Methylene blue is a dye which is prescribed in human and veterinary medicine to treat chemically-induced methemoglobinemia. MBLU is also used in obstetrics to determine the presence of twin amnionic sacs or premature rupture of the membranes. Clinical reports suggest MBLU may be fetotoxic and studies in pregnant rats indicate MBLU may have an adverse impact on fetal development. In the current study, we determined the effect of MBLU on maternal health and fetal growth, viability, and morphological development. Since MBLU readily distributes to all tissues of the body, it was administered (0, 50, 125, or 200 mg/kg/day) by gavage to pregnant Sprague-Dawley rats during the major period of organogenesis (gd 6-15). Maternal clinical signs, body weight, and food and water consumption were monitored from gd 0 to 20. On gd 20, fetuses were removed from the dams and examined for evidence of developmental toxicity.
In the dams, all doses of MBLU decreased relative food consumption from gd 6 to gd 12 and during the entire dosing period (gd 6 to 15). Relative food consumption subsequently returned to control levels until gd 18 to 20, when an increase was observed in the 125 and 200 mg/kg/day groups. Overall relative food consumption during the postdosing period (gd 15 to 20) was increased in all MBLU groups. Effects on relative water consumption were confined to increases from gd 15 to 20 and limited to the 125 and 200 mg/kg/day groups. All doses of MBLU decreased maternal body weight and body weight gain, especially during the first three days of dosing. Examination of maternal organs revealed dose-dependent increases in spleen weight which reached 127%, 168%, and 227% of control spleen weights in the 50, 125, and 200 mg/kg/day groups, respectively. Absolute liver weight was increased in the 200 mg/kg/day group and relative liver weight was increased at 125 and 200 mg/kg/day. Gravid uterine weight tended to be lower in the MBLU groups.
Examination of the uterine contents indicated MBLU was developmentally toxic. The principal effect of MBLU was to increase the percent resorptions per litter in the 200 mg/kg/day group; 4% in the controls vs. 25% at 200 mg/kg/day. This effect did not translate into a significant decrease in the average live litter size because there was an 8-9% increase in the number of corpora lutea/dam and implantation sites/dam in the 200 mg/kg/day group. Thus, although there was a treatment-related increase in the percent resorptions per litter, average live litter size was comparable to controls in the 200 mg/kg/day group. Average fetal body weight (male and female) was reduced 10-11% relative to controls in the 200 mg/kg/day group. No increase in the incidence of external, visceral, or skeletal malformations or variations was observed at any dose of MBLU.
These results indicate the lowest observed adverse effect level for MBLU-induced developmental toxicity in our study was 200 mg/kg/day and the no observed adverse effect level was 125 mg/kg/day. The LOAEL for maternal toxicity was less than or equal to 50 mg/kg/day; The NOAEL for maternal toxicity could not be determined from this study.