RESUMO
For years, compounds of natural origin have been the subject of extensive biomedical research due to very interesting, new ingredients potentially useful for various pharmaceutical, medical and industrial applications. The therapeutic properties and healing benefits of sea cucumbers may result from the presence of numerous, biologically active ingredients. Sperm subjected to processing and subsequent storage at low temperatures experience a number of damage, including the loss of the integrity of the cytoplasmic membrane, DNA and acrosome defragmentation. Therefore, the aim of this experiment was to investigate the cytoprotective potential of sea cucumber extract against cryopreserved sperm and semen fertility rate. Commercially available sea cucumber extract was taken from the cellulose shell, then 790 mg of powder was weighed out and placed in 3 glass tubes containing, respectively: 10 mL of water-glycerin solution (WG), water-ethanol (EC), glycerin-ethanol (GE), glycerin-DMSO (DG). Tubes were mixed with vortex for 3 min, then placed in a water bath and incubated for 16 h at 40 °C. Six simmental bulls, 3 years old, of known health status were used for the experiment. Semen was collected from each male once a week (for 18 weeks) using an artificial vagina. After an initial assessment of semen quality, the ejaculates were pooled to eliminate individual differences between males, then diluted to a final concentration of 80 × 106 sperm/mL with a commercial extender (Optixcell, IMV, L'Aigle, France) and divided into 16 equal samples. Control (C) without additive, the test samples contained 2, 4, 6, 8 and 10 µL WG, 2, 4, 6, 8 and 10 µL WE, 2, 4, 6, 8 and 10 µL GE, 2, 4, 6, 8 and 10 µL DG. Semen was frozen/thawed and assessed for motility, viability, DNA defragmentation, mitochondrial membrane potential and acrosome integrity. It was shown a positive effect of water-glycerin (WG) and glycerine-ethanol (GE) extracts on the efficiency of sperm preservation at low temperatures. Established that, depending on the type of prepared extract, the sea cucumber can have both cytoprotective (WG, GE, WE) and cytotoxic (DG) effects. Moreover, too high concentrations of the extract can adversely affect the sperm in terms of parameters such as viability, motility, mitochondrial potential, and the integrity of the acrosome or DNA of cells. The present study, thanks to the use of model animals to study the cytoprotective potential of the sea cucumber extract, proves that it can be a potential candidate for use in semen cryopreservation technology to improve the efficiency of storage at low temperatures. Further research is needed to optimize the composition of individual types of extracts and their effect on sperm. The highest effectiveness of female fertilization was observed when doses from GE groups (2 and 4) were used for insemination. The results of this analysis prove that the addition of the tested extract may improve the fertilization efficiency.
RESUMO
The aim of this study was to determine the concentration of proAKAP4 and other indicators of cryopotential of spermatozoa cryopreserved in extender with Holothuroidea extract addition. Nine Holstein Friesian bulls, 3.5 years old, of known health status, were used for the study. The animals were kept and fed equally. Semen was collected once a week using an artificial vagina. The commercially available Holothuroidea extract was used as a supplement to the commercial extender (0, 2, 4 and 6 µL/mL) before the freezing/thawing process. The viability, motility, motion parameters, and acrosome integrity of the sperm were analyzed with (test) or without (control) extract samples. Furthermore, the concentration of the proAKAP4 biomarker in frozen sperm was assessed. It was shown that the addition of 4 and 6 µL of the extract may have a positive effect on the quality parameters of the sperm after thawing. The results indicate that extender supplementation with the above extract modulates (increases) the concentration of proAKAP4 in sperm at all tested levels. Additionally, this indicator has become helpful in identifying sperm of poor biological quality. Moreover, it has been proven that the proAKAP4 biomarker can be successfully used to evaluate the effectiveness of the use of various extenders for semen cryopreservation.
