Effects of Ochratoxin A on nuclear and cytoplasmic maturation of oocytes obtained from prepubertal lambs

OchratoxinA (OTA) is a major mycotoxin produced by several species of Aspergillus and Penicillium fungi and has been reported as an ubiquitous natural contaminant found in food and feed[1].OTA plays reprotoxic, embryotoxic and teratogenic as well as nephrotoxic, neurotoxic, immunotoxic and carcinogenic activity as reported in either laboratory or farm animals [2]. Major mechanisms of action include inhibition of protein synthesis,toxic effect on mitochondrial (mt) function and calcium homeostasis with consequent oxidative stress, apoptosis induction and DNA adduct formation [2]. The aim of the present study was to evaluate the effects of OTA on nuclear and cytoplasmic maturation of oocytes from pre-pubertal lambs.Cumulus-oocyte complexes (COCs) were recovered at local slaughterhouses from the ovaries of prepubertal lambs (less than 6 months of age). During in vitro maturation (IVM) [3], COCs were exposed to 10 μM OTA, a concentration reported as effectivein a previous study in the mouse[4]. Control conditions were: vehicle controls (IVM medium with 1% methanol) and standard controls (IVM medium without vehicle). After IVM and the removal of cumulus cells, the oocytes were stained with MitoTracker Orange CMTM Ros, 2′,7′-dichlorodihydrofluorescein diacetate and Hoechst 33258 and fixed in 2% paraformaldehyde solution in PBS. Metaphase II oocytes were analyzed by laser scanning confocal microscopy for assessing their cytoplasmic maturation indicated by mt distribution pattern [2]. Data were analysed by Chi-square test and differences were statistically significant when P<0.05. A total of 218 oocytes were analyzed. Lack of vehicle-related effects was noticed (23/37, 62% vs 62/96, 65%, for oocytes cultured with or without the vehicle; P> 0.05). OTA caused a slight (P>0.05) reduction of the maturation rate (39/85, 46% vs 23/37, 62%, for exposed and controls, respectively). Instead, it affected oocyte bioenergetic status, as it reduced the rate of oocytes showing healthy perinuclear/pericortical mitochondrial distribution pattern (4/39, 10% vs 9/23, 39%, P<0.05). These data indicate that OTA, in the exposure conditions used in the present study,hinders nuclear and cytoplasmic maturation in prepubertal lamb oocytes