Parent R01 Research Summary: Pregnancy is a state of constant growth, proliferation, and modification in the immune system between mother and fetus. For this reason, the maternal immune system is at higher risk for exposures of toxicants during pregnancy. More specifically, toxicants from ambient air pollution are numerous and many are absorbed directly from the lungs into the blood stream, resulting in concentrations of toxicants increasing in pregnancy vs. no pregnancy. We hypothesize that exposure to ambient air pollution, specifically particulate matter less than 2.5 M (PM2.5), could increase the likelihood of immune dysregulation in pregnant women during, and in the short- and long-term periods after pregnancy. We will test this by the use of an extant set of samples already collected and stored from a prior NIEHS funded study (PO1). These stored samples are from cohorts (pregnant vs. non-pregnant) living in Fresno, CA continuously for more than 10 years in which individual estimate exposures of PM2.5 (and other pollutants) have been quantitated using validated and published methods. We have stored 200 blood samples during the 2nd trimester of pregnancy, and longitudinal samples from the mothers (at birth, at 1 yr post birth, and at 3 yrs post birth). We also have stored samples from an age matched set non-pregnant, non-parous females from the same area (n=200). Of the n=400 individuals, approximately 50% live in high vs. 50% live in low PM2.5 exposure areas. Leveraging this unique dataset, we hypothesize that the immune system (after a single pregnancy) —with a specific focus on regulatory T cells or Treg--- will be significantly dysregulated in cell subsets, gene expression, T cell receptor, and cytokine responses in pregnancy vs. no pregnancy and that the extent of this dysregulation will be worsened by exposure to PM2.5. In addition, we will test if these mechanistic differences are driven by underlying epigenetic changes. Moreover, the experimental design, which is longitudinal and covers year-to-year time points will be used to test exploratory hypotheses surrounding sustainability of these changes since we have collected short term vs. long term time points after pregnancy. Supplement Research Summary: The extent of PM2.5-induced gene expression changes in B-cells and Monocytes during pregnancy vs. no pregnancy is unknown. Therefore, we will conduct global gene expression profiling of monocytes and B-cell subsets and perform the comparative transcriptome analysis on sorted monocytes and B-cells from maternal blood vs. cord blood exposed to high vs. low PM2.5 levels (Subaim 1b). Our primary hypothesis is that during pregnancy high exposure to fine particulate matter PM2.5 modulates the epigenetic mark on maternal immune system, by inducing cell type specific epigenetic modifications. To determine the mechanism of PM2.5 mediated changes in pregnancy, we will perform integrative multi-omics analysis and reconstruct the regulatory map of...