Xpa and Xpa/p53+/- knockout mice: overview of available data.

DNA repair deficient Xpa-/- and Xpa-/-/p53+/- knock-out mice in a C57BL/6 genetic background, referred to as respectively the XPA and XPA/p53 model, were investigated in the international collaborative research program coordinated by International Life Sciences Institute (ILSI)/Health and Environmental Science Institute. From the selected list of 21 ILSI compounds, 13 were tested in the XPA model, and 10 in the XPA/p53 model. With one exception, all studies had a duration of 9 months (39 weeks). The observed spontaneous tumor incidence for the XPA model after 9 months was comparable to that of wild-type mice (total 6%). For the XPA/p53 model, this was somewhat higher (9%/13% for males/females). The 3 positive control compounds used, B[a]P, p-cresidine, and 2-AAF, gave positive and consistent tumor responses in both the XPA and XPA/p53 model, but no or lower responses in wild-type mice. From the 13 ILSI compounds tested, the single genotoxic carcinogen (phenacetin) was negative in both the XPA and XPA/p53 model. Positive tumor responses were observed for 4 compounds, the immunosuppressant cyclosporin A, the hormone carcinogens DES and estradiol, and the peroxisome proliferator WY-14,643. Negative results were obtained with 5 other nongenotoxic rodent carcinogens, and 2 noncarcinogens tested. As expected, both DNA repair deficient models respond to genotoxic carcinogens. Combined with previous results, 6 out of 7 (86%) of the genotoxic human and/or rodent carcinogens tested are positive in the XPA model. The positive results obtained with the 4 mentioned nongenotoxic ILSI compounds may point to other carcinogenic mechanisms involved, or may raise some doubts about their true nongenotoxic nature. In general. the XPA/p53 model appears to be more sensitive to carcinogens than the XPA model.

Sucralose: assessment of teratogenic potential in the rat and the rabbit.

The teratogenic potential of sucralose was examined following gavage administration to pregnant rats and rabbits during organogenesis. Groups of 20 mated rats were dosed on days 6-15 of gestation inclusive at 500, 1000 or 2000mg/kg/day; groups of 16 to 18 inseminated rabbits were dosed on days 6 to 19 of gestation inclusive at 175, 350 or 700mg/kg/day following preliminary studies at higher doses. Concurrent control groups received vehicle alone. Rats were killed on day 21 and rabbits on day 29 of gestation. Foetuses were evaluated at necropsy and after processing for possible soft tissue and skeletal alterations. There was no evidence of teratogenicity for either species. The only observed response to treatment in rats was a slight increase in water intake. Some adult rabbits receiving 700mg/kg/day exhibited marked gastrointestinal disturbance, also seen at higher doses in preliminary studies. Gastrointestinal effects such as these occur non-specifically in response to high doses of poorly absorbed compounds, and in the present study were considered to be responsible for two maternal deaths and four abortions. Full evaluation of rabbit foetuses in the main study (up to 700mg/kg/day) and necropsy of foetuses in a preliminary study with pregnant animals (up to 1000mg/kg/day) showed no evidence of adverse foetal response to sucralose. These teratology studies in both pregnant rodent and non-rodent animal models demonstrate that maternal consumption of high levels of sucralose during the period of organogenesis has no effect on normal foetal development in the rat or rabbit.

Red-wine polyphenols and inhibition of platelet aggregation: possible mechanisms, and potential use in health promotion and disease prevention.

An extract from red-wine grape fermentation, ANOX has been developed as a source of red-wine polyphenols, which are thought to inhibit several of the pathogenic pathways that lead to cardiovascular disease. New data indicate that this extract has a significantly greater effect than either red wine or red-wine powder on the inhibition of platelet aggregation in vitro. Based on this data, about 300 - 500 mg of the extract is equivalent to the daily dose of red-wine polyphenols that appears to protect against cardiovascular disease. The possible synergistic effect of red-wine polyphenols with vitamin C, their vasorelaxing activity and their possible role in preventing over-crosslinking of connective tissues (premature ageing) are considered. The extract contains standardized amounts of the whole spectrum of polyphenolic compounds found in red wine and may provide a valuable reference substance in clinical investigations of the physiological actions of plant polyphenols; its potential use in functional nutrition and preventive medicine is also discussed.

Absence of developmental effects in CF-1 mice exposed to aspartame in utero.

Aspartame (L-aspartyl-L-phenylalanine methyl ester) is a widely used high potency dipeptide sweetener. Developmental toxicology studies have been performed in several species documenting no effects of high doses of aspartame. Recently, a study by Mahalik and Gautieri [1984) Res. Commun. Psychol. Psychiatry Behav. 9, 385-403) reported a delay in the achievement age for the visual placing response in mice pups after maternal administration of high dosages of aspartame during late gestation. In the present study developmental parameters were determined in offspring of CF-1 mice after maternal administration of aspartame at 500, 1000, 2000, and 4000 mg/kg body wt by oral gavage. Aspartame was administered on Days 15 through 18 of gestation. Maternal body weight, food consumption, gestation length, reproductive indices, and litter size were not affected by aspartame treatment. In the pups, body weights, negative geotaxis, and surface and midair righting reflexes were not altered by treatment. There was no delay in the development of the visual placing response regardless of the method employed for assessment (grid or rope) or the manner by which the data were analyzed. There were also no changes in time of eye opening, reflex pupil closure, and ophthalmoscopic examination in the offspring. Thus, neither physical nor functional development was altered in mice after in utero exposure to extremely large dosages of aspartame. More specifically, in utero exposure to aspartame did not affect the development of the visual system in mice.

Reproductive studies of NY-198 in rats. I. Fertility study.

Lomefloxacin (NY-198), a new antibacterial agent, was administered daily by gavage to groups of 22 male and 22 female rats at dosages of 30, 100 or 300 mg/kg/day. Males were dosed for 71 days before pairing and then until termination, and females were dosed for 15 days before pairing, throughout mating and until Day 7 of gestation. Females were killed on Day 20 of gestation for examination of their uterine contents. Males were killed after approximately 14 weeks treatment and their reproductive organs were weighed and retained. At 300 mg/kg/day the majority of animals showed increased salivation, water intake was slightly increased throughout the treatment period in males and before pairing in females whereas food intake showed a slight, transient reduction during the first few days of treatment in both sexes. Body weight gain of males was marginally depressed during the first week of treatment, but no other signs of reaction to treatment were observed. At 30 and 100 mg/kg/day some animals exhibited increased salivation after being dosed. At all dosages, NY-198 was without adverse effects upon mating performance and fertility, or upon survival, growth and development in utero. On the basis of the above results it is considered that the no effect level with respect to reproduction and breeding performance of treated F0 animals and the in utero development of the foetuses was 300 mg/kg/day. A dosage of 100 mg/kg/day was considered to be the no effect level for somatic changes in the F0 animals, and even at the highest dosage of 300 mg/kg/day only slight effects were recorded on the F0 animals.

Reproductive studies of NY-198 in rats. II. Teratology study.

Lomefloxacin (NY-198), a new antibacterial agent, was administered daily by gavage to groups of 32 pregnant female rats of the CD strain at dosages of 30, 100 or 300 mg/kg/day from Day 7 to Day 17 of gestation. Twenty-one females in each group were killed on Day 20 of gestation for examination of their uterine contents. Eleven females in each group were allowed to deliver their litters and the offspring were examined for growth and functional development. At the highest dosage (300 mg/kg/day), there was a small reduction in maternal weight gain and a transient reduction in food intake during the treatment period. Foetal and placental weights were markedly reduced. However, survival, growth and development of F1 offspring were unaffected and, with the possible exception of a slight reduction in F2 foetal weight, their reproductive performance was unimpaired. At the intermediate level (100 mg/kg/day), maternal body weight gain and food intake during the treatment period were slightly reduced but, with this exception, the performance of F0 females and of the F1 generation was essentially similar to that of the vehicle controls. At the lowest dosage (30 mg/kg/day), no adverse effects were recorded on either the F0 females or the F1 generation. On the basis of the above results 30 mg/kg/day was considered to be the no effect level for the F0 females treated during gestation while 100 mg/kg/day administered during gestation to F0 females had no effect upon performance of the F1 generation.

Reproductive studies of NY-198 in rats. III. Perinatal and postnatal study.

Lomefloxacin (NY-198), a new antibacterial agent, was administered daily by gavage to groups of 22 pregnant female rats of the CD strain at dosages of 30, 100 or 300 mg/kg/day from Day 17 of gestation to Day 21 of lactation. Females were allowed to deliver their litters and the offspring were examined for growth and functional development. There was a slight maternal response at the highest dosage (300 mg/kg/day), including increased salivation after dosing, reduced food intake in the treated period of gestation and increased water intake during the lactation period. Gestation length was slightly increased, although remaining within the laboratory background control range; in consequence, body weight of F1 offspring at Day 1 post partum was slightly increased. At 100 mg/kg/day, a few females showed increased salivation after dosing and there was a slight increase in gestation length. Birth weight of F1 offspring was slightly increased at 30 and 100 mg/kg/day but all values were within laboratory background control ranges. The survival, functional responses and fertility of F1 offspring were essentially unaffected by NY-198. On the basis of the above results, 30 mg/kg/day was considered to be the no-effect level for the F0 females treated during late gestation and lactation whilst 300 mg/kg/day administered to the F0 females had no adverse effect upon their offspring.

The effect on fertility of immunizing female sheep with ram sperm acrosin and hyaluronidase.

Female sheep were injected with highly purified and partially purified preparations of ram sperm acrosin and hyaluronidase. The fertility and immune response of the sheep were monitored. Fertility was not significantly reduced in any single group, though a positive correlation was observed between high antibody titres against acrosin and reduced fertility. Studies on the direct action of sera from the ewes on ejaculated ram spermatozoa did not show any evidence of sperm agglutination or immobilization. Similar studies with denuded spermatozoa (detergent induced 'acrosome reaction') sometimes resulted in agglutination and enzyme inhibition was also seen; there was no correlation between any of these parameters and pregnancy.

The immune response of the genital tract of the female rabbit following systemic and local immunization.

Female rabbits were immunized with a model antigen, horseradish peroxidase (HRP), either locally in the reproductive tract or systemically via the intramuscular route. A sensitive immunoassay was developed to detect antibodies to the HRP in both serum and reproductive tract flushings. Systemic immunization resulted in circulating and reproductive tract antibodies of the IgG class, but none of the IgA class. Local immunization produced IgA and IgG antibodies in the reproductive tract, and also IgG antibodies in serum. Cell-mediated immunity was examined by circulating lymphocyte transformation and skin sensitivity to HRP. Cell-mediated immune responses occurred after systemic immunization, but not after local immunization. The relevance of the results to immunological methods of fertility control is discussed.

Polyamines and their biosynthetic decarboxylases in various tissues of the young rat during undernutrition.

1. Male weanling rats were maintained at a constant body-weight by feeding them reduced amounts of the normal diet for various periods up to 4 weeks. Control male rats were allowed free access to the normal diet and some were killed at the beginning of the experiment and others at the same ages as the experimental rats. 2. After killing by cervical dislocation the rats had their liver, quadriceps muscles and spleen removed. The tissues were weighed and the activities of the enzymes ornithine decarboxylase (ODC; EC 4.1.1.17) and S-adenosylmethionine decarboxylase (SAMD; EC 4.1.1.50) assayed in each tissue. In the liver the content of the polyamines (spermidine and spermine) and putrescine was also measured. 3. The liver and quadriceps muscles showed an over-all maintenance of weight during undernutrition, but the spleen lost weight during the first 7 d of undernutrition and then remained constant. The weight of the liver increased by approximately 50% following the daily maintenance feed, but returned to its prefeeding value by 24 h after feeding. 4. During the first 7 d of undernutrition ODC activity decreased in all three tissues, and remained fairly constant thereafter. In the liver there were marked increases in the activity of ODC during the first 4 h after the daily feed, but the activity then decreased to prefeeding values. SAMD activity tended to remain normal in the liver, decreased initially and then returned to normal in the quadriceps muscles, and remained normal initially and then decreased in the spleen. Hepatic SAMD activity showed no consistent response to the daily feed, but quadriceps SAMD activity increased significantly between 1 and 8 h after feeding. 5. Hepatic putrescine content remained constant during undernutrition whilst spermine increased slightly and was then maintained above normal for liver size. Hepatic spermidine content decreased initially and then remained constant. Putrescine increased slightly in response to the daily feed and spermidine increased considerably. Spermine content was unaffected by the daily feed. 6. It is suggested that the response of polyamine synthesis in the various tissues is primarily dependent upon the way in which nutrients are made available to the tissues. The maintenance of spermine content in the liver at the expense of spermidine may be related to differential changes in the nucleic acids.

Growth and skeletal development of the rat during prolonged undernutrition and subsequent rehabilitation.

Male rats of 150 g body weight were given restricted amounts of their normal diet up to the age of 85 weeks, such that they showed no significant change in body weight. They were then rehabilitated by allowing them unlimited access to the normal diet. During rehabilitation, the experimental rats reached a maximum body weight that was only 70% of that normally attained. Body length, however, exhibited better recovery, the experimental animals achieving 96% of the normal body length. The skeleton continued to grow during the first 7 weeks of undernutrition, but thereafter only the caudal half of the pelvic girdle continued to grow. On rehabilitation all skeletal dimensions increased, and all eventually reached the normal range of mature skeletal dimensions. Although the pelvic girdle returned to a normal size, it still manifested some alterations in shape.

Polyamines and their biosynthetic decarboxylases in various tissues of the young rat during recovery from undernutrition.

1. Weanling male and female rats were undernourished for 4 weeks and then rehabilitated by allowing ad libitum feeding. 2. During rehabilitation polyamine-biosynthetic enzymes were examined in the liver, spleen and quadriceps and gastrocnemius muscles. 3. During the first few hours of rehabilitiation there was a marked increase in liver weight, accompanied by a very marked increase in ornithine decarboxylase activity. Increases in the activity of this enzyme in other tissues did not occur until between 2 and 7 days of rehabilitation, at which time there were further increases in enzyme activity in the liver. 4. S-Adenosylmethionine decarboxylase activity also showed marked fluctuations in activity in all the tissues examined. 5. Hepatic putrescine and spermidine concentrations also varied during rehabilitation, but permine concentration remained relatively constant. Both spermine and spermidine were at normal concentrations in the liver from the 10th days of rehabilitation onwards. 6. In all of the tissues examined there were marked sex differences in the parameters studied, particularly in splenic and muscular ornithine decarboxylase activity. 7. In the tissues of the male rats, changes in polyamine synthesis paralled changes in nucleic acid and protein synthesis.

Foetal and placental ornithine decarboxylase activity in the rat--effect of maternal undernutrition.

The activity of the enzyme ornithine decarboxylase and the accumulation of nucleic acids were examined in the rat foetus and placenta during normal development and during maternal undernutrition. Maternal undernutrition resulted in a reduced rate of increase in weight of both the foetus and placenta towards the end of gestation. In the placenta the failure to increase in weight was accompanied by a failure of DNA to increase. In the foetus the amount of DNA failed to increase over a short period and then made a compensatory response to return to a normal amount by the end of gestation. Undernutrition failed to affect RNA in either the foetus or placenta. These results differ from those obtained during maternal protein deficiency. The activity of foetal and placental ornithine decarboxylase was normal during much of gestation in the undernourished group. However, at the same time as the compensatory increase in foetal DNA, there was a marked increase in enzyme activity in both tissues. It is suggested that a single compensatory stimulus is responsible for the changes in both ornithine decarboxylase and DNA.

The response of hind-limb muscles of the weanling rat to undernutrition and subsequent rehabilitation.

1. Weanling male rats were maintained at constant body-weight for 28 d by feeding them reduced amounts of their normal diet. They were rehabilitated for 0, 3, 7, 10 or 16 d, and compared with two sets of control groups, one set of the same body-weight and the other of the same age. 2. The quadriceps, gastrocnemius and anterior tibialis muscles from the left hind-limb were weighed, and DNA, RNA, extracellular protein and intracellular protein estimated in the quadriceps and gastrocnemius muscles. 3. Each muscle responded differently during undernutrition and rehabilitation if compared with "age controls", but if compared with "body-weight controls" there was a tendency for muscle weight to remain appropriate for body-weight. 4. The amount of DNA did not change in the gastrocnemius or quadriceps muscles during undernutrition, and on rehabilitation did not begin to increase until after 7 d. RNA decreased during undernutrition, but increased rapidly on rehabilitation. The rate of increase in RNA was greatest in the quadriceps during the first 3 d, but in the gastrocnemius was greatest between 3 and 7 d. The peak values for the rate of RNA increase corresponded with the initiation of intracellular protein accretion in each muscle. Extra cellular protein increased during undernutrition, and on rehabilitation, responded in the same way as DNA, that is, it increased only after 7 d. 5. It is concluded that the differences found between muscles of the hind-limb during rehabilitation are due mainly to differences in the rates of DNA and intracellular protein accretion. It is suggested that these differences are the result of a differential response of RNA to rehabilitation.