Effect of antioxidant supplementation on pig and horse gamete storage

According to recent studies, antioxidant supplementation on gamete processing and/or storage solutions improvesgamete quality parameters, after cooling or storage at sub zero temperature. The aim of the present study was to investigate the effects of antioxidant supplementation on pig and horse gamete storage. The first study aimed to determine the effects of resveratrol (RESV) on the apoptotic status of porcine oocytes vitrified by Cryotop method, evaluating phosphatidylserine (PS) exteriorization and caspases activation. RESV(2µM) was added during: IVM (A); 2 h post-warming incubation (B); vitrification/warming and 2 h post-warming incubation (C); all previous phases (D). The obtained data demonstrate that RESV supplementation in the various steps of IVM and vitrification/warming procedure can modulate the apoptotic process, improving the resistance of porcine oocytes to cryopreservation-induced damage. In the second work different concentrations of RESV (10, 20, 40, and 80µM) were added during liquid storage of stallion sperm for 24 hours at either 10°C or 4°C, under anaerobic conditions. Our findings demonstrate that RESV supplementation does not enhance sperm quality of stallion semen after 24 hours of storage. Moreover, the highest RESV concentrations tested (40 and 80µM) could damage sperm functional status, probably acting as pro-oxidant. Finally, in the third work other two antioxidants, ascorbic acid (AA) (100 µM) and glutathione (GSH) (5mM) were added on boar freezing and/or thawing solutions. In our study different sperm parameters were evaluated before freezing and at 30 and 240 minutes after thawing. Our results showed that GSH and AA significantly improved boar sperm cryotolerance, especially when supplemented together to both freezing and thawing media. This improvement was observed in sperm viability and acrosome integrity, sperm motility, and nucleoprotein structure. Although ROS levels were not much increased by freeze-thawing procedures, the addition of GSH and AA to both freezing and thawing extenders significantly decreased intracellular peroxide levels.

Effects of glyphosate and Roundup upon mammalian gametes

The wide use of glyphosate-based herbicides (GBHs) has become a controversial issue due to the potential harmful effects on human health. Commercial formulations, among which Roundup is the most famous one, contain a number of adjuvants inside; most of these are patented and not publicly known, therefore, they can act differently from glyphosate alone and might strengthen its toxic effect. Our study is focused on GBHs reproductive toxicity with a special regard to glyphosate and Roundup impact on male and female mammalian gametes after exposure to concentrations ranging from the one recommended for agricultural use (0.1% Roundup, containing 360 µg/mL glyphosate) to 70-fold lower or more. Sperm quality analysis, either on boar and stallion, showed that Roundup has much more detrimental impact than glyphosate at equivalent concentrations on spermatozoa function and survival. Basing on our results, the toxic effect of these pesticides on spermatozoa may be linked to an impairment in mitochondrial activity and a subsequent decrease in ATP production and/or alterations in the redox balance, which impact cell motility and plasma membrane stability. Moreover, a different species sensitivity to GBHs may exists as high doses of glyphosate affected sperm quality only in boar and not in stallion; furthermore, Roundup had deleterious effects at lower doses in the first compared to the latter. With regard to female gametes, we found that glyphosate and Roundup exposure during IVM detrimentally affect the subsequent developmental ability of swine embryos, providing further evidence of their potential toxic effect on female reproductive system. In addition, Roundup altered steroidogenesis and increased oocyte ROS levels. Therefore, according to our results, we can conclude that GBHs exert a negative impact on both male and female gametes and that Roundup adjuvants enhance glyphosate toxic effects and/or are biologically active in their side-effect.