Neonatal exposure to a glyphosate-based herbicide alters the histofunctional differentiation of the ovaries and uterus in lambs.

The aim of the present study was to compare the effect of oral and subcutaneous exposure to a glyphosate-based herbicide (GBH) on the female reproductive system, specifically in the ovaries and uterus of prepubertal lambs. To this end, ewe lambs were exposed to a s.c. (n: 5) or an oral (n: 5) environmentally relevant dose of GBH (2 mg/kg/day) or to vehicle (controls, n: 12), from postnatal day (PND) 1 to PND14. Serum glyphosate and aminomethylphosphonic acid (AMPA) concentrations were measured on PND15 and PND45. The ovaries and uterus were obtained and weighed on PND45. Ovarian follicular dynamics and uterine morphological features were determined by picrosirius-hematoxylin staining. The proliferation marker Ki67 was evaluated by immunohistochemistry in ovarian and uterine samples. Glyphosate but not AMPA was detected in serum of exposed lambs on PND15, whereas neither glyphosate nor AMPA were detected on PND45. Controls were negative for glyphosate and AMPA on PND15 and PND45. GBH exposure did not affect ovarian or uterine weight. However, on PND45, the ovary of GBH-exposed lambs showed altered follicular dynamics, increased proliferation of granulosa and theca cells, and decreased mRNA expression of FSHR and GDF9, whereas their uterus showed decreased cell proliferation but no alterations in the histomorphology or gene expression. In conclusion, GBH exposure altered the ovarian follicular dynamics and gene expression, and the proliferative activity of the ovaries and uterus of lambs. It is noteworthy that all the adverse effects found in the ovaries and uterus of both GBH-exposed groups were similar, independently of the administration route.

Perinatal exposure to a glyphosate-based herbicide impairs female reproductive outcomes and induces second-generation adverse effects in Wistar rats.

Glyphosate-based herbicides (GBHs) are the most globally used herbicides raising the risk of environmental exposition. Here, we investigated whether perinatal exposure to low doses of a GBH alters the female reproductive performance, and/or induced second-generation effects related to congenital anomalies or growth alterations. Pregnant rats (F0) received a GBH through food, in a dose of 2 mg (GBH-LD: GBH-low dose group) or 200 mg (GBH-HD: GBH-high dose group) of glyphosate/kg bw/day from gestational day (GD) 9 until weaning. Body weight gain and vaginal canal-opening of F1 females were recorded. Sexually mature F1 females were mated to evaluate their reproductive performance by assessing the pregnancy rate, and on GD19, the number of corpora lutea, the implantation sites (IS) and resorption sites. To analyze second-generation effects on F2 offspring, we analyzed the fetal morphology on GD19, and assessed the fetal length and weight, and the placental weight. GBH exposure neither altered the body weight gain of F1 females, nor vaginal opening onset. Although all GBH-exposed F1 rats became pregnant, a lower number of IS was detected. F2 offspring from both GBH groups showed delayed growth, evidenced by lower fetal weight and length, associated with a higher incidence of small for gestational age fetuses. In addition, higher placental weight and placental index were found in F2 offspring from GBH-HD dams. Surprisingly, structural congenital anomalies (conjoined fetuses and abnormally developed limbs) were detected in the F2 offspring from GBH-HD group. In conclusion, perinatal exposure to low doses of a GBH impaired female reproductive performance and induced fetal growth retardation and structural congenital anomalies in F2 offspring.

Simultaneous determination of losartan and hydrochlorothiazide in human plasma by LC/MS/MS with electrospray ionization and its application to pharmacokinetics.

A method based on a simple liquid-liquid extraction (LLE) followed by high-performance liquid chromatography with negative ion electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) detection was developed for the simultaneous determination of losartan (LOS) and hydrochlorothiazide (HCTZ) in human plasma, using valsartan (VAL) and chlorthalidone (CHTD) as an internal standard, respectively. The acquisition was performed in multiple reactions monitoring (MRM) and the limit of quantification was 4 ng/mL for both LOS and HCTZ. The method was linear in the studied range (4-800 ng/mL for LOS and 4-500 ng/mL for HCTZ). The intra-assay precisions ranged from 2.6-11.9% for LOS and 1.4-8.2% for HCTZ, while the inter-assay precisions ranged from 1.0-8.0% for LOS and 2.5-7.7% for HCTZ. The intra-assay accuracies ranged from 91.3 to 107.6% for LOS and 91.5 to 105.8% for HCTZ, while the inter-assay accuracies ranged from 99.9 to 106.4% for LOS and 97.4 to 101.4% for HCTZ. The analytical method was applied to a bioequivalence study, in which 28 healthy adult volunteers (14 men) received single oral doses (100 mg LOS + 25 mg HCTZ) of reference and test formulations, in an open, two-period, balanced randomized, crossover protocol. Based on the 90% confidence interval of the individual ratios for Cmax and AUC0-inf, it was concluded that the test formulation is bioequivalent to the reference Hyzaar formulation with respect to the rate and extent of absorption of both LOS and HCTZ.

Efficacy of DA-7218, a new oxazolidinone prodrug, in the treatment of experimental actinomycetoma produced by Nocardia brasiliensis.

Two recently synthesized oxazolidinones: (R)-3-(4-(2-(2-methyltetrazol-5-yl)-pyridin-5-yl)-3-fluorophenyl)-5-hydroxymethyloxazolidin-2-one (DA-7157) and its corresponding pro-drug (R)-3-(4-(2-(2-methyltetrazol-5-yl)-pyridin-5-yl)-3-fluorophenyl)-2-oxo-5-oxazolidinyl) methyl disodium phosphate (DA-7218), have shown very good activity against several Gram positive bacteria, including Nocardia and Mycobacterium. In the present work we evaluated the therapeutic in vivo effects of DA-7218 on Nocardia brasiliensis. We first determined the plasma concentration of the prodrug in BALB/c mice using several doses and then tested its activity in an in vivo experimental actinomycetoma murine model. At the end of treatment, there was a statistically significant difference between the three drug receiving groups (25, 12.5 and 5 mg/kg) and the control group(saline solution) (p=0.001), proving that DA-7218 is effective for the treatment of experimental murine actinomycetoma. This compound could be a potential option for patients affected with mycetoma by Nocardia brasiliensis.

Pharmacokinetics and the cardiovascular effects of irbesartan in aortic coarctated rats.

A pharmacokinetic-pharmacodynamic study of irbesartan (IRB) was performed in anesthetized sham-operated (SO) and aortic coarctated (ACo) rats. Anesthetized Wistar rats were used 7 days after ACo procedure or SO. A vascular shunt probe was inserted into the carotid artery for the study of plasma pharmacokinetics. In a separate experiment, a concentric probe was placed into the anterior hypothalamus for the study of the IRB distribution in the central nervous system. IRB (10 mg.kg(-1) i.v.) induced a rapid decrease of the heart rate (HR) in the ACo animals (Delta HR -19.2 +/- 2.0 bpm, n = 6; p < 0.05 vs. basal HR), but not in the SO rats (Delta HR -6.7 +/- 5.1 bpm, n = 6). Moreover, IRB reduced the mean arterial blood pressure in the animals of both experimental groups, but the hypotensive effect lasted longer in ACo rats than in SO animals. Analysis of blood samples showed a lower constant of elimination of IRB in ACo rats (Ke 0.67 +/- 0.28 h(-1), n = 5; p < 0.05) than in SO rats (Ke 1.72 +/- 0.30 h(-1), n = 6). Also, a greater distribution of IRB in the anterior hypothalamus was seen in the ACo rats (area under the curve 32 +/- 4 ng.ml(-1).h(-1), n = 6; p < 0.05) than in the SO rats (area under the curve 12 +/- 1 ng.ml(-1).h(-1)). The protein binding of IRB was similar in both experimental groups (SO rats 7.1 +/- 1.2%, n = 6; ACo rats 7.7 +/- 1.5%, n = 6). In conclusion, ACo reduces the plasma elimination of IRB, increasing the distribution in the central nervous system. The longer hypotensive effect of IRB observed in ACo animals could be explained by the slowest elimination of the drug in ACo rats. On the other hand, the effect of IRB on the HR suggested that angiotensin II modulates this parameter in ACo animals at an early stage of hypertension.

Validation of a new intraarterial microdialysis shunt probe for the estimation of pharmacokinetic parameters.

The aim of our study was to compare pharmacokinetic parameters of a highly bound protein drug, irbesartan, obtained from microdialysis data (MD) of arterial blood and conventional blood samples (BS). A new vascular shunt microdialysis probe was inserted into the carotid artery and one femoral vein was cannulated for i.v. administration of irbesartan. Microdialysis samples were collected every 15 min. Blood samples were taken every 15 min. Levels of drug were measured by HPLC. Pharmacokinetic parameters were estimated using TOPFIT program. Corrected MD were compared with BS taken at same time to determine protein binding. The irbesartan protein binding did not change during the experiment. The estimated Ke from MD and BS were similar (MD: 1.8+/-0.3 h(-1), n=5; BS: 1.7+/-0.2 h(-1), n=5). After protein binding correction for the MD, the estimated values of volume of distribution (Vd) (MD: 1.2+/-0.4 l, n=5; BS: 1.1+/-0.4 l, n=5), clearance (Cl) (MD: 32.3+/-7.3 ml min(-1), n=5; BS: 30.7+/-8.2 ml min(-1), n=5) and AUC (MD: 7.7+/-3.2 microg x ml(-1) h, n=5; BS: 8.8+/-3.4 microg x ml(-1) h, n=5) were similar between MD and BS. In conclusion, these results show that our new probe inserted in the carotid artery provides accurate MD to estimate pharmacokinetic parameters of a highly bound protein drug like irbesartan. On the other hand, MD were also useful to the in vivo study of drug protein binding and saturation in protein binding.