Effect of genistein on semen quality, antioxidant capacity, caspase-3 expression and DNA integrity in cryopreserved ram spermatozoa

Abstract

The present study aimed to assess the impacts of different concentrations of genistein on freezability and functional integrity of ram spermatozoa. Five ejaculates were collected from each ram (n = 6) twice weekly. Different levels of genistein (0, 1, 5, 10 and 100μM) were added to tris-egg yolk based extender (TBE) for extension and cryopreservation of semen. After thawing, samples were assessed for progressive forward motility (PFM), viability index, acrosomal and plasma membrane integrity. Furthermore, total antioxidant capacity (TAC), lipid peroxidation expressed as malondialdehyde (MDA), caspase-3 mRNA expression and comet assay for DNA integrity were estimated. PFM significantly (P < 0.05) lowered in 100 μM genistein at immediate, one and two hours post-thawing. However, post-thawing PFM at three hours significantly elevated (P < 0.05) in 10μM genistein concentration compared to that in control TBE and 100 μM genistein. Addition of 10μM genistein to TBE significantly (P < 0.05) upgraded viability index, acrosome integrity and plasma membrane integrity as compared to those in other treatments. Levels of TAC significantly (P<0.05) promoted in genistein (5, 10 and 100 μM). MDA significantly (P < 0.05) lowered in genistein (10 and 100 μM) than in control and 1 μM genistein. Caspase-3 expression revealed a significant (P < 0.001) fold (0.53 ± 0.09) decrease in semen supplemented by 10 μM genistein as compared to other treatments. DNA% in comet tail significantly promoted in 5 and 10 μM genistein groups. In conclusion, adding 10μM genistein to TBE ameliorated the criteria of cryopreserved ram semen as well as down regulated caspase-3 expression with maximal reduction in DNA fragmentation.