%) and expected (47-50%) false positive rates were essentially the same. While one should not overgeneralize the implications of these findings, these results should lessen concerns that elevated false positive rates resulting from extra-binomial within-study variability might be adversely affecting the interpretation of long-term laboratory animal carcinogenicity studies. On the other hand, these results reaffirm the conclusions of other investigators that (particularly for commonly occurring tumors) more stringent evidence than an isolated p less than 0.05 effect should be required before an increased tumor incidence is regarded as biologically significant; otherwise, the study may have an unacceptably high false positive rate.

Effect of subchronic dietary administration of butylated hydroxyanisole on canine stomach and hepatic tissue.

Beagle dogs, 29 males and 30 females, were assigned to feeding groups of 0, 1.0 and 1.3% butylated hydroxyanisole (BHA) for 180 days. Animals were observed daily for physical signs of pharmacological or toxicological effect. Except for the production of a reddish-brown urine, no physical signs attributable to BHA administration were observed. Both food consumption and body-weight gain were reduced in the BHA-treated animals. Fifty-one animals completed the study. At termination, tissues were examined for gross BHA-related effects, and specimens were taken for enzyme analysis, light microscopy, electron microscopy and morphometric analysis. The liver showed a significant weight increase over the control in both sexes at both BHA dose levels. Ultrastructural examination of the liver of BHA-treated animals revealed proliferation of smooth endoplasmic reticulum and hepatocytic cytoplasmic myelinoid bodies. Enzyme analysis of hepatic tissue showed a significant increase in mixed-function oxidases, UDP glucuronyl transferase, glutathione S-transferase and epoxide hydratase in the BHA-treated dogs compared with the controls. Light microscopy revealed no proliferative/hyperplastic lesions of the stomach/gastric epithelium. Electron microscopic examination of the lower oesophagus and stomach specimens from representative animals from each treatment group failed to reveal any treatment-related effect as compared with controls.