Accumulating evidence has demonstrated that exposure to man-made chemicals disrupts human reproductive health. Elucidation of the relevant mechanisms is critical for designing intervention strategies. The goal of the proposed work is to investigate the role of endocrine disrupting chemicals (EDCs) in modulating tryptophan catabolism as a novel mechanism of environmental exposure-induced adverse pregnancy outcomes. Tryptophan catabolism is a regulator of maternal immune tolerance during pregnancy. Failure to degrade tryptophan in the pregnant mother is linked to a significant proportion of human miscarriages, preeclampsia, and premature labor. A key event of this process is activation of the placental Ido1 gene that encodes for a major tryptophan-degrading enzyme. Our laboratory has shown that in utero exposure to a representative EDC, tetrabromobisphenol A (TBBPA), reduces placental Ido1 gene and protein expression, and is linked to increased rates of fetal resorption. In this proposal, we wish to investigate whether exposure to TBBPA alters pregnancy outcomes by perturbing tryptophan catabolism and reducing maternal-fetal immune tolerance. We will expose female mice to environmentally relevant doses of TBBPA (i.e., 5, 50, and 500 g/kg bw/day) prior to mating, during mating, and throughout pregnancy, and analyze the pregnant mice and conceptuses. We wish to determine if exposures are linked to reduced maternal immunity by analyzing a subset of immunosuppressive T cells called ?regulatory T cells? or ?Tregs?. Using flow cytometry, we will determine Treg number in the decidua and uterus of controls and exposed pregnant mice. Increased tryptophan catabolism during pregnancy has been linked to expansion of maternal Tregs. As elevated maternal Tregs favor pregnancy maintenance, we anticipate that TBBPA-exposed mice have reduced Tregs. To determine if increased rate of fetal loss in TBBPA-exposed pregnant mice is causatively linked to reduced maternal Treg number, we will perform an adoptive transfer study using Tregs from donor Foxp3-GFP mice. Additionally, we will perform liquid chromatography and mass spectrometry-based assays to measure levels of tryptophan and its catabolites in the pregnant dam, fetus, and placenta during sequential stages of pregnancy. We wish to elucidate whether TBBPA-induced effects on maternal immunity and pregnancy outcomes are linked to abnormal levels of tryptophan catabolites. Finally, our preliminary studies show that the Ido1 gene is subject to genomic imprinting. We will investigate effects of TBBPA exposure on imprinting regulation of the Ido1 locus by analyzing allele-specific gene expression, DNA methylation and/or posttranslational histone modifications through quantitative real time PCR, bisulfite sequencing, and chromatin immunoprecipitation.
The proposed application will use the mouse model to investigate how exposure to endocrine disrupting chemicals (EDCs) adversely influences pregnancy outcomes. Previous studies have shown that proper regulation of maternal fetal immune tolerance through tryptophan catabolism is essential for a healthy and successful pregnancy. In this work, we will define how exposure to tetrabromobisphenol A (TBBPA) disrupts tryptophan catabolism and study effects on pregnancy and fetal developmental outcomes.
