Introduction The addition of low-molecular-weight antioxidants during the freezing process improves post-thaw sperm quality. The high antioxidant potential of cryopreserved semen could have a positive effect on the motility, viability, and energy status of sperm cells and their ability to bind to the zona pellucida of oocytes. The aim of the study was to determine the effects of different concentrations and combinations of vitamins E and C in a semen extender on selected quality parameters of frozen-thawed canine spermatozoa. Material and Methods The experimental material was the semen of four mixed-breed dogs. Sperm viability (motility, plasma membrane integrity, and mitochondrial function) was examined at 0, 60, and 120 min in semen samples supplemented with the extender and in the controls. Results Combined supplementation with vitamins C + E at a concentration of 200 + 200 μM /1 × 10⁹ spermatozoa had the most profound effect on total sperm motility, linear motility, and the percentage of spermatozoa with intact plasma membrane and active mitochondria. Conclusion The synergistic activity of vitamins E and C had a more beneficial influence on the quality of frozen–thawed sperm than these non-enzymatic antioxidants applied separately.

Introduction: Nickel and iron are very commonly occurring metals. Nickel is used in industry, but nowadays it is also used in medical biomaterials. Iron is an element necessary for cell metabolism and is used in diet supplements and biomaterials, whence it may be released along with nickel. Material and Methods: BALB/3T3 and HepG2 cells were incubated with iron chloride or nickel chloride at concentrations ranging from 100 to 1,400 µM. The following mixtures were used: iron chloride 200 µM plus nickel chloride 1,000 µM, or iron chloride 1,000 µM plus nickel chloride 200 µM. The cell viability was determined with MTT, LHD, and NRU tests. Results: A decrease in cell viability was observed after incubating the BALB/3T3 and HepG2 cells with iron chloride or nickel chloride. A synergistic effect was observed after iron chloride 1,000 μM plus nickel chloride 200 μM treatment in all assays. Moreover, the same effect was observed in the pair iron chloride 200 μM plus nickel chloride 1,000 μM in the LDH and NRU assays. Conclusions: Iron (III) and nickel (II) decrease cell viability. Iron chloride at a concentration of 200 µM protects mitochondria from nickel chloride toxicity.

Inosine pranobex (Isoprinosine) stimulates cell-mediated immune responses to viral infections in humans and might have also therapeutic use in animals. The aim of this study was to compare three in vitro cytotoxicity assays on mouse embryo fibroblasts and liver cancer cells and determine their ability to detect early cytotoxic effects for inosine pranobex. BALB/3T3 clone A31and HepG2 cells were incubated with inosine pranobex at concentrations from 0.1 to 1,000 μg/mL. Cell viability was determined with the MTT reduction, the LHD release, and the NRU tests. A decrease in the cell viability was observed after incubating the BALB/3T3 clone A31and HepG2 cells with inosine pranobex. Based on the cytotoxicity endpoints measured in these investigations in BALB/3T3 clone A31cells, it can be concluded that the cell membrane may be the first part of the cell to be affected by inosine pranobex. The disintegration of lysosomes and mitochondria follows mitochondria damage. In HepG2 cells likewise, the cell membrane may be the first part of the cell to be affected by inosine pranobex. Also in liver cancer cells, the disintegration of mitochondria (assessed with the MTT reduction assay) and next of lysosomes (assessed with the NRU assay) follows mitochondria damage.