Phthalate esters (PEs) are synthetic plasticizers used for a wide range of consumer and industrial products including toys, bottles, household vinyl, automobiles, and medical devices. Previous studies have shown that chronic exposure to PEs is associated with lowered fertility in humans and animal models. For example, chronic exposure to PEs was associated with decreased pregnancy rates in women. Di-(2-ethylhexyl) phthalate (DEHP) caused decreased serum estradiol levels, prolonged estrous cycles, and anovulation in adult cycling rats. Monoethylhexyl phthalate (MEHP), the active metabolite of DEHP, decreased aromatase (Arom) expression in cultured rat granulosa cells. These observations indicate that PEs could target antral follicles in the ovary. Major functions of antral follicles are to produce oocytes and estrogen. Mammalian females are born with a finite number of primordial follicles, of which a small fraction survive and reach the penultimate antral stage, while most die via apoptosis. Any insult that affects the survival of antral follicles can cause fertility problems. Our previous in vitro studies have shown that PEs inhibited antral follicle growth and induced atresia by reducing the levels of estradiol. Further, we have also shown that estradiol supplement partially prevented growth inhibition induced by PEs and fully protected follicles from PE-induced atresia. The mechanism of these PE actions on follicles is unknown. Therefore, the goal of this study was to test the hypothesis that PEs (specifically MEHP and DEHP) act by reducing the expression of aromatase and cell cycle regulators in antral follicles. Antral follicles from adult cycling CD-1 mice were isolated and cultured in α-minimal essential media with or without MEHP (10μg/ml) or DEHP (100 μg/ml) and estradiol (E; 1 and 10 nM) for 96 hr. Dimethyl sulfoxide (DMSO) was used as vehicle control. Follicles without any treatment were also included to monitor the culture conditions. After culture, follicles were processed for qPCR to measure the levels of Arom, cyclin D2 (CCND2), and cyclin dependant kinase 4 (CDK4) genes. We found that DEHP and MEHP significantly reduced the expression of Arom compared to DMSO-treated controls and estradiol restored the expression of Arom in DEHP treated group (DEHP: DMSO=5.77±0.85, DEHP=0.70±0.09. E1nM=5.51±0.21, E10nM=6.89±2.09, E1nM+DEHP=4.09±0.68, and E10nM+DEHP=5.08±0.96; but not in the MEHP-treated group MEHP: DMSO=3.64±0.15, MEHP=0.41±0.05. E1nM=3.23±0.46, E10nM=2.81±0.38, E1nM+MEHP=0.84±0.07, and E10nM+MEHP=0.83±0.09; n=3; p≤0.05). Further, DEHP and MEHP significantly decreased the CCND2 and CDK4 expression compared to DMSO treated controls and estradiol restored the expression of CCND2 and CDK4 in DEHP and MEHP treated groups (CCND2; for DEHP: DMSO=1.96±0.44, DEHP=0.61±0.02. E1nM=2.32±0.15, E10nM=2±0.29, E1nM+DEHP=2.04±0.17, E10nM+DEHP=2.05±0.12, and for MEHP: DMSO=2.67±0.23, MEHP=0.69±0.10. E1nM=2.94±0.72, E10nM=2.91±0.34, E1nM+MEHP=1.89±0.48, and E10nM+MEHP=1.63±0.12; CDK4; for DEHP: DMSO=3.20±0.73, DEHP=0.60±0.09. E1nM=2.35±0.05, E10nM=2.71±0.35, E1nM+DEHP=2.53±0.08, E10nM+DEHP=2.49±0.36, and for MEHP: DMSO=1.75±0.21, MEHP=0.41±0.05. E1nM=2.19±0.15, E10nM=2.29±0.35, E1nM+MEHP=1.96±0.36, E10nM+MEHP=1.55±0.10 n=3; p≤0.05). Collectively, these data indicate that DEHP and MEHP may inhibit the growth and induce atresia of antral follicles by affecting the estradiol biosynthesis and the cell cycle regulators. Support: NIHHD46861 (poster)