Veterinary Research Forum

Abstract Testicular ischemia-reperfusion (I/R) injury during testicular torsion is strongly influenced by oxidative stress caused by excessive accumulation of uncaptured reactive oxygen species (ROS). Kisspeptin-10, a biologically active fragment of the kisspeptin peptide family, has demonstrated significant antioxidant and anti-apoptotic properties. Recent studies indicate that kisspeptin-10 can mitigate oxidative stress by reducing reactive oxygen species levels and enhancing the activity of endogenous antioxidant enzymes such as superoxide dismutase (SOD) and catalase. This study examined the effects of kisspeptin-10 on I/R injury in testicular torsion/detorsion (T/D) of male rats. Twenty male rats were divided into four groups: the control group and three treatment groups (720° T/D, 720° T/D+ 0.50 µg kg-1 kisspeptin-10, 720° T/D+ 1.00 µg kg-1 kisspeptin-10). After inducing 720° clockwise testicular torsion for 2 hr, various factors such as sperm parameters, number, total motility, viability, DNA damage and hypoosmotic test were evaluated. The results showed that 720° T/D can increase sperm DNA damage. In addition, it also had negative effects on overall motility and other properties such as viability and plasma membrane functionality. The results also showed that administration of kisspeptin-10 to T/D rats can reduce DNA damage. These effects could also increase overall motility, viability and plasma membrane functionality compared to the T/D group. Based on our results, kisspeptin-10 provides significant protection against acute T/D injury to the testis when administered after spermatic cord torsion in rat.

Abstract Sterilization in animals serves multiple purposes, such as behavior control, performance improvement, and population management. Chemical sterilization has emerged as a promising non-surgical alternative to traditional methods. This study aimed to investigate the effects of intra-testicular injection of zinc-doped carbon dots (Zn-CDs) nanoparticles as a chemical sterilant in mature rats. Twenty-five rats were randomly divided into five groups, including a control group without injection, a sham group receiving 0.50 mL distilled water, and three treatment groups administered respectively 0.50, 2.00, and 8.00 mg kg^-1 of Zn-CDs synthesized through a hydrothermal process. Following anesthesia with ketamine and xylazine, and aseptic preparation, intra-testicular injections were administered bilaterally. At 60th day post-injections, blood samples were collected to measure serum testosterone levels using chemiluminescence immunoassay. The rats were then surgically castrated to assess sperm parameters and testicular histopathology. Testicular oxidant/anti-oxidant status was also evaluated. The results revealed a dose-dependent reduction in sperm viability, membrane integrity, and motility, accompanied by increased sperm DNA damage. The highest Zn-CDs dose caused the most significant decrease in sperm concentration, as well as severe testicular tissue damage. In addition, anti-oxidant capacity, seminiferous tubules maturation, testosterone production, and spermatogenesis declined with increasing Zn-CDs concentrations in a dose-dependent manner. These findings indicate that intra-testicular injection of Zn-CDs effectively induces infertility in mature rats and holds potential as a chemical sterilization method. With further studies to evaluate safety and efficacy, this approach could be developed as a practical solution for large-scale in situ castration, offering a non-surgical alternative for over-population control programs.

Abstract Buildup of reactive oxygen species during testicular torsion causes oxidative stress and ischemia-reperfusion (I/R) injury in testis. The purpose of this study was to investigate influence of β-cryptoxanthin (BCX) on I/R injury in testicular torsion/detorsion in mature rats. Thirty mature male Wistar rats were divided into five groups of six animals each, including sham group: In this group, midline incision of the scrotum was performed and the testicles were taken out for 2 hr with a 720-degree rotation, I/R group: In this group, midline incision of the scrotum was performed and the testicles were taken out and undergone ischemia for 2 hr with a 720-degree rotation, I/R/Oil group: In this group, a midline scrotum cut was performed, the testicles were taken out, ischemia was created for 2 hr with a 720-degree rotation, and at the end of ischemia 100 µL of corn oil (BCX solvent) was injected intraperitoneally, I/R/BCX10 group: The same as I/R/Oil group, as well as intraperitoneal administration of 100 µL of BCX (10.00 µg kg^-1) at the end of ischemia, and I/R/BCX40: The same as I/R/Oil group, as well as intraperitoneal administration of 100 µL of BCX (40.00 µg kg^-1) at the end of ischemia. Evaluations were based on histopathological and spermatological parameters and oxidative stress assessments. Histopathological spermatological and oxidative stress parameters values obtained from I/R/BCX40 were significantly different from those of other groups (p < 0.05). It could be concluded that BCX could ameliorate testicular injuries in acute testicular torsion/detorsion in mature rats.

Abstract Testicular ischemia-reperfusion (I/R) injury during testicular torsion is strongly influenced by oxidative stress caused by excessive accumulation of unscavenged reactive oxygen species. This study aimed to investigate the effects of intra-peritoneal administration of Mito-TEMPO (MT) on I/R injury in testicular torsion/detorsion (T/D) in mice. Forty-two male mice were divided into seven groups including 1 control and 6 treatment groups (360° T/D, 720° T/D, 360° T/D + 0.70 mg kg^-1 MT, 360° T/D + 1.00 mg kg^-1 MT, 720° T/D + 0.70 mg kg^-1 MT, and 720° T/D + 1.00 mg kg^-1 MT). After inducing 360° and 720° clockwise testicular torsions for 2 hr, sperm parameters, apoptosis-related genes expression, and in vivo fertility index were evaluated. The results showed that 720° T/D can lead to increased abnormal sperm morphology, sperm DNA damage, and Bax expression, while the Bcl-2 expression was reduced compared to the other groups. In addition, it also had negative effects on sperm total and progressive motilities as well as viability and plasma membrane functionality (PMF). The results also showed that administration of MT to T/D mice can result in a reduction in abnormal sperm morphology, DNA damage, and Bax expression. It could also increase sperm total and progressive motilities, viability and PMF, Bcl-2 expression, and in vivo fertility index. Based on our results, it is concluded that MT, when administered after spermatic cord torsion in mice, provides significant protection against acute testicular T/D injury.

Abstract The cooling procedure markedly diminishes the quality of guinea pig (Cavia porcellus) sperms, primarily because their membranes are highly susceptible to this process. This susceptibility triggers the generation of reactive oxygen species and free radicals, ultimately leading to lipid peroxidation in the sperm membrane. Surprisingly, there has been a lack of research on the use of Tris-based extenders to safeguard guinea pig sperm under refrigeration conditions. This study aimed to assess the viability of guinea pig spermatozoa diluted in Tris buffer-based extenders during storage at 5.00 ˚C. Sperm collection was carried out through castration of the animals. For this study, 10 adult male guinea pigs were utilized, being divided into four groups including phosphate-buffered saline (PBS), human tubal fluid (HTF), Tris-citric-glucose (TCG), and Tris-fructose-yolk (TFY) cultures. Evaluations including sperm motility, morphology, plasma membrane integrity, viability, and total count were conducted at 0, 24, and 48 hr after sampling. The results obtained indicated that at the 24-hr and 48-hr marks of the experiment, both overall and progressive motility percentages, viability, plasma membrane integrity, and morphology of sperms in the PBS and HTF cultures exhibited a significant increase in comparison with the TCG and TFY cultures. Consequently, it can be inferred that PBS and HTF cultures are highly effective in preserving the quality of guinea pig spermatozoa.

Abstract Busulfan is known to cause several adverse effects including reproductive toxicity in humans. Garlic (Allium sativum), a widely distributed medicinal plant, is highly regarded for its medicinal activities including antioxidant property.This study was conducted to assess whether garlic extract could serve as protective agents against testicular toxicity during busulfan treatment in a mice model.Seventy-two adult male mice were randomly divided into nine groups. In groups 1,2 and 3, distilled water, busulfan, and dimethyl sulfoxide and in the treatment groups hydro-alcoholic extract of garlic was administered orally at different doses per day (groups 4, 5 and 6; 200, 400, 800 mg kg^-1 respectively). Groups 7, 8 and 9 were treated with the extract (200, 400 and 800 mg kg^-1, respectively) plus busulfan. Following euthanasia, blood samples and epididymal sperm were collected.The busulfan-treated group showed significant decreases in sperm qualityparameters, and serum levels of testosterone, LH and FSH was observed in the busulfan-treated mice. In addition, the TAC levels and antioxidant enzymes activities were reduced and malondialdehyde (MDA) levels were increased in the busulfan-treated mice. Notably, garlic extract co-administration caused a considerable recovery in sperm qualityparameters, TAC levels, antioxidant enzymes activities, hormonal changes and MDA level. Based on our results, garlic has antioxidant effects against busulfan-induced testicular damages in mice.

Abstract Bisphenol-S (BPS) is a new bisphenol-A substitute widely used in many plastic products. Bisphenol-A as a main member of bisphenol family has been known as an endocrine system disrupter chemical compound. Like other members of bisphenol family, there is public health concern about the toxic effects of BPS on reproductive system, thus, we examined BPS effects on in vitro fertilization (IVF) potential and oxidative stress status in a murine model. Adult female mice (n = 70) were randomly divided into control and BPS-treated groups. Bisphenol-S was administered at doses of 0, 1, 5, 10, 50 and 100 µg kg^-1 body weight per day intraperitoneally for 21 consecutive days. Twenty-Four hr after the last treatment, five mice in each group were super-ovulated and the oocytes were harvested for IVF. All ovaries were collected and used for biochemical factors analyses. Bisphenol-S exposure at doses more than 10 µg kg^-1 induced developmental arrest of pre-implantation embryos. Further, lipid peroxidation measurement in ovaries indicated that all doses of BPS cause oxidative stress in female mice. In conclusion, BPS administration even in low doses can result in female reproductive toxicities and oxidative stress in mice.