Incidence and potentiation of external and internal fetal anomalies resulting from chlordiazepoxide and amitriptyline alone and in combination

The teratogenic potential of a combination of chlordiazepoxide (Cdz) and amitriptyline (Amt) was examined with regard to both internal and external anomalies. Timed pregnant golden hamsters were given a single intraperitoneal injection on day 8 of gestation of one of the following: chlordiazepoxide hydrochloride (28.5 mg/kg), amitriptyline hydrochloride (70.3 mg/kg), Cdz-Amt combination (28.5 mg/kg Cdz + 70.3 mg/kg Amt, in order to retain the 1:2.5 dose ratio utilized in a clinically-used preparation of these agents), or saline vehicle (control). Fetuses were recovered on gestation day 15 following maternal sacrifice. Cranial malformations were analyzed in Bouin's-fixed fetuses by making 1-mm coronal sections through each head, whereas visceral anomalies were examined following general dissection of each body. Amt alone produced a significant (P less than 0.05) incidence of bent tail and encephalocele, whereas Cdz significantly (P less than 0.05) altered the male:female ratio of surviving fetuses when compared with saline-injected controls. The Cdz-Amt combination caused significant increases in cranial malformations, open eye, bent tail, abnormal lung, and urogenital anomalies. The teratogenic effects of potentiation between the components of this combination are discussed in terms of external and internal malformations.     

Action of the phenothiazine derivative methophenazine on prenatal development in rats.

Single doses of 100-400 mg/kg or multiple doses of 10 or 50 mg/kg of the phenothiazine derivative methophenzaine were given per osto Wistar rats at various times on the 7th-14th days fo gestation and the fetuses examined near term. Results indicated that methophenazine was mainly embryolethal when administered on the 8th-11th days, and was teratogenic at later times, producing types of malformations that depended on the day of treatment, the most susceptible period being the 13th and 14th days of gestation. Teratogenicity occurred only when the dosages were highly toxic to the pregnant rats. Ribovlavin given ip on the 14th day significantly reduced the embryolethal but not the teratogenic action of methophenazine.     

Teratogenic effects of nigericin, a carboxylic ionophore

Nigericin (Na+ salt) was given intraperitoneally at doses of 5.0 or 7.0 mg/kg on one of gestation days 7-12 to pregnant CD-1 mice. Additional mice were injected ip with 2.5 mg/kg on day 11 or 12 only. Injections on single gestation days reduced fetal growth and increased prenatal deaths. Additional signs of toxicity to the conceptus included treatment-related extra ribs and delayed ossification. Treatment was also associated with gross and skeletal malformations, such as median facial cleft, exencephaly, encephalocele, fused ribs, and anomalous vertebrae and exoccipitals. With the possible exception of the 5.0 mg/kg dose given on gestation day 8, nigericin doses associated with gross or skeletal malformations also resulted in observable maternal toxicity.     

Cocaine as a cause of congenital malformations of vascular origin: experimental evidence in the rat

Cocaine hydrochloride was administered to pregnant Sprague-Dawley rats as a single intraperitoneal dose or as two doses 1-4 hours apart. A single dose administered on day 16 of gestation was teratogenic in a dose-dependent manner, with 40 mg/kg being a no-effect dose and 50 mg/kg the lowest teratogenic dose; 80 mg/kg was lethal to the dam. Forty-eight hours after exposure to a teratogenic dose on day 16 of pregnancy, the fetuses showed severe hemorrhage and edema in the their extremities, particularly the footplates, tail, genital tubercle, and upper lip/nose. When the fetuses were examined on day 21 of gestation, the main externally visible malformations were reduction deformities of the limbs and tail. When two doses of cocaine were administered 1-4 hours apart, the incidence of affected fetuses increased as the time interval between the two doses decreased. Two doses of cocaine administered 2 hours apart were not teratogenic on day 9, 10, 11, 12, 13, or 14 of gestation but did induce reduction deformities on days 15, 16, 17, 18, or 19. The same dose administered 1 hour apart was teratogenic on days 14-19. In general, cocaine administration on gestational days 14, 15, or 16 induced more severe and more widespread hemorrhage and edema than administration on days 17, 18, or 19. In the latter cases, damage was restricted to the distal parts of the hindlimb digits and the tail. The results show that in the rat cocaine is only teratogenic during the late organogenic or postorganogenic period.(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of maternal stress on uranium-induced developmental toxicity in rats

It has been demonstrated that uranium is an embryo/fetal toxicant when given orally or subcutaneously to pregnant mice. On the other hand, maternal stress has been shown to enhance the developmental toxicity of a number of metals. In this study, maternal toxicity and developmental effects of a concurrent exposure to uranyl acetate dihydrate (UAD) and restraint stress were evaluated in rats. Four groups of pregnant animals were given subcutaneous injections of UAD at 0.415 and 0.830 mg/kg/day on Days 6 to 15 of gestation. Animals in two of these groups were also subjected to restraint for 2 hr/day during the same gestational days. Control groups included restrained and unrestrained pregnant rats not exposed to UAD. Cesarean sections were performed on gestation Day 20, and the fetuses were weighed and examined for malformations and variations. Maternal toxicity and embryotoxicity were noted at 0.830 mg/kg/day of UAD, while fetotoxicity was evidenced at 0.415 and 0.830 mg/kg/day of UAD by significant reductions in fetal body weight and increases in the total number of skeletally affected fetuses. No teratogenic effects were noted in any group. Maternal restraint enhanced uranium-induced embryo/fetal toxicity only at 0.830 mg/kg/day, a dose that was also significantly toxic to the dams. As in previous studies with other metals, maternal stress enhances uranium-induced developmental toxicity at uranium doses that are highly toxic to the dams; however, at doses that are less acutely toxic the role of maternal stress would not be significant.     

Developmental toxicity of dichloroacetate in the rat.

Dichloroacetic acid (DCA) is a principal by-product of the chlorine disinfection of water containing humic and fulvic acids, and is also a drug of interest in the therapeutic management of metabolic disorders. The developmental effects of DCA were evaluated in the pregnant Long-Evans rat. In two separate studies, animals were dosed by oral intubation on gestation days 6-15 (plug = 0) with 0, 900, 1,400, 1,900 or 2,400 mg/kg/day and 0, 14, 140, or 400 mg/kg/day. The vehicle control was distilled water. Maternal observations included clinical signs, weight change, and gross evaluation of organ weights and uterine contents at necropsy (day 20). Corpora lutea were counted and uteri stained for implantation sites. Live fetuses were examined for external, skeletal, and soft tissue malformations. Seven dams died during treatment (1,400 mg 1/19, 1,900 mg 2/19, 2,400 mg 4/21), and maternal weight gain was reduced at all except the lowest treatment levels. Liver, spleen, and kidney weights increased in a dose-related manner. The mean percentage of resorbed implants per litter was significantly elevated at greater than or equal to 900 mg/kg/day. Live fetuses showed dose-dependent reductions in weight and length at doses above 140 mg/kg. Statistically significant frequencies of soft tissue malformations ranged from 2.6% (140 mg/kg) to 73% (2,400 mg/kg). These were principally in the cardiovascular system and predominantly comprised defects between the ascending aorta and the right ventricle. Skeletal malformations were not observed in significant numbers in any dose group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Teratogenic effect of D-amphetamine sulphate: histodifferentiation and electrocardiogram pattern of mouse embryonic heart

Pregnant albino mice (ICR random-bred strain) received daily injections of 50 or 100 mg/kg body weight of D-amphetamine sulphate between days 9 and 11 of gestation. Parallel control animals were injected with saline solution. Treated mice were sacrificed on day 15 or 19 of gestation. The embryos were examined for gross malformations and direct embryonic ECG recordings were made; they were weighed, and their hearts were carefully dissected. Microscopic sections of the heart were stained with hematoxylin and eosin. It was found that high doses of D-amphetamine raised the incidence of mortality in the treated pregnant mice up to 40%. The resorption rate in the survivors was high (up to 58%) following the high dose of the drug. Up to 15% of the embryos from the treated groups showed gross malformations, including skeletal and eye malformations and exencephaly. The electrocardiogram (ECG) recordings in most 19-day-old embryos from both treated groups showed a pattern with prolonged Q-T interval (PQT), similar to that of control embryos in the intermediate developmental stages (days 14-16 of gestation). The ECG of control embryos (from day 17 on) resembled that of prenatal fetuses. Microscopically, the hearts of treated embryos showed a large number of undifferentiated cardiac myoblasts. It can be inferred that high doses of D-amphetamine affect embryonic development generally and delay the histodifferentiation of the myocardium, resulting in incomplete maturation of the cardiac muscles, thus leading to the immature ECG pattern, with PQT intervals.     

Potentiating effects of caffeine on teratogenicity of alkylating agents in mice

Teratogenic to subteratogenic doses of x-ray, mitomycin C, MNNG, thio-TEPA, cyclophosphamide, and chlorambucil were administered to pregnant ICR mice together with caffeine at doses of 12.5, 25, or 50 mg/kg on day 11 of gestation. Fetuses were examined for gross malformations on day 18 of gestation. The teratogenicity of mitomycin C was significantly potentiated by caffeine at a dose as low as 12.5 mg/kg. The teratogenicity of chlorambucil was also significantly potentiated by caffeine at 50 mg/kg, but similar potentiation was not observed for x-ray, MNNG, thio-TEPA, and cyclophosphamide.     

Teratogenic Effects of the Organophosphorus Compound Fenchlorphos in Rabbits

Fenchlorphos was administered orally in doses of 0, 12.5, 25 and 50 mg per kg to pregnant rabbits from day 6 to 18 of gestation. No effect on implantation efficacy, number of live fetuses, or fetal weight was observed. The incidence of major malformations such as cardiovascular and brain anomalies was, however, increased in the medicated groups. Major skeletal malformations were more frequent in the medicated groups; minor skeletal variation were about equal in all groups. Dose-relationship was observed for cardiovascular malformations and cerebellar hypoplasia.     

Teratogenic effects of a single oral administration of methylmercuric chloride in mice.

The teratogenic effects of methylmercuric chloride (MMC) given orally as a single dose to pregnant ICR mice on day 10 of gestation were examined. The doses tested were 25, 20, 15 and 10 mg/kg. Controls received distilled water orally. Each group consisted of 20 females. Fetuses were taken on day 18 of gestation for teratological study. The number of resorbed or dead embryos was moderately increased in the 25 mg/kg group. Fetuses from dams given 25, 20 and 15 mg/kg MMC weighed significantly less than those in the control group. Many fetuses with malformations were observed in the treated groups; cleft palate occurred in 100, 58.6 and 28.0% of fetuses from dams given 25, 20 and 15 mg/kg MMC, respectively (statistically significant). Hydronephrosis appeared in 23.8 and 18.5% of fetuses from dams given 25 and 20 mg/kg MMC, respectively (statistically significant). Skeletal variations, incomplete ossification of sternebrae, for example, were also observed in the treated groups. These results indicate that MMC is teratogenic so far as cleft palate is concerned and embryotoxic in ICR mice.     

Effects of eflornithine hydrochloride (DFMO) on fetal development in rats and rabbits

Eflornithine hydrochloride (DFMO) is a highly selective, enzyme-activated, irreversible inhibitor of the enzyme L-ornithine decarboxylase (ODC). Because of its role in the biosynthetic pathway of polyamines, ODC is essential for the growth and development of newly implanted embryonic tissue. In order to assess its effect on embryonic growth and fetal development, at various stages of gestation, DFMO was administered in the drinking water to pregnant rats and rabbits at several concentrations (from 0.03% to 3.0%) and times (from days 7, 10, or 11 through days 18 or 19). Rats were killed on day 21 and rabbits on day 29 of pregnancy (day 1 = day of insemination), and the implantations and fetuses were examined. At a concentration of 1.0% (approximately 1,270 mg/kg/day) in rats and 3.0% (approximately 915 mg/kg/day) in rabbits, maternal food and water consumption and body weight gain were significantly reduced during the treatment period, and all implantations were aborted or resorbed. At lower doses (approximately 200-600 mg/kg/day) fetuses survived to term, though in reduced numbers, and a marked reduction in average fetal weight was seen. At levels of 60 mg/kg/day or lower, there were no deleterious effects to the dams or their offspring. Few malformations were detected at any dose level by gross teratologic examination; nor were any considered to have been drug induced because of their sporadic incidence. The embryotoxicity and severe growth retardation demonstrated by these studies verify that adequate polyamine levels are essential for normal embryonic and fetal development.     

Developmental toxicity of pentostatin (2'-deoxycoformycin) in rats and rabbits

The developmental toxicity of the potent adenosine deaminase (ADA) inhibitor, pentostatin (2'-deoxycoformycin), was investigated in pregnant rats and rabbits administered daily iv doses during organogenesis. Rats received 0, 0.01, 0.10, or 0.75 mg/kg on gestation days 6-15 and rabbits received 0, 0.005, 0.01, or 0.02 mg/kg on gestation days 6-18 and maternal and fetal parameters were evaluated on gestation day 21 (rats) or 30 (rabbits). Live fetuses were examined for external, visceral, and skeletal malformations and variations. In rats, maternal body weight gain and food consumption were significantly suppressed at doses of 0.10 and 0.75 mg/kg during the treatment period but returned to control levels during posttreatment. Increased postimplantation loss and decreased numbers of live fetuses, litter size, and fetal body weight were observed at 0.75 mg/kg. A statistically significant increase in the incidence of vertebral malformations occurred at 0.75 mg/kg. The incidence of certain skeletal variations (extra presacral vertebrae, extra ribs, hypoplastic vertebrae) was also increased at 0.75 mg/kg. Ossification of cervical centra was reduced at 0.75 mg/kg compared with controls. In rabbits, marked maternal toxicity (death, body weight loss, and decreased food consumption) and reproductive toxicity (abortion and premature delivery) occurred in all pentostatin-treated groups. However, there were no significant effects on number of live fetuses, pre- or postimplantation loss, litter size, or fetal body weights in the animals with live litters. There was also no apparent increase in the incidence of malformations or variations in the live fetuses of pentostatin-treated rabbits. Thus, these studies demonstrate developmental toxicity of pentostatin in rats and rabbits, and teratogenicity in rats, at maternally toxic doses.     

Inhibition of isotretinoin teratogenicity by acetylsalicylic acid pretreatment in mice.

Although isotretinoin (ITR) has been suggested to cause malformations via cytopathic effects on embryonic cells, the molecular mechanisms of ITR cytotoxicity in teratogenesis are not clear. Since ITR undergoes metabolism by prostaglandin synthase to a potentially cytotoxic peroxyl free radical, the possible role of prostaglandin synthase metabolism as a modulator of ITR teratogenicity was evaluated. Craniofacial and limb abnormalities were noted in fetuses on day 18.5 of gestation following administration of ITR to pregnant CD-1 mice in a three dose regimen of 100 mg/kg at 4 hr intervals on day 10.5 of gestation (plug day = day 0.5 of gestation). Mice were also treated with acetylsalicylic acid (ASA), an irreversible inhibitor of the cyclooxygenase component of prostaglandin synthase, at doses of 20 and 60 mg/kg body weight 2 hr prior to each ITR dose. ASA pretreatment of mice receiving ITR treatment showed a dose-dependent decrease in the overall incidence of malformations, number of defects per fetus, and the incidence of specific craniofacial and limb defects. Equivalent doses of ASA given to control mice did not cause malformations or alter the incidence of resorptions. These results demonstrate that ASA is able to ameliorate the teratogenic effects of ITR observed in fetal mice near term and indicate that prostaglandin metabolism could play a mechanistic role in ITR teratogenicity.     

Strain differences in teratogenic susceptibility to trypan blue between WM and BDIX rat strains

Female rats of WM (Wistar-Mishima)/Nem strain were mated with WM/Nem (group W) or BDIX/Nem males (group WB), and BDIX/Nem females were mated with BDIX/Nem (group B) or WM/Nem males (group BW). On day 8 of gestation, pregnant females were treated intraperitoneally with 1% aqueous solution of trypan blue at a dose of between 20 and 120 mg/kg of body weight. On day 20 of gestation, fetuses were examined for external, visceral, and skeletal malformations. In group W, fetal mortality increased dose dependently at doses higher than 20 mg/kg, and incidences of external, visceral, and skeletal malformations were significantly higher than control at doses of 30 mg/kg and more. In group B, fetal mortality and the incidence of external malformations were significantly higher than control only in the group treated with 120 mg/kg, and no significant increase of visceral and skeletal malformations was shown. It was confirmed that BDIX strain is much more resistant to trypan blue teratogenicity than WM strain. In group BW, nearly the same teratogenic effects were shown as in group W in terms of fetal mortality and incidence of malformations. However, in group WB, teratogenic effects were not so remarkable as in group BW, suggesting patroclinous effects in teratogenic susceptibility to trypan blue. In group BW, sex differences in teratogenic susceptibility were found; male fetuses were more susceptible to trypan blue than females.     

The Potential of Cr3 [Triaqua‐μ3‐Oxo‐Hexa‐μ‐Propionatotrichromium(III) Chloride] to Reduce Birth Defects in the Offspring of Diabetic CD‐1 Mice

Diabetes mellitus is a growing concern worldwide and leads to multiple complications during pregnancy. Pharmacologic doses of chromium (Cr) have been linked with improving insulin sensitivity and other positive benefits in the treatment of diabetes in animal models. By using streptozotocin induced hyperglycemia in female CD‐1 mice, reproductive outcomes of diabetic and chromium‐dosed diabetic females were examined. After dosing 10 mg/kg Cr in the form of triaqua‐μ₃‐oxo‐hexa‐μ‐propionatotrichromium(III) chloride or Cr3 during gestation days 8–16 (GD8–GD16), all females were sacrificed on gestation day 17 (GD17) and examined for maternal weight gain. The fetuses were examined for gross malformations and for skeletal malformations. The offspring of Cr3‐dosed females tended to have a reduction in the incidence of supernumerary ribs. While hyperglycemia still had negative impacts on the health of dams and their offspring, administration of Cr led to an apparent trend in the reduction in the number of malformations and incidence of supernumerary ribs compared to those of untreated diabetic mothers     

Developmental abnormalities induced by 6-mercaptopurine in the hamster.

Pregnant hamsters were given varying doses of 6-mercaptopurine (6MP) at various times during gestation. The fetuses were examined for both gross and histological malformations which showed that the toxic and teratogenic effects of 6MP were dose and time dependent. The most severe gross malformations induced by 6MP were cleft palate, micrognathia and agnathia, microglossia, short limbs, and gut herniation. Grossly normal appearing fetuses, greated during late gestation, showed malformations at the tissue and cellular level. The effects of 6MP in hamster was compared with other species, and with other growth-supressive agents, and it was deduced that the teratogenicity of 6MP is species and tissue specific. Also, it was recommended that histological observations be made an integral part of the teratological safety analysis.     

Effect of WR-2721 on fetal development in the rat

The radioprotector WR-2721 has been shown to radioprotect all tissues studied except the central nervous system. However, it has not yet been used to radioprotect the fetus. In this study we determined that [14C]WR-2721 injected into pregnant rats quickly passed the placenta and was concentrated by the fetus. In addition, we evaluated the toxicity of WR-2721 (2.5-600 mg/kg) to pregnant rats and their fetuses during the period of major organogenesis at Days 9, 11, and 14 postconception. Pregnant animals were only slightly more (10%) sensitive (LD50, 580 mg/kg) to WR-2721 than nonpregnant cohort animals (LD50, 640 mg/kg). At concentrations of 50 mg/kg or less there was a small but statistically significant increase in fetal deaths, while at doses greater than 300 mg/kg a larger degree of fetal mortality occurred. Maximal fetal weight loss, to about 84% of control, was found at 500 mg/kg. No changes in head dimensions or gross malformations of the surviving fetuses were detected at any time or concentration. Of all the parameters measured in this study none demonstrated a predilection for any specific period of major organogenesis. The results of this study indicate that while WR-2721 demonstrates a dose-related embryotoxicity it is not teratogenic.     

Teratogenic activity of trichloroacetic acid in the rat

Trichloroacetic acid (TCA) is a by-product of the chlorine disinfection of water containing natural organic material. It is detectable in finished drinking water at levels comparable to the trihalomethanes (30-160 micrograms/L). TCA is also formed in vivo after ingestion of hypochlorite and has been identified as a major metabolite of chlorinated hydrocarbons such as trichloroethylene. The developmental effects of TCA were evaluated in the pregnant Long-Evans rat. Animals were dosed by oral intubation on gestation days 6-15 (plug = 0) with 0, 330, 800, 1,200, or 1,800 mg/kg/day. The vehicle control was distilled water. Maternal observations included clinical signs, weight change, and gross evaluation of organ weights and uterine contents at necropsy (day 20). Live fetuses were examined for external, skeletal, and soft tissue malformation. There were no maternal deaths associated with toxicity prior to sacrifice. Weight gain during treatment was reduced at 800, 1,200, and 1,800 mg/kg. Spleen and kidney weights were increased in a dose-related manner. The mean percent of resorbed implants per litter was 34, 62, and 90 at 800, 1,200, and 1,800 mg/kg, respectively. Live fetuses showed dose-dependent reductions in weight and length. The mean frequency of soft tissue malformations ranged from 9% at the low dose to 97% at the high dose. These were principally in the cardiovascular system (interventricular septal defect, levocardia). Skeletal malformations were found only at 1,200 and 1,800 mg/kg and were mainly in the orbit. Based on these observations TCA was considered to be developmentally toxic in the pregnant rat at doses of 330 mg/kg and above.     

Severe embryolethality of artesunate related to pharmacokinetics following intravenous and intramuscular doses in pregnant rats

Artesunate (AS), a rapid, effective, and safe antimalarial drug, has been used for the treatment of malaria for decades. However, severe embryolethality was found for injectable AS in pregnant animals. In the present study, pregnant rats were selected and dosed with AS (GMP product) intravenously (IV) and intramuscularly (IM) at varied doses daily for 13 days from gestation day (GD) 6 to 18. In addition, a toxic dose of 1.2 mg/kg/day was subsequently tested in the GD 6–10, GD 11–15, and GD 16–20 periods of rat pregnancy. A pharmacokinetic study was also conducted to evaluate the bioavailability of AS following the IM administrations. Results showed that no significant adverse effects were found in maternal rats. All of the fetuses were either damaged or reabsorbed by placentas in treated pregnant rats, but doses did not show an adverse effect at 0.4 and 0.5 mg/kg after IV and IM administrations, respectively. The survival rate of fetuses is dose-dependent and the 50% fetus re-absorption doses (FRD₅₀) were 0.61 and 0.60 mg/kg following the IV and IM, respectively. The most drug-sensitive period, showing severe embryotoxicity, was between GD 11 and 15 for injectable AS. When calculated with total concentrations of AS and dihydroartemisinin, an active metabolite of AS, the bioavailability of 97.8% after intramuscular injection was fulfilled to a bioequivalence of that in intravenous treatment. The fact that injectable AS exhibited severe embryolethality after both IV and IM injections seems related to their comparable pharmacokinetic profiles that indicate high peak concentrations in pregnant animals. Birth Defects Res (Part B) 86:385–393, 2009. Published 2009 Wiley-Liss, Inc.     

Teratogenic effect of AY 9944 in rats: importance of the day of administration and maternal plasma cholesterol level

An inhibitor of cholesterol synthesis, AY 9944 (trans-1,4-bis(2-chlorobenzylaminomethyl) cyclohexane dihydrochloride) is teratogenic. A single dose of AY 9944 (50 mg/kg or 75 mg/kg) given to Wistar pregnant rats on the second, fourth, sixth, seventh, or eighth day of gestation induced malformations such as holoprosencephaly. They were often limited to isolated pituitary agenesis. The highest percentage of holoprosencephalic fetuses was found when AY 9944 was given on the fourth day of gestation. Whatever the dose and the day of administration, the lower the maternal plasma cholesterol level, the more frequent were holoprosencephalic fetuses. Therefore, it is suggested that the decrease in maternal plasma cholesterol level is at least one of the factors provoking holoprosencephaly.