RESUMEN
Cryopreservation has adverse effects on the post-thaw sperm quality due to oxidative stress and the presence of bacteria. To minimize such effects, plant extracts have been included in the composition of the semen diluents. The objective of the present study was to evaluate the antimicrobial and antioxidant effects of Moringa oleifera seed extract (MOSE) on cryopreserved ram semen, as well as its impact on in vitro fertilization. Semen from six hair rams was treated with five treatments before cryopreservation: Control (without any antibiotic), Standard (conventional antibiotic), 1.0, 10.0, and 50.0 mg/ml of MOSE. Post-thawing sperm characteristics were evaluated by the computer-assisted semen analysis. Antimicrobial activity was assessed by counting colony-forming units (CFU) and the antioxidant capacity by the ferric reducing antioxidant power method. A heterologous in vitro fertilization technique was implemented to measure the fertilization rate. Progressive and rapid motility, membrane and acrosome integrity, and active mitochondria were higher (p < .05) in the 10.0 mg/ml treatment compared with Standard after thawing. All M. oleifera treatments showed inhibition of CFU. The antioxidant capacity of M. oleifera seed extract was higher in the 10.0 and 50.0 mg/ml treatments. Fertilization rate (cleavage percentage) was higher (p < .05) in the 10.0 mg/ml (82.9 ± 10.0) and Control (82.5 ± 9.9) treatments compared with Standard (73.7 ± 9.1). The addition of 10.0 mg/ml of MOSE to ram semen inhibits the development of microorganisms and improves sperm characteristics and the in vitro fertility of the semen.
Asunto(s)
Moringa oleifera , Preservación de Semen , Masculino , Ovinos , Animales , Preservación de Semen/veterinaria , Preservación de Semen/métodos , Motilidad Espermática , Antioxidantes/farmacología , Semillas , Criopreservación/veterinaria , Criopreservación/métodos , Espermatozoides/fisiología , Fertilización In Vitro/veterinaria , Extractos Vegetales/farmacología , Antibacterianos/farmacología , Crioprotectores/farmacologíaRESUMEN
Meiotic recombination is a critical process for sexually reproducing organisms. This exchange of genetic information between homologous chromosomes during meiosis is important not only because it generates genetic diversity, but also because it is often required for proper chromosome segregation. Consequently, the frequency and distribution of crossovers are tightly controlled to ensure fertility and offspring viability. However, in many systems, it has been shown that environmental factors can alter the frequency of crossover events. Two studies in flies and yeast point to nutritional status affecting the frequency of crossing over. However, this question remains unexplored in mammals. Here, we test how crossover frequency varies in response to diet in Mus musculus males. We use immunohistochemistry to estimate crossover frequency in multiple genotypes under two diet treatments. Our results indicate that while crossover frequency was unaffected by diet in some strains, other strains were sensitive even to small composition changes between two common laboratory chows. Therefore, recombination is both resistant and sensitive to certain dietary changes in a strain-dependent manner and, hence, this response is genetically determined. Our study is the first to report a nutrition effect on genome-wide levels of recombination. Moreover, our work highlights the importance of controlling diet in recombination studies and may point to diet as a potential source of variability among studies, which is relevant for reproducibility.
Asunto(s)
MeiosisRESUMEN
Capacitation-like changes affect sperm of several species, such as ram, reducing cell survival and fertilizing competence. Proteins from seminal plasma stabilize sperm plasma membranes, being an interesting focus to develop strategies for improving cryopreserved ram semen performance. To date, biotechnologies are focused to reduce damage in frozen-thawed ram spermatozoa through the addition of bioactives. Serine Protease Inhibitor Kazal-type 3 (SPINK3) is a little protein synthesized by mouse seminal vesicle and secreted to seminal plasma. While attached to the sperm, this protein binds to non-capacitated sperm and blocks calcium entry, avoiding a premature physiological capacitation and consequently, acrosome reaction. Due to these characteristics, SPINK3 has been proposed as a decapacitating factor. The aim of this work was to assess whether heterologous SPINK3 is able to protect ram sperm from the well-known cell damages produced by freezing/thawing and to understand the mechanisms by which it is acting. Sperm were supplemented with 13 µM SPINK3 before freezing in an egg yolk-based extender or after thawing and selection. Under both conditions, SPINK3 decreased intracellular calcium content (p < 0.05) and reduced the 25 kDa tyrosine phosphorylated protein demonstrating a decapacitating effect, although the addition of the protein before cryopreservation was not enough to improve other sperm parameters. However, the addition of SPINK3 post thawing was able to significantly ameliorate viability, motility, mitochondrial status and to avoid the increase of lipid peroxidation (p < 0.05). Moreover, sperm treated with SPINK3 was not only still capable to fertilize, but also improved it, as evidenced by an increase in the oocyte cleavage rate (p < 0.05) although, the embryo development and embryo quality were not affected. Our findings would contribute to develop a strategy for improving sperm quality by using decapacitating proteins. In fact, the outcomes of this work demonstrate that SPINK3 is able to reduce sperm cryo-injuries when is added after thawing, improving functionality and thus in vitro fertilization results.