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Cryopreservation of rooster semen is a reproductive technology carried out to boost genetic gain and productivity in commercial flocks of chicken. However, semen freezing significantly reduces the quality and fertilizing potential of spermatozoa. This study examined cryoprotective effects of the mitochondria-targeted antioxidant mitoquinol mesylate added to the freezing extender by assessing post-thaw characteristics of rooster sperm. Semen samples were diluted in the Beltsville extender supplemented with 0, 1, 10, 100 or 1000 nM of mitoquinol mesylate. Following the thawing of cryopreserved semen doses, we evaluated the following sperm parameters: motility and morphology, membrane integrity and mitochondrial function, acrosome integrity, apoptosis status, lipid peroxidation, DNA fragmentation, reactive oxygen species (ROS) accumulation, epigenetic patterns (DNA methylation and histone modifications), and fertilizing ability. Semen diluted in extenders containing 10 or 100 nM of mitoquinol mesylate significantly exceeded all other groups of frozen-thawed semen samples in sperm motility, average path velocity as well as membrane/acrosome integrity, mitochondrial function indices and DNA/histone modification. Moreover, the addition of 10 and 100 nM of mitoquinol mesylate significantly reduced lipid peroxidation, apoptosis rate, DNA fragmentation and ROS concentrations compared with all other dilutions and was associated with a higher (P ≤ 0.05) fertilization rate compared with a non-supplemented control group, albeit it was still significantly lower compared with that obtained using fresh semen. It can be, however, concluded that mitoquinol mesylate significantly improved an array of quality parameters and fertilizing potential of rooster semen, and hence can be recommended for use as a diluent additive in semen cryopreservation procedures employed in commercial poultry operations., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to disclose., (Copyright © 2025 Elsevier Inc. All rights reserved.)

Mammalian genome editing has utilized expensive and highly specialized electroporator devices. The "Gene Pulser XCell," a modular electroporation system for transfecting all cell types, has not been used extensively in mammalian embryo genome editing. The present experiment was undertaken to determine the usefulness of the Gene Pulser XCell for inserting the CRISPR/Cas9 system into intact zygotes in order to obtain the enhanced green fluorescent protein reporter rats (eGFP-R). An electroporation pulse response test using mCherry mRNA was performed to optimize the settings of the electroporator. Forty-five combinations of five pulse voltages (15, 25, 30, 35 and 40 V), three pulse durations (5, 10 and 25 ms), and three pulse frequencies (2, 5 and 6 pulses) applied at a constant 100-ms pulse interval and temperature of 37.5 °C were evaluated. The test revealed that the 35 V was the only voltage suitable for insertion of mCherry mRNA into intact rat zygotes and the only one that resulted in the production of embryos attaining the blastocyst stage. The incorporation of mCherry mRNA increased but the survival of the electroporated embryos declined with an increment in the number of pulses. Subsequent transfer of 1112 surviving Sprague Dawley rat embryos (after 8 h of incubating 1800 zygotes electroporated with the CRISPR/Cas9) resulted in the production of 287 offspring (25.8%). Ensuing PCR and phenotypic evaluation confirmed that twenty animals (6.96%) expressed eGFP in all body organs/tissues except for blood and blood vessels. The mortality of males and females before the attainment of puberty was 2 and 3 pups, respectively, and the final number/ratio of male to female of offspring was 9:11. All the surviving rats mated naturally and successfully transmitted the GFP transgene to their progeny. The Gene Pulser XCell total system with the settings predetermined in the present experiment can effectively be used to produce transgenic rats through the CRISPR/Cas9-mediated genome editing of zygotes., (Copyright © 2023 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

Semen cryopreservation can provide genetic resources for a large number of females from a small number of superior males. Optimization of cryopreservation media to achieve the highest quality of post-thaw semen is crucial. Soybean lecithin has evaluated as a plant-based cryoprotectant for substitution of egg yolk in ram semen extender. Flow cytometric and fertility assessments were applied following cryopreservation procedure in two experimental groups (SL group: extender containing 1% w/v soybean lecithin and EY group: extender containing 20% v/v egg yolk). The higher percentage of live sperm and the lower percentage of dead sperm were obtained in SL (47.66 ± 1.38, 52.33 ± 1.69, respectively) extender compared to EY (41.16 ± 1.38, 58.83 ± 1.69). For motion characteristics, plasma membrane integrity, acrosome integrity and mitochondria activity, no significant difference was observed between SL and EY extenders. In artificial insemination experiment, there was no significant difference in pregnancy rate, lambing rate and twining rate between SL and EY extenders. It can be concluded that SL extender can be an efficient alternative extender to preserve ram sperm during cryopreservation procedure without adverse effects., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Hepatitis C virus (HCV) is an important cause of acute and chronic hepatitis which is a disorder with a high worldwide prevalence. HCV core protein was considered as immunogenic counterpart of the HCV vaccine and it is an ideal candidate for HCV vaccine. Since cytokines such as IL-6, TNF-alpha and IFN-Gamma are responsible for the prevention of viral infection, this study aimed to evaluate the effectiveness of HCV core protein as a vaccine. Ten BALB/c mice were immunized with HCV core protein and after 42 days the splenocytes were isolated and the IL-6 and INF-gamma secretion were measured using ELISpot technique, at the same time TNF-alpha was determined by ELISA in the sera. The MTT assay was done to assess the viability of the cultured splenocytes. For evaluating the humoral immune response against the recombinant HCV core protein the DOT Blot test was used. Data was compared using one-way ANOVA test and significant results were considered at p < 0.05. In the present study the IL-6, INF-gamma and TNF-alpha levels were dramatically higher in the immunized mice compared to the control group (respectively, 22.9 +/- 1.26; 18.53 +/- 3.87; 53.96 +/- 4.54 and p < 0.05). The immunized mice with recombinant HCV core protein showed higher amount of IL-6, INF-gamma and TNF-alpha in the current study. Since the level of IL-6, TNF-alpha and IFN-gamma is high in patients with acute HCV infection, thus a vaccine which could stimulate the secretion of these cytokines in advance may have a preventive role.