Embryotoxic and teratogenic effects of petroleum hydrocarbons in mallards (Anas platyrhynchos)

Egg surface applications of microliter quantities of crude and refined oils of high aromatic content are embryotoxic to mallards (Anas platyrhynchos) and other avian species; applications of aliphatic hydrocarbons have virtually no effect. Mallard eggs at 72 h of development were exposed to a mixture of aromatic hydrocarbons or to aromatic compounds representative to those present in crude oil to assess their toxicity. The class composition of the mixture was similar to that of South Louisiana crude oil, an American Petroleum Institute reference oil. Application of 20 microliter of the mixture reduced embryonic survival by nearly 70%. The temporal pattern of embryonic death was similar to that after exposure to South Louisiana crude oil. Embryonic growth was stunted, as reflected by weight, crown-rump length, and bill length, and there was a significant increase in the incidence of abnormal survivors. When individual classes of aromatic hydrocarbons were tested, tetracyclics caused some embryonic death at the concentrations in the mixture. When classes were tested in all possible combinations of two, no combination appeared to be as toxic as the entire mixture. Addition of the tetracyclic compound chrysene to the aromatic mixture considerably enhanced embryotoxicity, but could not completely account for the toxicity of the crude oil. The presence of additional unidentified polycyclic aromatic hydrocarbons as well as methylated derivatives of polycyclic aromatic compounds such as chrysene may further account for the embryotoxicity of the crude oil.     

Embryotoxic and teratogenic effects of indium chloride in rats and rabbits

Daily indium chloride doses of control (0), 50, 100, 200, or 400 mg/kg were administered orally to Sprague-Dawley rats by gavage, on d 6-15 of gestation, and daily metal doses of control (0), 50, 100, or 200 mg/kg were administered to New Zealand rabbits on d 6-20 of gestation. Further groups of pregnant rats were treated with control (0) or 400 mg/kg indium chloride orally on one of d 8, 9, 10, 11, 12, 13, 14, or 15 of gestation. The dams and fetuses were examined on d 21 (rats) and 30 (rabbits) of gestation, using standard teratological methods. Indium concentration was determined in the maternal and fetal blood, as well as in the amniotic fluid, by atomic absorption spectrometry. Indium was found to cross the placenta and appeared in fetal blood in proportion to the metal concentration of the maternal blood. In the amniotic fluid, indium concentrations remained below the detection limit. In rats, indium chloride produced dose-dependent maternal toxic effects, with a dose of 400 mg/kg inducing embryotoxicity (embryolethality) and teratogenicity. Doses of 200 and 100 mg/kg were embryotoxic (retarding) and teratogenic, causing skeletal and visceral anomalies in addition to external anomalies (rudimentary or missing tail, syndactylia, clubfoot, exencephalia) in rats. In rabbits, 200 mg/kg indium chloride was lethal for the dams and the embryos (some of the animals died, and the number of abortions and full resorptions increased). This dose was found to be teratogenic (caused gross renal anomalies) and increased the frequency of fetuses with skeletal retardation. In rats, the effects of indium chloride causing fetal retardation was found to be independent of exposure time. The teratogenic effects were the highest on d 11 and 12 of gestation, when indium chloride caused gross external malformations. Data suggest that the teratogenic effects of indium chloride can be attributed primarily to a direct cytotoxic action of indium resulting from placental transfer, but the effect is not a selective one, as it appears only in the presence of maternal toxic effects.     

Embryotoxic and teratogenic effects of intraperitoneally administered metavanadate in mice.

Metavanadate was evaluated for developmental toxicity in pregnant Swiss mice. Sodium metavanadate (NaVO3) was administered intraperitoneally on d 6-15 of gestation at doses of 0, 2, 4, or 8 mg/kg/d. On gestation d 18, all live fetuses were examined for external, visceral, and skeletal malformations and variations. Maternal toxicity was observed at 2, 4, and 8 mg/kg/d as evidenced by decreased weight gain during treatment. Increased resorptions and dead fetuses, increased percentage postimplantation loss, and reduced fetal body weight per litter were observed at 4 and 8 mg/kg/d. There were no significant increases in the type or incidence of external and skeletal anomalies, but a significant increase in the incidence of cleft palate was detected at 8 mg/kg/d. The lowest-observed-adverse-effect level (LOAEL) for maternal toxicity was 2 mg NaVO3/kg/d, while 2 mg/kg/d was also the no-observed-adverse-effect level (NOAEL) for significant developmental toxicity.     

Embryotoxic,teratogenic, and metabolic effects of ribavirin in mice

To gain comparative perspective on the teratogenic mechanisms of action of drugs that are precursor analogs or antimetabolites of nucleic acids, we employed a new antiviral agent—ribavirin—which is known in other systems to inhibit the biosynthesis of guanine nucleotides. Pregnant ICR mice were injected at 10th–13th days of gestation (stages 14–20) with a single ip dose of ribavirin in the range of 10–200 mg/kg. All dosages in excess of 25 mg/kg were teratogenic. The optimal teratogenic dose varied with the stage of development, being higher at advanced stages of development. Depending on the dose and stage of treatment, virtually all parts of the skeleton including the craniofacial and limb bones were susceptible to ribavirin. Both the frequency and multiplicity of skeletal defects increased as the dose was raised. The stage dependency of defects in the orofacial bones was markedly apparent. Treatment on Day 10.5 resulted in shortened maxilla in all survivors, while treatment on either Day 11 or 11.5 resulted in a high frequency of reduction in the length of both upper and lowerjaws. Treatment on the 12th day resulted in a very low incidence of effect on the maxilla (4%) but a high frequency (88–100%) of reduction and deformation of the mandible. This enhanced susceptibility of individual facial bones at different stages of development with virtually no overlap provides an experimental model to study cellular phenomena underlying normal and abnormal facial development. Ribavirin, both in vivo and in vitro, inhibited embryonic DNA synthesis. The inhibition was transitory and did not seem to be directly related to the embryolethal activity of the drug. Although the role of metabolic inhibition in precipitating teratogenesis is not clear, cytotoxic action of ribavirin against proliferating limb bud mesenchymal cells is directly associated with the origin of limb deformities.     

Embryotoxic and teratogenic effects of interaction of cadmium and lindane in rats

Embryotoxic and teratogenic effects of cadmium and lindane were studied in rats. Cadmium or lindane alone did not produce any significant malformations in the 20 day old foetuses whereas their combination caused significant reduction in the body weight of dams and pups and increased total embryonic deaths. Skeletal deformities like wavy ribs, reduced skull ossification and reduced caudal vertebrae were potentiated in coexposure group. The study indicated that the simultaneous exposure to cadmium and lindane may have more deleterious effects on the developing foetus compared to either alone.     

Embryotoxic and teratogenic effect of actinomycin D in the Syrian hamster

Actinomycin D determines in the hamster polymorphic malformations involving the eye, the nervous system, the face and the skeleton. The anomalies are associated with a general growth retardation.     

Embryotoxic and teratogenic effects of pesticides in chick embryos: A comparative study using two commercial formulations

Developmental toxicity of two different classes of commercial formulations of insecticides was studied by in ovo treatment of fertilized Rhode Island Red eggs. The first one was a combination of chlorpyrifos and cypermethrin and the second one was spinosad, a fermentation product of soil bacterium, Actinomycetes. In this study, the combination pesticide and spinosad of different concentrations were administered as a single dose in ovo in volumes of 50 μL per each egg on day “0” of incubation. Embryonic growth and development, morphological and skeletal malformations, and hatchability were assessed. The combination insecticide induced explicit alterations in the embryonic growth and development and resulted in malformations particularly to the axial and appendicular skeletal structures, whereas the changes were trivial in case of the spinosad exposure. © 2010 Wiley Periodicals, Inc. Environ Toxicol, 2012.     

Potentiation of the teratogenic effects and altered disposition of diphenylhydantoin in mice fed a purified diet

The effect of a standardized purified diet (AIN-76) on the teratogenic response to diphenylhydantoin (DPH) was studied in mice. Mice were fed either the purified diet or Purina Rodent Laboratory Chow for 2-4 weeks prior to mating, and were treated with either saline or 50 mg/kg of DPH on Days 12, 13, and 14 of gestation (copulatory plug = Day 0). The teratogenic response to DPH was found to be markedly potentiated in mice fed the purified diet (75% cleft palate) as compared to mice fed rodent chow (21% cleft palate). The potentiated teratogenic response to DPH correlated with markedly higher plasma DPH levels in pregnant mice fed the purified diet, indicating that the disposition of DPH was impaired. These effects were attributed to a decreased basal level of drug-metabolizing enzymes in mice fed the purified diet, as indicated by markedly prolonged hexobarbital sleeping times. Modifications of the purified diet, which included the replacement of soluble carbohydrate (50% sucrose) in the purified diet with either cornstarch or casein, did not alter the high incidence of cleft palate. A reduction in the incidence of cleft palate was observed, however, when corn oil in the purified diet was replaced with linseed oil. The replacement of corn oil with linseed oil in the purified diet also restored the hexobarbital sleeping times to those observed in mice fed rodent chow. It is concluded that mice fed purified diets have decreased basal levels of drug-metabolizing activity that alter the disposition of DPH and, as a consequence, potentiate its teratogenic effects.     

Embryotoxic effects of thalidomide derivatives in the non-human primateCallithrix jacchus

The teratogenic potency of the thalidomide (Thd) derivative phthalimidophthalimide (Phtpht) was assessed in the common marmoset (Callithrix jacchus), by oral administration of the relatively high daily dose of 50 mg Phtpht/kg body wt, during the susceptible period (days 48–61 of pregnancy). Since in this species daily doses of only 100 μg/kg body wt of the Thd derivative EM12 already induce typical gross structural abnormalities in nearly 100% of the fetuses, investigations with a small number of these New World monkeys allow a rough estimation of the teratogenic potency of Thd-type substances. Macroscopic inspection and skeletal evaluation of ten fetuses gave no indication of dysmorphogenesis following treatment with Phtpht. We conclude that Phtpht has little, if any, Thd-type teratogenic potency in this non-human primate.     

Embryotoxic effects of crude oil in mallard ducks and chicks

Recent studies in this laboratory have revealed that surface applications of microliter amounts of some crude and fuel oils that coat less than 10% of the egg surface reduce hatching considerably in different avian species. Applications of paraffin compounds that coat equal areas of the egg surface do not reduce hatching suggesting that toxicity is due to causes other than asphyxia. In the present study, 1–10 μl of South Louisiana crude oil, an API reference oil, were applied to the surface of fertile mallard (Anas platyrhynchos) and chicken (Gallus gallus) eggs. Early embryolethality was greater in mallard embryos than in chick embryos, but later embryolethality that coincided with the time of rapid outgrowth of the chorioallantoic membrane was more prevalent in chick embryos. The overall incidence of embryolethality was similar in both species. Retardation of growth as reflected by embryonic body weight, crown-rump length, beak length, and general appearance was more pronounced in chick than mallard embryos. Teratogenic defects were more frequent in chick embryos, and incomplete or abnormal ossification of the skull was the most common. External application of equivalent amounts of a mixture of paraffin compounds present in crude oil had virtually no embryotoxic effects in either species, suggesting that other components including aromatic hydrocarbons and organometallics may cause the embryotoxicity.     

Embryotoxic effects of ethylene glycol monomethyl ether in mice

An embryotoxicity study on ethylene glycol monomethyl ether (EGM) was carried out in ICR mice. They were given EGM daily at 6 dose levels (31.25, 62.5, 125, 250, 500 or 1000 mg/kg body wt) by gastric intubation on days 7 through 14 gestation. On day 18 of gestation all fetuses were examined. Marked and dose-related embryotoxic effects were observed. Skeletal and gross anomalies, reduced fetal weight and feral death were all observed at lower dosages of EGM, while marked leucopenia of the dams occured at the highest dose.     

Embryotoxic effects of thalidomide derivatives in the non-human primate Callithrix jacchus

The response of pregnant marmosets (Callithrix jacchus) to the thalidomide derivative EM 12 was evaluated. EM 12 was selected for these studies because it is more active than thalidomide and is much more stable for hydrolysis. Skeletal gross structural abnormalities were observed when EM 12 was given to marmosets for 3–7 days during the period between days 49 and 60 post ovulation. Using the treatment schedule finally adapted in our laboratory, i.e. treatment during days 51–57 post ovulation, doses of 5 (or 10) mg EM 12/kg body wt induced the typical limb abnormalities known from man with an 80–100% certainty. In some animals we could observe the typical pattern of abnormalities even with doses as low as 1 mg EM 12/kg body wt. Abnormalities of the skeleton induced during this sensitive period are described. None of these (except some bifurcations of ribs) were seen in any of the ten litters (23 fetuses) serving as controls during the period of the study.     

Embryolethal and teratogenic effects of bromofenofos in rats

Bromofenofos, an organophosphorus anthelmintic, was administered by gavage to rats as a single dose (50 mg/kg) on one of days 6 through 14 of pregnancy. The dams were killed on day 21, and the fetuses were removed, weighed and examined by routine teratological methods. A significant increase in fetal resorptions occurred after administration on days 9 through 13, with a maximum on day 10. Approximately 72% of the implants were resorbed after administration on day 10. Fetal body weights were significantly decreased when dams were treated on day 8 or later. The greatest decrease in fetal body weights was observed on day 10, when the fetuses weighed less than the controls by about 44% on the average. The incidence of fetuses with gross, skeletal and internal malformations was significantly increased on days 8 through 10, on days 8 through 11 and on days 8 and 9, respectively. Although various types of malformations were observed, most of them occurred on day 8, when no significant increase in fetal resorptions did occur. Cleft lip, short tail, brachygnathia, anal atresia, absence of genital tubercle, fused pelvic legs and perineal testicles were seen on day 8 as gross malformations. Skeletal malformations mainly affected the vertebrae and ribs. Major internal malformations on day 8 were hydronephrosis, hydroureter, anophthalmia, cleft palate, agenesis of the bladder and renal agenesis. Anophthalmia and/or microphthalmia were observed on days 8 through 10, with the highest incidence on day 9. To further determine the no-effect levels for embryolethal and teratogenic effects, a single dose of 10, 20, 30 or 40 mg/kg was administered by gavage to rats on days 8 or 10 of pregnancy. The no-effect levels of single oral dose for embryolethal and teratogenic effects were considered to be 40 and 30 mg/kg, respectively.     

Embryotoxic and long-term effects of cadmium exposure during embryogenesis in rats

The present study was performed to evaluate the long-term behavioral effect in offspring of a subteratogenic Cd dose administered by the oral route to Wistar rat during organogenesis. First, the teratogenic Cd dose was determined by treating pregnant rats with 20 mg/kg Cd from Day 6 to Day 14 of pregnancy and by visceral and skeletal analysis of their fetuses. In a second experiment, pregnant rats treated with this Cd dose were allowed to give birth and nurture their offspring. The physical and behavioral parameters of the offspring were analyzed in infancy and during adulthood. Results showed that Cd treatment during organogenesis (1) was not able to induce maternal toxicity; (2) induced external malformations; (3) increased significantly fetus anomalies and malformations, with reduced metacarpus ossification, cleft palate and right or left renal cavitation being observed in these animals; (4) did not modify pup body weight or weight gain during the lactation period; (5) improved testis descent and delayed the vaginal opening of pups; (6) did not modify ear unfolding, incisor eruption, eye opening, negative geotaxis or palmar grasp; (7) did not modify the open-field parameters and the stereotyped behavior of male or female pups; and (8) modified male sexual behavior and (9) reduced female sexual behavior. We conclude that prenatal exposure to a teratogenic Cd dose induced in the survivor animals several deleterious effects in their development as well as in adult behaviors, mainly in the sexual sphere.     

Embryotoxic effects of salicylates: role of biotransformation

The three major metabolites of salicylate, o- hydroxyhippurate ( salicylglycine , salicyluric acid), 2,5-dihydroxybenzoate (gentisic acid), and 2,3-dihydroxybenzoate, were examined for their capacities to elicit dysmorphogenesis, embryolethality , and growth retarding effects in an embryo culture system. The effects were compared with those produced by the parent salicylate. At the highest concentrations tested (1.9 mM), none of the three metabolites produced significant increases in the number of malformed embryos or in embryolethality . At the same concentration, all three agents reduced crown-rump lengths and somite numbers slightly but significantly (p less than 0.01), and the dihydroxy metabolites also reduced the embryonic protein content (p less than 0.01). In contrast, the parent salicylate produced large increases in embryolethality ( embryolethality in controls was 6% or less) and malformed embryos at equivalent or lower concentrations. Preincubation of the parent salicylate with various biotransforming systems did not affect embryotoxicity significantly. The most rapid biotransformation of salicylate in vitro was achieved with mitochondrial preparations of monkey kidney as the enzyme source but quantities metabolized were not sufficient to prevent malformations in the culture system. Increased serum protein concentration in the culture medium, however, markedly reduced the capacity of added salicylate to cause malformations. An examination of the kinetics of the dysmorphogenic effects of parent salicylate indicated that 5 hr of exposure elicited nonsignificant increases in numbers of malformations. A significant malformation rate was produced by 9 hr of exposure. In contrast, effects on embryonic growth parameters and embryolethality were greatest after a 24-hr exposure period. The results strongly suggest that the parent salicylate, rather than generated metabolites, was primarily or solely responsible for the malformations observed and that the duration of exposure of embryos to unmetabolized salicylate may be the critical factor for determining teratogenic outcome.     

Embryotoxic effects of thalidomide derivatives on the non-human primateCallithrix jacchus

The teratogenic potencies of the enantiomers of 2-(2,6-dioxopiperidine-3-yl)-phthalimidine (= EM 12), a teratogenic thalidomide analogue, were investigated inCallithrix jacchus, a primate very sensitive to the teratogenic action of this thalidomide analogue. The results indicate that the S-(–)-form of EM 12 is clearly more teratogenic than the R-(+)-form. The interpretation of the studies designed to evaluate stereo-selective differences in the teratogenicity of the enantiomers becomes difficult, since both enantiomers racemise in vivo with appreciable rates (Schmahl et al. 1988a, b). Therefore, it cannot be concluded as yet that the R-(+)-form lacks all teratogenic potential.Abbreviations EM 12 2-(2,6-dioxopiperidine-3-yl)-phthalimidine - S-EM 12 S-(–) enantiomer of EM 12 - R-EM 12 R-(+) enantiomer of EM 12