The researcher point of view: evaluating follicle quality and chances to pregnancy David Albertini Kansas University Medical Center Kansas City, USA Oocyte Qualiy As A Developmental Continuum Ovarian Development Ovulation MEIOSIS-GENES-IMPRINTS Transmission of genetic errors….? Embryogenesis MITOSIS-GENES-EPIGENES Determinants of OQ Epi/genetic Integrity • Chromatin and Chromosome-based • DNA damage response(DDR) is active • Aneuploidy is common Metabolic Support • Derived from somatic cells via direct contact • Negatively impacted by FSH and LH • Sustains meiotic cell cycle Repeated COH causes a progressive decrease in mouse oocyte ATP content TZP density is sensitive to FSH/LH stimulation Less contact Less ATP Both methods lead to oocyte chromatin condensation Control Slow Freeze Vitrified DNA damage incurred by cryo can be repaired Bovine strips Slow freeze 24 hr post thaw culture The DDR is initiated and completed in bovine follicles contained within cultured ovarian strips after slow freeze but NOT vitrification. Is metabolic recovery influenced by the type of freezing used?? Lesson 1:Compromised metabolism leads to compromised function How does cryopreservation and subsequent culture impact the follicle’s ability to maintain oocyte metabolism? The dark side of aerobic metabolism • Oxidative stress and the preantral follicle • Oxidative stress and the well-vascularized antral follicle • Intrinsic defense mechanisms • Extrinsic manipulations to foster follicle viability and oogenesis Coupling Oxidative Stress to Survival ROS ATM PAR/LKBp kinase AMPKp AUTOPHAGY TSC2 mTOR Autophagy is a good thing • Involves breakdown of organelles, such as peroxisomes, lysosomes, mitochondria for storage and recycling of membrane and protein precursors. • Is probably part of an intrinsic pathway to maintain oocytes. • Is pharmacologically accessible Lesson 2: Metabolic support from the follicle requires maintenance of TZPs As manifestations of granulosa cell polarity designed to serve and protect the oocyte from genetic or epigenetic damage sustained during cryo and/or culture The researchers perspective • Minimizing gonadotropin exposure at all costs will maintain follicle in an oocentrically favorable environment-in vivo or in vitro • Counteracting oxidative stress continues to be essential via exposure to reducing equivalents or ideally, shifting metabolism away from OX/PHOS • We should soon be able to manipulate ovarian tissue survival pharmacologically Take Home Message • Frozen-thawed ovarian tissues undergo stress adaptive changes that will influence follicle survival and function following transplantation or culture. • The challenge ahead is to minimize, rectify, and/or rehabilitate follicular function and in so doing support oogenesis that results in the production of developmentally competent and genetically stable oocytes. The road to fertility preservation Oocyte Cryopreservation P63, c-abl, imantinib PTEN/AKT/ROS Plausible Mechanisms for Preventing or Ameliorating Damage gammaH2AX/IR Cytoxan/DOX Chemo and IR-induced DNA Damage Ovarian Tissue Cryopreservation Oocyte and Somatic Cell DNA Damage