Cryopreservation of ram semen: baicalein efficiency on oxidative stress, chromatin integrity, viability and motility post thaw.

Baicalein (B) has potential antioxidant properties, but it has not been tested as a ram semen extender. This study aimed to assess the impact of B on various sperm parameters and determine its potential influence on semen quality after the freeze-thawing process. During the breeding season, ejaculates were obtained from four rams with the aid of an artificial vagina. The collected mixed semen samples were divided into four groups: control (C; 0), B0.5 (0.5 mM), B1 (1 mM), and B2 (2 mM). After semen extension, the samples were loaded into 0.25 mL straws and stored for 2 h at 4°C prior to freezing in liquid nitrogen vapor and thawed in a water bath at 37°C. Among the groups, B0.5 demonstrated the highest progressive motility results, while B1 and B2 exhibited reduced motility (p < 0.05). In terms of high mitochondrial membrane potential, plasma membrane and acrosome integrity, and viability, B0.5 showed significantly superior outcomes to the other B groups (p < 0.05), although it was not significantly better than C. B1 displayed the highest plasma membrane integrity levels (p < 0.05). Notably, B2 displayed the lowest total antioxidant status levels among the groups (p < 0.05). The findings of this study suggested that the in vitro spermatological characteristics of ram spermatozoa such as progressive motility and chromatin integrity can be protected from the freeze-thawing process by using the 0.5 mM dose of baicalein as a semen extender. The treatment of sperm freezing might benefit from further in-depth research on the role of B in the improvement of cryoinjury and its underlying processes.

Evaluation of Cryopreserved Ram Sperm with Nano-Ozone Solution and Post-Thaw Life Span by Flow Cytometric Analysis.

Ozone has been used as a therapy tool in medical science for conditions such as ulcers, peritonitis, wounds, and mostly joint problems. Ozone therapy strengthens the resistance to infections by kick-starting antioxidant, anti-inflammatory, and immune modulation systems. Ozone creates a defensive response against oxidative stress in membranes and protects metabolism against reactive oxygen species (ROS). Sperm membranes are one of ROS's main targets; therefore, the cells' cryopreservation process requires more defensive elements for better results. This study aimed to investigate the protective effect of nano-ozone solution (NOS) on ram sperm cryopreservation and the influence of the process on various sperm parameters for post-thaw (0 hour) and postincubation (6 hours) time points. Samples were collected from six Merino rams in the breeding season by electroejaculation five times at 3-day intervals. The study was conducted by cryopreservation of the samples using a tris citric acid-egg yolk-based extender. The samples were subjected to freezing in control and NOS (0.5, 1, and 2 µg/mL nano-ozone supplemented). Post-thaw motility, hypo-osmotic swelling test, acrosome (fluorescein isothiocyanate-conjugated Pisum sativum agglutinin [PSA-FITC]), and DNA integrities (terminal deoxynucleotidyl transferase dUTP nick end labeling [TUNEL]) were evaluated with a phase-contrast microscope. Mitochondrial membrane potential (MMP) assessments were conducted by JC1-PI dual staining with a flow cytometer. Malondialdehyde and glutathione (GSH) levels were measured by a spectrophotometer. Sperm kinematics were investigated by a computer-assisted sperm analyzer (CASA) at the post-thaw time point. Compared with the control, relatively low doses of NOS (0.5 and 1 µg/mL) yielded better results in many parameters (motility, membrane and acrosomal integrities, MMP, various sperm kinematics, and GSH levels) (p < 0.05). The addition of low ozone doses to cryopreservation extenders improved the results compared with the control group at post-thaw and postincubation time points. Despite the valuable potential of nano-ozone supplementation in ram sperm cryopreservation, this subject requires further investigations with fertility trials soon.

Investigation of the effects of syringic acid supplemented to Tris semen diluent on ram semen freezability.

To the best of our knowledge, no research has been conducted to test the effects of syringic acid (SA) on ram semen freezing within the scope of natural antioxidants added to semen extenders. Therefore, this study had two main objectives. First, to test whether adding SA to ram semen freezing extender has a protective effect and contributes positively to sperm kinetic, plasma and acrosome integrity, mitochondrial membrane potential, lipid peroxidation, oxidant and antioxidant and DNA damage parameters after thawing. Second, it was to determine at what concentration the SA supplemented to the extender could be applied by in vitro studies by preserving the fertilization ability of frozen semen at the highest level. In the study, six individuals of Sönmez rams were used. The semen was collected from the rams using an artificial vagina and pooled. The pooled semen was divided into five different groups and extended with 0, 0.5, 1, 2 and 4 mM SA (control C, SA0.5, SA1, SA2 and SA4, respectively). After dilution, the semen samples were kept at 4°C for 3 h, then loaded into 0.25 mL straws and frozen in liquid nitrogen vapour. The SA1 and SA2 groups were higher plasma membrane and acrosome integrity (PMAI), high mitochondrial membrane potential (HMMP), plasma membrane integrity and motility compared to other groups (p < .05). It was observed that SA supplemented to the Tris extender significantly reduced DNA damage, and the lowest values were obtained especially in the SA1 and SA2 treatments (p < .05). Also, lowest MDA level was determined at the SA1 and this was statistically significant compared to SA4 and C (p < .05). In conclusion, it was revealed that SA added to Tris semen extender at 1 and 2 mM treatment doses increased progressive and total motility and preserved PMAI, plasma membrane integrity, HMMP and DNA integrity.

Investigation of Changes in Spermatozoon Characteristics, Chromatin Structure, and Antioxidant/Oxidant Parameters after Freeze-Thawing of Hesperidin (Vitamin P) Doses Added to Ram Semen.

We conducted this study to determine the potential cryopreservative effects of different hesperidin (vitamin P; H) doses on ram semen after freeze-thawing. Semen samples were obtained from Sönmez rams using an artificial vagina. The samples were divided into six groups: control, 10, 50, 100, 250, and 500 µg/mL H (C, H10, H50, H100, H250, and H500, respectively). At the end of the study, sperm motility and kinetic parameters, acrosome integrity (AI), mitochondrial membrane potential (MMP), viability, lipid peroxidation levels (LPL), chromatin damage, oxidant parameters, and antioxidant parameters were assayed. None of the doses of H added to the semen extender showed any enhancing effects on progressive motility compared to C (p > 0.05). In fact, H500 had negative effects (p < 0.05). Moreover, AI was the highest at the H10 dose, while LPL values were the lowest at the same dose (p < 0.05). The doses of H10 and H50 added to the Tris extender medium showed positive effects on sperm cell chromatin damage. Consequently, we can say that H doses used in this study are not effective on semen progressive motility, but the H10 dose is effective on AI and chromatin damage by reducing LPL.

Thymoquinone improves motility, plasma membrane integrity and DNA integrity of frozen-thawed ram semen.

The aim of this study was to determine the effects of thymoquinone (TQ), which is the most essential active compound of Nigella sativa, on the spermatological parameters of ram semen during cryopreservation. Ejaculates were collected from five Sonmez rams using an artificial vagina and extended with Tris-based extender not containing TQ (control, 0 µg/ml TQ) and containing 10, 25, 50 and 100 µg/ml TQ. The extended semen samples were equilibrated in a + 4°C cold cabinet for 2 h. After 2 h, the samples were loaded into 0.25 ml French straws. The straws were frozen by liquid nitrogen vapour and stored in a liquid nitrogen container (-196°C). The frozen straws were thawed in a water bath (37°C for 30 s) and evaluated in terms of motility characteristics, plasma membrane and acrosome integrity, mitochondrial reactive oxygen species levels, lipid peroxidation levels, DNA damage and biochemical alterations (oxidative stress index, malondialdehyde and glutathione). TQ100 had higher total motility (53.59 ± 3.01) and progressive motility (19.84 ± 1.44; not significantly different from TQ25 and TQ50) compared to the control and TQ10 (p Ë‚ 0.05). According to the results of the analyses on motility characteristics, there were significant differences between the groups in terms of curvilinear velocity (VCL), amplitude of lateral head displacement (ALH) and linearity (LIN; p Ë‚ 0.05). The highest DNA damage was detected in the control group (p Ë‚ 0.05). TQ50 had higher plasma membrane and acrosome integrity (59.56 ± 5.92) compared to the control and TQ25 (p < 0.05) but not significantly different from TQ10 and TQ100. The lowest mitochondrial reactive oxygen species levels were detected in TQ50 and TQ100 (p Ë‚ 0.05). There were no significant differences among the groups in terms of their oxidative stress index, lipid peroxidation, malondialdehyde and glutathione levels (p > 0.05). According to the results, it could be concluded that supplementing 50 or 100 µg/ml TQ to Tris extenders that were used for ram semen cryopreservation showed a positive effect on motility, plasma membrane integrity and acrosome integrity, and it reduced DNA damage and mitochondrial reactive oxygen species levels.

Protective effects of dantrolene and methylprednisolone against spinal cord injury-induced early oxidative damage in rabbit bladder: A comparative experimental study.

BACKGROUND: Spinal cord injury (SCI) may cause dysfunction in the bladder and many distal organs due to systemic inflammatory response and oxidative stress-related injury. OBJECTIVES: We investigated the preventive effects of dantrolene (DNT) and methylprednisolone (MP) on stress-induced tissue damage in rabbit bladder with SCI. MATERIAL AND METHODS: A total of 35 rabbits were included in this study and they were divided into 5 groups: group 1 - control, group 2 - SCI only, group 3 - SCI and DNT, group 4 - SCI and MP, and group 5 - SCI and DNT+MP. Twenty-four hours after SCI, the bladders of these rabbits were removed and the histopathologic changes in the bladder were examined under a light microscope. Additionally, malondialdehyde (MDA), glutathione (GSH), and nitric oxide (NO) levels were evaluated as antioxidant agents both in bladder tissue and in blood. RESULTS: Compared to the control group, there was an increase in edema and congestion in all groups. The least amount of edema was observed in the group receiving DNT and the least amount of congestion was observed in the group receiving combined treatment (group 5). No superiority was found between the drug-receiving groups in terms of reducing MDA level in blood and tissue after SCI. The most successful group was the group receiving combined drug therapy in terms of increasing the blood GSH level, which was significantly decreased after SCI. After SCI, blood NO level increased significantly in all groups. Nitric oxide levels in the bladder tissue significantly decreased in the groups receiving DNT and combination therapy and fell in the control group. CONCLUSIONS: Dantrolene and MP may have potential benefits against oxidative damage in the bladder after SCIs because of their anti-inflammatory and antioxidant effects. In particular, the combined use of DNT and MP at different doses can be considered a treatment strategy.

Protective effect of polydatin, a natural precursor of resveratrol, against cisplatin-induced toxicity in rats.

The aim of the present study was to evaluate the possible protective effect of polydatin (PD) on cisplatin (Cis) induced oxidative stress in rats. Totally, thirty male Wistar albino rats were fed standard rodent diet and divided into 5 equal groups: the control group (vehicle treated) was treated with physiological saline for ten days both orally and intraperitoneally (i.p.), the second group was orally treated with physiological saline and 7 mg/kg single i.p. injection of Cis on the seventh day, and third, fourth, and fifth groups were treated orally PD at 25, 50, and 100 mg/kg/day, respectively for 10 days starting seven days before Cis injection and 7 mg/kg single i.p. Cis was injected on the seventh day. Cis resulted in significant increase malondialdehyde levels and decreased glutathione levels. In addition, Cis treatment decreased superoxide dismutase and catalase activities in erythrocyte and tissues. Also, Cis treatment caused to increase DNA damage and affected serum biochemical parameters whereas slightly decreased AchE activity. However, treatment of PD resulted in reversal of Cis-induced oxidative stress, lipid peroxidation, and activities of antioxidant enzymes. In conclusion, PD has protective effect in rats against Cis-induced oxidative stress, enhances antioxidant defence mechanism, and regenerates their tissues.