MITOQUINONE RESCUES STRESS INDUCED MITOPHAGY IN AGED OOCYTES

Abstract

It is well established that decreased oocyte quality from advanced maternal age (AMA) women are associated with dysfunctional mitochondria. Mitophagy is a critical pathway to regulate mitochondrial quality. Our previous research demonstrated reduction in key mitophagy protein Parkin (PRKN) and its activity in oocytes from AMA women. The objective of this study was to investigate if maturing aged mouse oocytes with Mitoquinone (MQ) would improve oocyte mitochondrial quality by rescuing mitophagy. First, oocytes from young (2 months) and aged (13.5 months) mice were matured with or without MQ and the levels of p-PRKN and PRKN were analyzed by Western blot (Jess, ProteinSimple). In the second experiment, young oocytes with or without MQ were matured under normal (6.5%) or high oxygen (20%) environment, and the expression of p-PRKN and PRKN, mitochondria DNA (mtDNA) copy number (analyzed by qPCR) and blastocyst development after in vitro fertilization, blastocyst hatching, and inner cell mass (ICM) ratio were examined. We observed a significant decrease in p-PRKN (0.32±0.09, p≤0.01) and PRKN (0.30±0.1, p≤0.05) in aged compared to young mouse oocytes (normalized to 1). Treatment of aged oocytes with MQ rescued p-PRKN (1.11±0.19 p≤0.01) and PRKN (0.67±0.04, p≤0.05) protein expression. Oocytes matured under high oxygen environment exhibited significantly reduced p-PRKN (0.5±0.1, p≤0.05) compared to oocytes matured under normal oxygen conditions. This coincided with an increase in mtDNA copy number (Young: 56957, High oxygen: 72758 copies). MQ treatment significantly increased p-PRKN protein expression (1.2±0.2, p≤0.05) as well as decreased mtDNA copy number (50312 copies) in oxidatively stressed oocytes. Addition of MQ significantly improved blastocyst development (High O2: 39.5%, High O2+MQ: 70%, p≤0.05), hatching (High O2: 59%, High O2+MQ: 93%, p≤0.05), and ICM ratio (High O2: 14.61%, High O2+MQ: 22.5%, p≤0.0001) of oxidatively stressed oocytes that showed decreased trends in blastocyst development potential. We have demonstrated previously that oocytes from AMA women (>38 years old) exhibited significantly less p-PRKN and PRKN than oocytes from young women (<32 years old). Similar PRKN expression pattern was obtained in young and aged mice and we also observed decreased p-PRKN in oxidatively stressed mouse oocytes, indicating that mitophagy via Parkin pathway is altered in aged and oxidatively stressed oocytes. We have shown for the first time that treating aged and oxidatively stressed oocytes with MQ rescues p-PRKN and PRKN protein expression to ameliorate mitophagy. MQ also improved blastocyst development potential of oxidatively stressed oocytes.