RESUMEN
Introduction: Embryo cryopreservation is a valuable technique used for preserving genetic resources for long periods. However, the survival rate of embryos is dependent on the method used. Therefore, in this study, we evaluated the efficiency of slow freezing method but with an additional dehydration step prior to freezing to overcome the formation of ice crystals. Methods: Oocytes collected from the ovaries of native Korean cattle subjected to in vitro fertilization were cultured for 7 days until the formation of expanded blastocysts. Before freezing, the blastocysts were placed in four pre-equilibration media: a control medium with no addition of sucrose, and three experimental media with the addition of 0.1, 0.25, and 0.5 M sucrose, respectively. Then, the pre-equilibrated embryos were frozen. Embryo survival and hatching rates were evaluated morphologically at 24, 48, and 72 h after thawing. Immunofluorescence staining, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, and gene expression analysis of the re-expanded blastocytes were examined 24 h after freeze-thawing. Results: The survival rate was significantly higher in the 0.1 M group than in the control group (p < 0.05), and the hatching rate at 72 h was significantly higher in the 0.25 and 0.5 M groups than in the control group (p < 0.05). TUNEL-positive cells were significantly lower in the 0.25 M group than in the control group (12.5 ± 0.9 vs. 8.3 ± 0.8; p < 0.05). The gene expression of BCL2 associated X, heat shock protein 70 kDa, and aquaporin 3 in the 0.25 M group was significantly lower than that in the control group (p < 0.05). Conclusion: Our study revealed that treatment with 0.25 M sucrose before slow freezing improved the viability of bovine embryos after freeze-thawing.
RESUMEN
The antimicrobial activity of lysosomes, a cell organelle, against a range of test microorganisms was examined in this study. The lysosomes isolated from Saccharomyces cerevisiae showed antimicrobial activity to Escherichia coli that positively correlated with the pH of the phosphate buffer as a dissolving solvent. The lysosomes from S. cerevisiae exhibited optimal activity at a concentration of 40%, at pH 4.0 of phosphate buffer, and at broad range temperature, except of over 50 degrees C. It was also found that the lysosomes have antimicrobial activity against seven different microorganisms including E. coli. In addition, S. cerevisiae were exposed by a treatment with H(2)O(2) and lysosomes were isolated from H(2)O(2) exposed S. cerevisiae. We found that fluorescent intensities of each isolated lysosomes were increased depending on the increment of treated H(2)O(2) concentration, and the lysosomes from 20 mM H(2)O(2) treated S. cerevisiae showed higher antimicrobial activity than those from normal S. cerevisiae. Therefore, it suggests that lysosomes isolated from S. cerevisiae can be used as an antimicrobial agent. In addition, lysosomes activated by H(2)O(2) enhanced its antimicrobial activity.
