Exposure to Benzo(a)pyrene in Early Pregnant Mice Impairs Ovarian Corpus Luteum Function by Interfering with Cell Cycle Regulation Proteins

This study was aimed to explore the effects and mechanism of B(a)P [Benzo(a)Ppyrene] on the function of ovarian corpus luteum during the early pregnancy of mice. For in vivo model, the female and male Kunming mice were mated to produce pregnancy at a ratio of 3:1 every night. At the next morning, the female mice with vaginal copulation plugs were recorded as “pregnant day 1 (d1)”. The pregnant mice were randomly divided into control group and B(a)P-treated group. The B(a)P-treated group received 0.2 mg/(kg·d) B(a)P daily by oral gavage at 0.1 mL/10 g of body weight from d1, and the control group received corn oil. The pregnant mice were sacrificed by cervical dislocation on d4 and d7, and the ovaries were collected immediately. In vitro model, the cultured mouse ovarian granule KK-1 cells were divided into three groups and treated with vehicle alone (control group, 0.1% DMSO), 1.0 IU/mL HCG (HCG group) and 1.0 IU/mL HCG plus 0.5 μmol/L BPDE (HCG+BPDE group) simultaneously for 24 h. ELISA was used to detect the levels of serum estrogen and progesterone. The mRNA levels of 3β-HSD, 17β-HSD and P450SCC were determined by qRT-PCR. Immunohistochemistry was used to detect Ki67 and PCNA expression in ovarian tissue sections, and CCK-8 was used to detect the proliferation of KK-1 cells. Western blot, immunohistochemistry and immunofluorescence were used to measure the expression levels of CyclinA1, CDK2, CDK4, CyclinB1 and GAS1. The morphology of mito chondria was observed by transmission electron microscope and Mitotracprobe. B(a)P decreased the levels of serum estrogen and progesterone compared with the control group. The mRNA levels of 3β-HSD, 17β-HSD and P450SCC were down-regulated by B(a)P. The cell activity was weakened after BPDE exposure in vitro. B(a)P exposure resulted in the down-regulation of Ki67 and PCNA expression. B(a)P or BPDE exposure decreased the expression of CyclinA1, CDK2, CDK4, CyclinB1 and GAS1. Electron microscopy and immunofluorescence results revealed that BPDE exposure could lead to abnormal mitochondria morphology. Ba)P exposure might disturb the cell cycle and affect mitochondrial function, which leaded to luteal dysfunction in early pregnancy mice.