Influence of Various Antioxidants on Microscopic-Oxidative Stress Indicators and Fertilizing Ability of Frozen-Thawed Bull Semen

Abstract

Cryopreservation is associated with the production of reactive oxygen substances (ROS), which lead to lipid peroxidation of sperm membranes, resulting in a loss of sperm motility, viability and fertility. The aim of this study was to determine effects of the antioxidants of oxidized glutathione (GSSG), reduced glutathione (GSH) and bovine serum albumin (BSA) on standard semen indicators (motility, acrosome and total abnormalities, HOST), endogenous antioxidant enzyme activities and fertilizing ability of frozen-thawed bull semen.Eighteen ejaculates from each of 3 Holstein bulls were collected using an artificial vagina and 9 replicates of the ejaculates were diluted with a Bioxcell ® -based extender supplemented with antioxidants, including BSA (5 mg/ml), GSH (2 mM), GGSG (2 mM), and an extender containing no antioxidants (control). Insemination doses (1.5 × 10 7 sperm/0.25 ml straw) were prepared for the insemination of cows at observed oestrus. Supplementation with antioxidants led to lower percentages of acrosome damage (4.0 ± 0.5%, 4.4 ± 0.5%, 4.0 ± 0.3%, respectively) and total abnormalities (10.3 ± 0.7%, 9.7 ± 0.8%, 10.4 ± 0.6%), compared to the controls (6.5 ± 0.6 and 14.9 ± 1.1% P < 0.01). Pregnancy rate after insemination was highest (72.2%) in the group which was given BSA (P < 0.05). There were no significant differences among groups in GSH and glutathione peroxidase (GSH-PX) enzyme activities. Superoxide dismutase (SOD) activities (0.05 ± 0.005, 0.03 ± 0.005, 0.05 ± 0.009 μkat/g protein, respectively) in all of the experimental groups with antioxidants were lower than the control group (0.11 ± 0.024 μkat/g protein, P < 0.001). Furthermore, BSA increased (P < 0.001) the activity of catalase (CAT, 304.23 ± 114.69 μkat/g protein), following the freezing-thawing process. Bull semen, freezing, artificial insemination, antioxidant activities, lipid peroxidationSemen cryopreservation has allowed specific opportunities for the conservation and widespread dissemination of valuable genetic resources through sperm banks, the guarantee of a constant commercial supply of semen, and collaboration in breed improvement programs by means of artificial insemination (AI) (Holt et al. 1997). Some of the problems encountered using AI in pigs, sheep and exotic species include decreased motility, membrane integrity and fertilizing potential of spermatozoa during cryopreservation, premature activation in the female genital tract, and altered membrane responses to physiological stimuli of sperm that survive the freezing-thawing process (Viswanath and Shannon 2000). It is well known that phospholipids in the sperm plasma membrane undergo peroxidation, which results in the formation of reactive oxygen substances (ROS, namely, superoxide anion radical, hydrogen peroxide) and lipid hydroperoxides (Alvarez and Storey 2005). Excessive generation of ROS by immature and abnormal spermatozoa, and sperm processing (e.g., extending, freezingthawing process), accompanied by low scavenging and antioxidant concentrations in semina...

Improving cryopreservation capacity of ram spermatozoa by supplementing the diluent with melatonin

“…Additionally, the percentages of post-thaw primary and secondary sperm abnormalities were 2.6 and 3.6 higher in the control group than those of MelatoninHD group, respectively. These results are in conformity with those reported previously in different species (Sariözkan et al 2009;El-Raey et al, 2014).…”

“…This, in turn, induces sperm premature activation in the female genital tract and altered membrane responses to physiological stimuli of sperm that survive the freezing-thawing process (Viswanath and Shannon 2000) and , consequently, a loss of sperm motility, viability and fertility (Sariözkan et al 2009). Furthermore, extending and freezing-thawing of semen decrease the antioxidant capacity of semen (Sariözkan et al 2009). …”

“…In addition to its multiple actions on different physiological processes, melatonin and its metabolites are considered powerful antioxidants due to their ability to scavenge excessive ROS and, thus, protect the spermatozoa (Reiter and Tan, 2003;Ahn and Bae, 2004;Adriaens et al, 2006;Kang et al, 2009). Therefore, melatonin supplementation was reported to counteract the drastic effects of lipid peroxidation on cryopreserved semen parameters in different species (Sariözkan et al 2009;El-Raey et al, 2014). …”

Positive effect of BSA supplemented soybean lecithin based extender on liquid storage of ram semen at 5C

“…On the other hand, main disadvantages of using an animal-derived material in semen extender can be listed as: failure to prevent individual differences (lack of homogeneity), risk of contamination and transmission of diseases (16,38). The low-density lipoprotein fraction such as lecithin (35,38) or multifunctional macromolecules such as bovine serum albumin (BSA) (19,31,33) could be substitutes for egg yolk in a semen extender.…”

“…It is known that, in addition to antioxidant capacity, BSA accelerates the motility (36,37), protects the membrane (36) and DNA integrity (33), increases the survivability of spermatozoa, and induces the capacitation (40) and acrosome reaction (26). As a consequence of these, BSA supplemented extenders are used for cryopreservation or liquid storage of bull (37), ram (35), goat (6,7), stallion (9), buffalo (2), rabbit (34) and turkey (8) semen.…”