Dithiothreitol prevents age-associated decrease in oocyte/conceptus viability in vitro

The present study was designed to ascertain whether the negative effects onreproductive potential of post-ovulatory ageing in vitro of oocytes can beprevented by antioxidant therapy. Mouse metaphase II (MII) oocytes wereaged in vitro for 12 h prior to insemination in the presence of varyingconcentrations of L-ascorbic acid, 6-methoxy-2,5,7,8-tetramethylchromane-2-carboxylic acid (Trolox), L-cystinedihydrochloride, ethylenediaminetetraacetic acid (EDTA), beta-mercaptoethanol and DL-dithiothreitol (DTT). In-vitro ageing of oocytes wasassociated with lower fertilization rate, higher proportion of conceptiexhibiting cellular fragmentation at 24 h post-insemination and lowerpercentage of concepti reaching the blastocyst stage. Ascorbic acid, Troloxand EDTA had no effect on cellular fragmentation or potential of oocytesfor development. However, the probability of an oocyte reaching theblastocyst stage was decreased (P < or = or = 0.05) in oocytes incubatedin the presence of L-cystine (50 and 500 microM) and beta-mercaptoethanol(5, 50 and 500 microM) when compared to control aged oocytes.Age-associated cellular fragmentation at 24 h post-insemination waspartially prevented (P < or = 0.05) by incubating oocytes in thepresence of beta-mercaptoethanol (500 microM). DTT (50 and 500 microM)increased (P < or = 0.05) fertilization rate and number of cells at 81 hpost-insemination to levels similar to those exhibited by control oocytes.Furthermore, both age-associated fragmentation at 24 h post-insemination (P< or = 0.05) and decreased potential of oocytes for development to theblastocyst stage (P < or = 0.05) were prevented, at least in part, byculturing oocytes in the presence of DTT (50 microM). Although themechanism by which DTT exerts its beneficial effects on aged oocytesremains to be elucidated, it may protect oocytes by preventing oxidation offree thiol groups and/or altering a redox-independent signalling pathwaythat mediates cellular fragmentation and death.