Ana C. Boareto

Toxicity of artemisinin [Artemisia annua L.] in two different periods of pregnancy in Wistar rats

Artemisinin compounds are important for treating multidrug-resistant malaria; however, the possible resorption and abnormalities observed in animal reproduction studies may contraindicate artemisinin use during the first trimester. To evaluate whether artemisinin interferes with developmental outcomes at different periods of pregnancy, Wistar rats were treated by gavage with increasing doses of 7, 35 and 70 mg/kg/day from gestational day [GD] 7 to 13 or 14 to 20. Viable embryos and post-implantation losses, and progestagens and testosterone levels, were monitored in the former treatment group and pregnancy and outcomes data, post-implantation losses and male and female developmental endpoints of the offspring were evaluated in the latter treatment group. Results indicate toxicity for both periods of treatment, with lower sensitivity at later stages of pregnancy. The results showed that dosing with 35 or 75 mg/kg of artemisinin caused high percentages of post-implantation losses that correlated with a trend to lower maternal progestagens and a significant maternal testosterone decrease. These findings demonstrate that oral administration of artemisinin can adversely effect post-implantation development and pregnancy in the rat.

Morinda citrifolia Linn (Noni): In vivo and in vitro reproductive toxicology

UNLABELLED Morinda citrifolia Linn (syn. Noni) is a plant widely used as food and medicine worldwide but there are no toxicological tests about this plant focused on reproduction. AIM OF THE STUDY To investigate possible endocrine activity and toxic effect on the reproductive system of Wistar rats by exposure of aqueous extract of the Morinda citrifolia. MATERIALS AND METHODS Two experimental protocols in vivo were developed, (a) uterotrophic assay and (b) in utero and lactational assay, and one test in vitro to investigate the effect on the contractility of pregnant uteri isolated from rats (doses of the extract: 7.5, 75 and 750 mg/kg). RESULTS The uterotrophic assay indicates presence of in vivo antiestrogenic activity of extract at doses of 7.5 and 750 mg/kg. The in utero and lactation exposure showed that the treatment with extract at the dose of 7.5mg/kg induced a reduction of 50% in parturition index and an increase of 74% in postimplantation losses index. The in vitro test showed that uteri from rats treated with 7.5mg/kg of the extract presented a 50% reduction on contraction induced by arachidonic acid. CONCLUSION The exposure of aqueous extract of Morinda citrifolia in Wistar rats induced reproductive toxicity in nonlinear dose-response.

In vivo and in vitro estrogenic activity of the antidepressant fluoxetine.

Recent years have seen an increase in the use of antidepressant drugs, especially fluoxetine (FLX), in sensitive populations, such as pregnant and lactating women. Although some evidence suggests a possible endocrine action of FLX, no specific studies have been performed to investigate this hypothesis. In the present study, we investigated the possible (anti)androgenic and (anti)estrogenic actions of FLX using Hershberger, uterotrophic (0.4, 1.7, and 17mg/kg), and reporter gene (7.6-129μM) assays. In the Hershberger assay, no differences were observed in androgen-dependent organ weights. However, the uterotrophic and gene reporter assays indicated a possible estrogenic action of FLX. Uterine weight increased in the 1.7 and 17mg/kg/day groups in the 3-day uterotrophic assay in immature rats. Additionally, noncytotoxic concentrations of FLX induced estrogenic responses and increased the estrogenic response of estradiol in MCF-7 breast cancer cells transfected with luciferase.

Screening for in vivo (anti)estrogenic and (anti)androgenic activities of Tropaeolum majus L. and its effect on uterine contractility.

ETHNOPHARMACOLOGICAL RELEVANCE Tropaeolum majus L. (Tropaeolaceae) is a medicinal herb popularly used in Brazil for treatment of inflammatory and cardiovascular diseases. Despite some published data on its efficacy, there are still few toxicological data describing the safety of this plant. The aim of this study was to evaluate the (anti)estrogenic and (anti)androgenic activity of the hydroethanolic extract obtained from Tropaeolum majus L. (HETM), as well as its possible effects on uterine contractility. MATERIALS AND METHODS Three experimental protocols were performed, (a) uterotrophic assay, (b) Hershberger assay and (c) an ex vivo test to investigate the effects of maternal administration of HETM on uterine contractility at the end of pregnancy. In all protocols three doses of the HETM were administered to Wistar rats: 3, 30 and 300mg/kg. RESULTS In vivo tests for detection of (anti)androgenic and (anti)estrogenic activities did not show any significant alterations. Similarly, no alterations were observed on uterine contractility induced by oxytocin and arachidonic acid. CONCLUSIONS HETM was unable to produce (anti)estrogenic or (anti)androgenic activities in the short-term in vivo screening assays performed. In addition, there was no evidence that HETM can affect uterine contractility following gestational exposure of rats.

Study on the developmental toxicity of combined artesunate and mefloquine antimalarial drugs on rats.

Antimalarial drug combinations containing artemisinins (ACTs) have become first choice therapies for Plasmodium falciparum malaria. Data on safety of ACTs in pregnancy are limited and no previous study has been conducted on the developmental toxicity of artesunate-mefloquine combinations on the first trimester of gestation. To evaluate the developmental toxicity of an artesunate/mefloquine combination, pregnant rats were treated orally with artesunate (15 and 40 mg/kg bwt/day), mefloquine (30 and 80 mg/kg bwt/day) and artesunate/mefloquine (15/30 and 40/80 mg/kg bwt/day) on gestation days 9-11. Dams were C-sectioned on day 20, and their uteri and fetuses removed and examined for soft tissue and skeleton abnormalities. Artesunate increased embryolethality and the incidence of limb long bone malformations on the absence of overt maternal toxicity. Mefloquine (80 mg/kg bwt/day) was maternally toxic and enhanced fetal variations. Combination of artesunate and mefloquine did not enhance their toxicity compared to the toxicity observed after its separate administration. Embryotoxicity of artesunate was apparently attenuated when it is co-administered with mefloquine.

Clinical and non-clinical safety of artemisinin derivatives in pregnancy

Malaria in pregnancy is a clinically wasting infectious disease, where drug therapy has to be promptly initiated. Currently, the treatment of this infection depends on the use of artemisinin derivatives. The World Health Organization does not recommend the use of these drugs in the first trimester of pregnancy due to non-clinical findings that have shown embryolethality and teratogenic effects. Nevertheless, until now, this toxicity has not been proved in humans. Artemisinin derivatives mechanisms of embryotoxicity are related to depletion of circulating embryonic primitive erythroblasts. Species differences in this sensitive period for toxicity and the presence of malaria infection, which could reduce drug distribution to the fetus, are significant to the risk assessment of artemisinin derivatives treatment to pregnant women. In this review we aimed to assess the results of non-clinical and clinical studies with artemisinin derivatives, their mechanisms of embryotoxicity and discuss the safety of their use during pregnancy.

Influence of oily vehicles on fetal testis and lipid profile of rats exposed to di-butyl phthalate

It has been hypothesized that oils containing high levels of omega-3 polyunsaturated fatty acids, such as canola and fish oil, could counteract some of the adverse effects induced by phthalates. In the present study, the influence of different oily vehicles on di-butyl phthalate (DBP)-induced testicular toxicity and lipid profile was investigated. Pregnant Wistar rats were treated by oral gavage from gestation days 13 to 20 with DBP (500 mg/kg/day) diluted in three different vehicles: corn, canola or fish oil. Male fetuses were analyzed on gestation day 20. DBP exposure lowered intratesticular testosterone levels and anogenital distance, regardless of the vehicle used. The percentage of seminiferous cords containing multinucleated gonocytes and cord diameter was increased in DBP-exposed groups, compared with vehicle controls, with no difference between the three DBP-exposed groups. Clustering of Leydig cells was seen in all DBP groups. Lipid profile indicated that administration of canola and fish oil can increase the content of omega-3 fatty acids in rat testis. However, content of omega-3 was diminished in DBP-treated groups. Overall, our results indicate that different oily vehicles did not alter fetal rat testicular toxicity induced by a high DBP dose.

Effects of the combined artesunate and mefloquine antimalarial drugs on rat embryos

Artemisinins combination therapy (ACT) is the first choice therapy for falciparum malaria. Data on the safety of ACTs in pregnancy are limited and controversial and the use is not recommended on the first trimester. To evaluate the effects of isolated and combined artesunate (AS)/mefloquine (MQ) on embryo rats, pregnant rats were treated orally with AS (15 and 40 mg/kg body weight (bwt)/day), MQ (30 and 80 mg/kg bwt/day) and AS/MQ (15/30 and 40/80 mg/kg bwt/day) on days 9–11 post coitum (pc). The dams were euthanized on day 12 pc and gestational and embryos histological parameters were evaluated. Embryolethality and histopathological anomalies were significant when AS was given alone or combined with MQ. Combination of AS and MQ did not enhance their toxicity compared to their separate administrations; on the other side, there was a reduction in the toxic effects of the AS when combined with MQ. Isolated MQ did not induce developmental toxicity.

In Utero and Lactational Exposure to Fluoxetine in Wistar Rats: Pregnancy Outcomes and Sexual Development

This study evaluated the reproductive effects of fluoxetine exposure in utero and during lactation on pregnancy outcomes and the sexual development of offspring. Pregnant Wistar rats were treated daily with fluoxetine (0.4, 1.7 and 17 mg/kg/day) or distilled water by gavage from gestation day (GD) 7 to lactation day (LD) 21. A significant reduction in maternal body weight was observed during pregnancy and lactation in dams exposed to 17 mg/kg fluoxetine. Hormone analysis revealed an increase in progestagen and glucocorticoid metabolites on GD 15 and oestrogen and progestagen metabolites on LD 7 in dams treated with 17 mg/kg fluoxetine. Oestrogen metabolites also were increased on LD 7 in dams treated with 0.4 mg/kg fluoxetine. Besides that, an increase in the weight of the adrenal glands and a reduction in uterine weight in dams exposed to highest dose of fluoxetine were observed. Finally, pup birthweight and the viability and weaning indices also were reduced in animals exposed to 17 mg/kg fluoxetine. Overall, maternal hormonal changes were only observed at the highest dose tested, which also induced maternal and foetal toxicity. No significant changes were seen in dams or offspring exposed to therapeutic‐like doses.

Effects of Angiotensin-Converting Enzyme Inhibitor Derived from Tropaeolum majus L. in Rat Preimplantation Embryos: Evidence for the Dehydroepiandrosterone and Estradiol Role

Although several studies have shown the inhibitory effects of Tropaeolum majus extracts (HETM) on angiotensin-converting enzyme (ACE) activity, no studies have been carried out during the beginning of pregnancy, when humoral and hormonal imbalance may affect zygote and early embryo transport. This study investigates whether HETM can affect embryonic development when administered during the one-cell-blastocyst period. Pregnant Wistar rats received orally the HETM (3, 30, and 300 mg/kg/day) from the 1st to the 7th gestational day. Rats were killed on the 8th day of pregnancy and the following parameters were evaluated: clinical symptoms of toxicity (including organ weights), number of corpora lutea, implants per group, preimplantation losses ratio, and the serum levels of dehydroepiandrosterone (DHEA), estradiol, and progesterone. No clinical symptoms of maternal toxicity were evidenced. On the 8th day of pregnancy, the levels of DHEA and estradiol were increased and significant preimplantation losses were observed at all doses used. The present study reveals that the HETM can raise levels of DHEA and estradiol and induce difficulty in the embryo implantation in the early stages of pregnancy. The data contributes significantly to the safety aspects of using this natural product when trying to get pregnant or during pregnancy.