Zinc oxide and selenium nanoparticles can improve semen quality and heat shock protein expression in cryopreserved goat (Capra hircus) spermatozoa.

BACKGROUND: Reactive oxygen species (ROS) are strongly linked with oxidative stress (OS) generated during the process of sperm cryopreservation. Indeed, cellular damage from ROS has been implicated during sperm cryopreservation which causes deterioration in sperm quality and antioxidant nanoparticles (NPs) have been successful in preventing such damage. The interaction of NPs with sperm cells has been less frequently explored in farm animals. OBJECTIVE: The present study explored the effect of NP supplementation on sperm ultrastructure, potential interaction with sperm membrane (plasma and acrosome membrane), heat shock protein (HSP) gene expression levels and sperm quality in cryopreserved buck semen. MATERIALS AND METHODS: Thirty-two (32) ejaculates were collected from four (4) adult male bucks and then diluted in Tris- citric acid- fructose- egg yolk (TCFY) extender containing the Zinc-oxide (ZnO) and Selenium (Se) NP treatments (T0: Control; TZn: 0.1 mg/mL ZnO NPs and TSe: 1 µg/mL Se NPs) after initial evaluation. Diluted semen was packed in 0.25 mL French mini straws and then stored in liquid nitrogen (LN2). Sperm parameters, lipid peroxidation (LPO) profile, sperm head morphology ultrastructural classification under transmission electron microscope (TEM), potential interaction of NPs with sperm membrane and expression of HSP genes were evaluated in the different treatment groups. RESULTS: We found a significant (p < 0.05) increase in the percentage of spermatozoa with intact plasma membrane, and intact acrosome in the ZnO (0.1 mg/mL) and Se (1 µg/mL) NP supplemented groups in comparison to the frozen control group. TEM assessment revealed no internalization of both ZnO and Se NPs into the sperm structure. Few occasional contacts of ZnO NPs with the sperm membrane and a few agglomerates of Se NPs around the area of damaged membranes were visualized. HSP70 and HSP90 mRNA levels were significantly (p < 0.001) higher in the NP supplemented groups in comparison to the control. HSP70 and HSP90 mRNA levels had a strong positive association with sperm motility and a weak to moderate association with other sperm parameters. CONCLUSIONS: Current findings indicated that ZnO NPs are more potent than Se NPs in ameliorating peroxidative damages during sperm cryopreservation, increases semen quality parameters possibly by increasing the expression levels of HSP genes in buck semen. Furthermore, NP supplementation may have a potential role in preserving sperm head ultrastructure by acting as an antioxidant and reducing OS during various degrees of cellular insults, which needs to be further explored.

In Vitro and In Vivo Studies on the Efficacy of Zinc-Oxide and Selenium Nanoparticle in Cryopreserved Goat (Capra hircus) Spermatozoa.

Different nanoparticles (NPs) are currently being investigated for their potential role as cryoprotectant during semen cryopreservation in several mammalian species. It may be possible to improve semen quality following cryopreservation by supplementation of NPs in the freezing extenders. The present study was carried out in semen collected from four (4) Assam Hill Goat bucks (10 ejaculates per buck) to investigate the effect of supplementing zinc oxide (ZnO) and selenium (Se) NPs in Tris-citric acid-fructose yolk (TCFY) extender on in vitro sperm quality and in vivo fertility rate after freeze-thawing. The size morphology and zeta potential of ZnO and Se NPs were evaluated prior to its incorporation in the freezing extender. Qualified semen samples (> 70% progressive motility) were divided into five (5) aliquots and then diluted in TCFY extender containing ZnO and Se NP supplementation at different concentrations (T0, control; T1, 0.1 mg/mL ZnO NPs; T2, 0.5 mg/mL ZnO NPs; T3, 0.5 µg/mL Se NPs; and T4, 1 µg/mL Se NPs). Diluted semen was packed in 0.25 mL straws and then stored in liquid nitrogen. After thawing, post-thaw in vitro sperm attributes were evaluated. Finally, the effect of NPs on in vivo fertility rate was checked in heat-synched does (n = 70) by artificial insemination (AI) using straws that showed superior results during the in vitro study. Results showed that ZnO and Se NPs were poly-crystalline in nature with particle size below 100 nm (nm). The evaluated post-thaw sperm in vitro attributes were significantly (p < 0.001) higher in T1 in comparison to T0. The antioxidant enzyme activities were significantly (p < 0.001) higher in T1. Lipid peroxidation (LPO) profile was significantly (p < 0.001) lower in T1. Sperm motility and mitochondrial membrane potential (MMP) had a highly significant (r = 0.580, p < 0.05) association in T1. No significant (p > 0.05) differences in pregnancy rates were recorded after AI in the different treatments. In conclusion, extender supplemented with 0.1 mg/mL ZnO NPs improved post-thaw semen quality of goat spermatozoa consequently by increasing activities of endogenous antioxidant enzymes thereby lowering LPO levels. However, improved in vitro outcomes might not correspond to improved field fertility outcomes.

Effect of selenium nanoparticles on the quality and fertility of short-term preserved boar semen.

This study was carried out to investigate the effect of different concentrations of selenium nanoparticles (Se-NPs) in the Beltsville Thawing Solution (BTS) extender on the semen quality and fertility of Hampshire crossbred pigs. For the study, semen was collected from four boars (10 ejaculates/boar) by the gloved hand method. Each ejaculate was extended @ 1:2 with the BTS extender and split into four aliquots. The control (C) samples were without the supplementation of Se-NPs, whereas the other three were supplemented with 0.5 (T1), 1 (T2), and 2 µg ml-1 of Se-NPs (T3) and stored at 15°C in a BOD incubator. Extended semen was evaluated at 0 (immediately after dilution), 24, 48, 72, and 96 h of storage for sperm motility, live sperm, plasma membrane integrity, acrosome integrity, DNA integrity, and mitochondrial membrane potential (MMP). The mean percentage of sperm motility, live sperm, and sperm with intact plasma membrane and acrosome, and MMPs were significantly (p < 0.01) higher in all treated groups in comparison to control at 24, 48, 72, and 96 h of storage. Sperm with intact DNA in all treated groups increased significantly at 48 (p < 0.05), and 72 and 96 (p < 0.01) h of storage in comparison to the control group. The concentration of 1 µg ml-1 of Se-NPs was found to be the best among other concentrations. In each group, 10 sows were artificially inseminated with the liquid semen preserved for 72 h at 15°C. Supplementation of 1 µg ml-1 of Se-NPs yielded the highest conception rate in comparison to other groups. In conclusion, supplementation of 1 µg ml-1 of Se-NPs in the BTS extender resulted in the best semen quality and conception rate during the short-time liquid preservation of boar semen.