Vliv gasotransmiterů na kvalitu a mrazitelnost kančích spermií.

Mgr. Marija Jovičić

Doctoral thesis

Abstract:

The successful reproductive biotechnology and assisted reproduction methods also depends on the quality of semen. During collection and dilution of boar semen is important to do it carefully and properly to maintain its quality during storage. Boar sperm contains high concentrations of polyunsaturated fatty acids which are very susceptible to lipid peroxidation and serve as prefered substrates of reactive oxygen species. Oxidative stress is important factor which decreases the quality of semen. Using chemicals which have antioxidant properties is useful to decrease oxidative stress. It is known that gasotransmitters such as nitric oxide (NO) and hydrogen sulphide (H2S) have a role in physiology. NO has dual effect, low concentration mostly has positive effect, high concentrations negative. In this study we evaluate the effect of donor NO, sodium nitroprusside (SNP) and Nω-Nitro-L-arginine methyl ester hydrochloride (L-NAME) or aminoguanidine (AG) as non-selective and selective nitric oxide synthase (NOS) inhibitors, respectively. The aim of this study was to evaluate the effect of NO on boar sperm motility, membrane integrity and acrosomal status during 72 h of semen storage at 17ºC. The effect on non-selective nitric oxide synthase (NOS) inhibitor L-NAME on boar semen stored at 17 °C in the concentration 0.625 mM did not show effect on quality, in contrast higher concentrations showed negative effect. This results suggests that low NO levels are necessary for boar sperm physiology. A significant increase of the amplitude of lateral sperm head displacement (ALH), and both curvilinear and straight-line velocity (VCL and VSL, respectively) was observed at 72 h of semen storage in samples treated with 0.625 mM selective AG, probably because of the antioxidant properties of this selective NOS inhibitor.Concerning NO donor, we found that SNP concentrations from 18.75 till 150 μM had mostly no or only negative effects on boar sperm parameters during semen storage.We also evaluated the in vitro effect of NOS inhibitor AG on boar sperm cells under induced oxidative stress. Our results demonstrate that AG mostly abolishes the deleterious effects of oxidative stress in vitro in boar spermatozoa. In virtue of its antioxidant capacities, AG preserves the boar sperm motility, reduces the lipid peroxidation, protects the plasma membrane and acrosome integrity under oxidative stress. Interestingly, AG 1 mM mostly  eliminates the negative effects of oxidative stress given that, except for the lipid peroxidation, all sperm parameters did not differ or even showed better performance than those of control samples without the reactive oxygen species (ROS) generating system.It is known that H2S has role in many diseases, but his role in sperm physiology still is not known. That´s way we evaluated effect of H2S donors on boar semen under induced oxidative stress. The aimed of this study was to determine the total antioxidant capacities of sodium sulfide (Na2S) and morpholin-4-ium 4-methoxyphenyl (morpholino) phosphinodithioate (GYY4137), which are fast- and slow-releasing H2S donors, respectively, and to test whether H2S donors are able to protect spermatozoa against oxidative stress. We used slow releaser based on his rate of H2S release, GYY4137, and fast releaser Na2S. Within the range of concentration tested (3-300µM), GYY413 showed positive effect on boar sperm motility, whereas Na2S was beneficial at the lowest concentration but detrimental at the highest. It is interesting that this two donors showed different effects on boar semen. Moreover both H2S donors preserve sperm motility and reduce acrosome loss, although the effects were both dose and donor dependent.

Keywords

Boar spermatozoa, Nitric oxide, Nitric oxide synthases, Oxidative stress, Hydrogen sulphide, Gasotransmitters, Sperm kinetics, Cryopreservation, Antioxidants

Language used: English

Date on which the thesis was submitted / produced: 25. 5. 2020

Thesis defence

• Supervisor: prof. Ing. Mgr. Markéta Sedmíková, Ph.D.

Czech University of Life Sciences Prague

Faculty of Agrobiology, Food and Natural Resources

Doctoral programme / field:
Animal Science / General Animal Science