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MicroRNAs (miRNAs) are short non-coding RNA species that play key roles in post-transcriptional regulation of gene expression. MiRNAs also serve as a promising source of early biomarkers for different environmental exposures and health effects, although there is limited information linking miRNA changes to specific target pathways. In this study, we measured liver miRNAs in male B6C3F1 mice exposed to a known chemical activator of the peroxisome proliferator-activated receptor alpha (PPARα) pathway, di(2-ethylhexyl) phthalate (DEHP), for 7 and 28 days at concentrations of 0, 750, 1500, 3000, or 6000 ppm in feed. At the highest dose tested, DEHP altered 61 miRNAs after 7 days and 171 miRNAs after 28 days of exposure, with 48 overlapping miRNAs between timepoints. Analysis of these 48 common miRNAs indicated enrichment in PPARα–related targets and other pathways related to liver injury and cancer. Four of the 10 miRNAs exhibiting a clear dose trend were linked to the PPARα pathway: mmu-miRs-125a-5p, -182−5p, -20a−5p, and -378a−3p. mmu-miRs-182−5p and -378a−3p were subsequently measured using digital drop PCR across a dose range for DEHP and two related phthalates with weaker PPARα activity, di-n-octyl phthalate and n-butyl benzyl phthalate, following 7-day exposures. Analysis of mmu-miRs-182−5p and -378a−3p by transcriptional benchmark dose analysis correctly identified DEHP as having the greatest potency. However, benchmark dose estimates for DEHP based on these miRNAs (average 163; range 126−202 mg/kg-day) were higher on average than values for PPARα target genes (average 74; range 29−183 mg/kg-day). These findings identify putative miRNA biomarkers of PPARα pathway activity and suggest that early miRNA changes may be used to stratify chemical potency.

Objective: The importance of the placenta in mediating the pre- and post-natal consequences of fetal growth restriction has been increasingly recognized. However, the influence of placental sexual dimorphism on driving these outcomes has received little attention. The purpose of this study was to characterize how sex contributes to the relationship between placental metabolism and fetal programming utilizing a novel rodent model of growth restriction. Methods: Fetal growth restriction was induced by maternal inhalation of 0.8 ppm ozone (4 h/day) during implantation receptivity (gestation days [GDs] 5 and 6) in Long-Evans rats. Control rats were exposed to filtered air. At GD 21, placental and fetal tissues were obtained for metabolic and genomic assessments. Results: Growth-restricted male placentae exhibited increased mitochondrial biogenesis, increased oxygen consumption, and reduced nutrient storage. Male growth-restricted fetuses also had evidence of reduced adiposity and downregulation of hepatic metabolic signaling. In contrast, placentae from growth-restricted females had elevated markers of autophagy accompanied by an observed protection against hepatic metabolic perturbations. Despite this, growth restriction in females induced a greater number of hypothalamic gene and pathway alterations compared to growth-restricted males. Conclusions: Increases in mitochondrial metabolism in growth-restricted male placentae likely initiates a sequela of adaptations that promote poor nutrient availability and adiposity. Divergently, the female placenta expresses protective mechanisms that may serve to increase nutrient availability to support fetal metabolic development. Collectively, this work emphasizes the importance of sex in mediating alterations in placental metabolism and fetal programming.