Poster Session 2 - R6
1,2,3Rosanne McQuaid, 1,2Kyunga Kim, 2Elina Kadriu, 1Nicole Zhang, 1Taline Naranian, 3Razqallah Hakem, 1,2,4Andrea Jurisicova
1 Dept. of Physiology, University of Toronto; 2 Lunenfeld-Tanenbaum Research Institute; 3 University Health Network; 4 Obstetrics & Gynaecology, University of Toronto
Objective: Women that carry germline mutations of the breast cancer associated gene 1 (BRCA1) appear to exhibit accelerated reproductive aging and premature infertility. The maternal aging process induces the deterioration of the ovarian reserve and oocyte quality, which can be detrimental to a woman’s reproductive success. BRCA1 regulates several cellular processes that can influence the oocyte’s maturation including the regulation of DNA replication and repair; transcription and translation; cell cycle checkpoints; genomic integrity; ubiquitination; and mitochondrial metabolism. This study investigates the role of BRCA1 haploinsufficiency on reproductive performance and whether BRCA1 could contribute to the mitochondrial dysfunction that is associated with the aging oocyte phenotype.
Methods: Five-month old BRCA1 heterozygous mice with a mutation in BRCA1 exons 5-6 (Brca1f5−6 ) were compared to wild-type littermates to assess ovarian function and reproductive success. Reproductive performance was monitored for 15-weeks of mice mated to ICR males. The ovarian reserve was evaluated from through serial-histological sections of ovaries from haploinsufficient BRCA1 (n=9) and wild-type mice (n=9). These animals were superovulated, and the matured metaphase II oocytes (MII) were retrieved from the oviducts. MII oocyte quality was microscopically analyzed for markers of DNA damage (γ-H2AX, Cell Signaling) and ubiquitination (Enzo Life Sciences). To evaluate the mitochondrial content and function, the oocytes were probed for total and respiring mitochondria (Mitotracker), heat production (Mito thermo yellow), lipid droplet accumulation (Bodipy), and outputs of oxidative phosphorylation: ATP, superoxides (MitoSox), total ROS (H2DCFDA), and mitochondrial membrane potential (DePsipher).
Results: BRCA1 haploinsuffiency results in reduced reproductive success, producing fewer offspring (20.25 ± 3.783, n=8) than wild-type animals (38.88 ± 2.341, n=8), (p<0.0001). We observed a reduced number of primary (p<0.05) and tertiary (p<0.001) ovarian follicles in the ovarian reserve, and poorer ovulation rates (p<0.05). Total mitochondrial content (p<0.0001) and respiring mitochondria (p<0.0001) are increased with BRCA1 haploinsufficiency. While there is no difference to ATP production, superoxide production (p<0.0001) and total ROS (p<0.001) are significantly reduced. Heterozygous BRCA1 oocytes exhibit a decreased accumulation of lipid droplets (p<0.05) and increased generation of heat (p<0.01).
Conclusion: BRCA1 mutations reduce the reproductive performance and negatively impact the ovarian reserve in a middle-aged mouse model. BRCA1 has a novel role in mitochondrial metabolism, and can influence lipid metabolism and the mitochondrial uncoupling of the oocyte to produce heat potentially through BRCA1’s interaction with acetyl-a-carboxylase. Elucidating the roles of BRCA1 mutations of the ovary are essential in women that test positive for mutations to preserve their fertility to improve reprodutive success, as they are also at risk for breast and ovarian cancer.