Immunofluorescence and biochemical investigation of the protective effects of naringin and diosmin on the freezability of merino ram semen.

BACKGROUND: Various antioxidant substances are added to sperm extenders to protect spermatozoa against oxidative stress and cryodamage. OBJECTIVE: To investigate the effects of the flavonoid diosmin (DIO) and a flavanone glycoside naringin (NAR) on the freezability of ram semen. MATERIALS AND METHODS: In this study, six Merino rams were used during the breeding season. The ejaculates were pooled after collection from the rams. Pooled ejaculates were divided into six groups: control, NAR 1 mM, NAR 2 mM, NAR 4 mM, DIO 2 mM, and DIO 4 mM, and then diluted with a TRIS-based diluent. The pooled semen was equilibrated, placed in 0.25 mL pipettes with 10 × 10 7 sperm cells in each pipette, and frozen in liquid nitrogen vapor. After 24 h, the pipettes were thawed at 37 degree C for 25 s and analyzed in terms of spermatological parameters. RESULTS: The highest plasma membrane integrity ratio was found in the DIO 4 mM group, whereas a statistically significant difference was found between the NAR 1 mM and NAR 2 mM groups (p < 0.05). While the DIO 4 mM group had the highest acrosome integrity rate, a statistically significant difference was found between the other groups (p < 0.05). Mitochondrial activity was the highest in the NAR 4 mM, DIO 4 mM and DIO 2 mM groups (p < 0.05). In the analysis of the sperm membrane lipid profile, it was observed that the DIO group had the highest lipid-phospholipid ratio. In sperm membrane protein profile analysis, it was found that both additives exerted protective effects at different levels. The highest total protein content was seen in the DIO 4 mM and NAR 4 mM groups. 8-hydroxydeoxyguanosine (8-OhDG) positivity was more common in the control group than in the DIO and NAR groups. Cu-Zn superoxide dismutase (SOD) expression was lower in the control group and more intense in all other groups. Positive results were especially observed in the acrosome of the sperm cells. CONCLUSION: The addition of NAR and DIO to the ram semen extender increased the quality of sperm parameters after the freeze-thaw process. Doi.org/10.54680/fr24510110412.

Rutin ameliorates cisplatin-induced reproductive damage via suppression of oxidative stress and apoptosis in adult male rats.

Cisplatin (CP) treatment causes damage in the male reproductive system. Rutin (RUT) is a naturally occurring flavonoid glycoside that has antioxidant and anti-inflammatory properties. This study aimed to investigate effects of RUT against cisplatin-induced reproductive toxicity in male rats. Twenty-one adult male Sprague Dawley rats were used. The control group received physiological saline with oral gavage during 14 days, and physiological saline was injected intraperitoneally (IP) in 10th days of study. CP Group received physiological saline during 14 days, and 10 mg kg-1 CP was injected IP in 10th day. RUT + CP group received RUT (150 mg kg-1 ) during 14 days, and 10 mg kg-1 CP was injected IP in 10th day. Spermatological parameters (including motility, cauda epididymal sperm density, dead sperm percentage and morphological sperm abnormalities), biochemical (MDA, GSH, GSH-px, SOD and CAT), histological (H&E dye) and immunochemistry evaluations of testicles were evaluated. CP treatment caused damage on some spermatological parameters, increased the oxidative stress and induced testicular degeneration and apoptosis when compared to the control group. However, RUT treatment mitigates these side effects when compared to the CP alone group. IT is concluded that RUT treatment may reduce CP-induced reproductive toxicity as a potential antioxidant compound.

Mitigation of paracetamol-induced reproductive damage by chrysin in male rats via reducing oxidative stress.

Paracetamol (PRC) is a nonsteroidal anti-inflammatory drug used widely as a painkiller for various diseases and as the symptomatic flu cure in several countries worldwide. PRC toxicity may occur under conditions of the overdose usage. Chrysin (CR) is a flavonoid that is naturally present in several plants, honey and propolis. The aim of this study was to investigate the effects of CR (at the doses of 25 mg kg-1 and 50 mg kg-1 ) pre-treatment over seven consecutive days against PRC-induced reproductive toxicity in male rats. Our results showed that PRC toxicity decreased the sperm motility, and increased dead sperm rate, abnormal sperm cell rate, apoptosis and MDA levels in testicular tissues. Pre-treatment with CR at the dose of 25 and 50 mg kg-1 for 7 days mitigated side effects of acute PRC toxicity in male reproductive system proportionally in a dose-dependent manner. This possible protection mechanism might be dependent on the antioxidant activity of CR. In conclusion, pre-treatment with CR at the dose of 25 and 50 mg kg-1 for 7 days can be the beneficial against PRC-induced reproductive toxicity proportionally in a dose-dependent manner.

3,3 diindolylmethane leads to apoptosis, decreases sperm quality, affects blood estradiol 17 ß and testosterone, oestrogen (α and ß) and androgen receptor levels in the reproductive system in male rats.

3,3 Diindolylmethane (DIM) is a major digestive product of indole-3 carbinol, obtained from Brassica family vegetables such as broccoli, cabbage and Brussels sprouts. This study aimed to investigate the effects of DIM on sperm parameters, histological structures of testicular tissues, blood testosterone (T) and estradiol 17-ß (E2) in male rats. Thirty-eight male Sprague Dawley rats were used. Rats were divided into four groups: Group I: referred as Control group, received corn oil only; Group II: as DIM-10, rats received 10 mg kg-1 DIM; Group III: as DIM-50, rats received 50 mg kg-1 DIM; Group IV: as DIM-100, received 100 mg kg-1 DIM during 53 days. Spermatological parameters, malondialdehyde (MDA) levels of testes and serum T and E2 levels were assayed. Histopathological examinations of tests were done. DIM caused an increase in MDA levels. It decreased motility and live sperm rates and increased degeneration of testicular tissues. While DIM-10 did not affect abnormal sperm rate, higher concentrations increased the abnormalities. Sperm density was higher in DIM-10 groups when compared to both other groups. Only DIM-50 had an anti-androgenic effect among all groups. Only, DIM-10 showed anti-estrogenic activity as compared to higher DIM groups. In conclusion, DIM (i) had side effect on some sperm characteristics, (ii) increased the MDA levels and (iii) led to histological degeneration of testicular tissues and apoptosis in a dose-dependent manner.

Evaluation of the effects of photostimulation on freeze-thawed bull sperm cells in terms of reproductive potential.

The aim of this study was to determine the time-dependent effectiveness of photo-stimulation against bovine sperm cells using a multi-wavelength LED (Light Emitting Diode). Spermatological parameters were evaluated for viability, acrosome structure and motility. In addition, the effect of photo-stimulation on frozen-thawed sperm cells subjected to in vitro capacitation was evaluated in terms of changes in mitochondrial membrane potential. The study consisted of two separate experiments and a total of 32 sperm samples obtained from separate bulls were used. All sperm samples were obtained from Holstein bulls using an artificial vagina. Semen was diluted to a final concentration of 92 x 106 spermatozoa per ml in 0.25 ml straws. The sperm cells were frozen using the conventional method. Straws were kept in a 37°C water bath for 20 seconds and diluted 1:4 in phosphate buffered saline (PBS) to eliminate the potentially deleterious effect of glycerol, the main permeable cryoprotectant in the freezing medium for bull sperm. This dilution also helped in the evaluation of sperm quality parameters. In the first experiment, whereas the 15-10-15 showed no differences with the control, other treatments such as 10-10-10, 5-5-5, and 3-1-3 exhibited significantly higher percentages of viable spermatozoa at 24h. The results obtained for acrosome integrity were pretty much similar to those observed in the sperm viability assessment. In effect, while the treatment consisting of 15-10-15 had no positive effects, shorter treatments exerted a much more positive effect. The percentages of acrosome-intact spermatozoa in 2-1-2 and 1-1-1 were significantly higher than those obtained in the control. The significant differences in mitochondrial membrane potential were observed at 0, 2, 4 and 24h post-photo-stimulation in all treatments, except 15-10-15. The highest increase in the percentage of spermatozoa exhibiting high mitochondrial membrane potential was found in 10-10-10, 5-5-5 and 3-1-3 treatments. With regard to total and progressive motility, whereas 10-10-10 was the best regime, 5-5-5 and 3-1-3 treatments also had a positive effect. However, 15-10-15 appeared to have a stimulating effect upon progressive motility at 2h and 4h but later declined and showed no significant differences with regard to the control at 24h. In the second experiment, not immediately after thawing but after having been kept at room temperature for up to 24h, it was observed that there was no statistical difference in terms of viability, acrosome integrity and total/progressive motility between photostimulation and the control group. This indicates that photo- stimulation is less able to exert a beneficial effect when post-thawed sperm are not immediately stimulated. As a result it was determined that photo-stimulation at a pattern of 10-10-10, 5-5-5, 3-1-3 and, to a lesser extent 2-1-2, increases the resilience of frozen-thawed bull sperm when applied upon thawing.

Activity of resveratrol on the influence of aflatoxin B1 on the testes of Sprague dawley rats.

Twenty eight male Sprague Dawley rats (aged 3 months) were used in the study. The animals were given feed and water as ad libitum. Sprague dawley rats were randomly divided into 4 groups as 7 rats in each group. Except for the control one, aflatoxin B1 (7.5 µg / 200 g), resveratrol (60 mg / kg) was administered to rats of 3 other groups. At the end of the 16th day, blood, semen and tissue specimens were taken by decapitation under ether anesthesia. When we evaluate the spermatological parameters, it is understood that resveratrol has a statistically significant difference in terms of sperm motility and viability (membrane integrity) compared to the control group and aflatoxin B1 administration groups, indicating a protective effect on spermatological parameters. In terms of pathological parameters - histopathological examination - in the control and resveratrol groups, seminiferous tubules were observed to be in normal structure. In the group treated with aflatoxin, the regular structure of the spermatogenic cells deteriorated and the seminiferous tubules became necrotic and degenerative. In the group treated with Afb1 + res, the decreasing of necrotic and degenerative changes were determined compared with in the group treated with aflatoxin. As immunohistochemical examination, cleaved caspase 3 expression was found to be very low in the control and resveratrol groups. Cleaved caspase 3 expression was severely exacerbated in seminiferous tubules in aflatoxin group but cleaved caspase 3 expression level decreased in Afb1 + res. In the biochemical direction, resveratrol has been shown to inhibit the adverse effects of aflatoxin on antioxidant levels and to show a protective effect. For this purpose, the use of resveratrol with antioxidant activity was investigated in preventing or ameliorating damage to aflatoxin B1. It has been concluded that resveratrol effectively prevent the aflatoxin-induced testicular damage and lipid peroxidation. It has also been shown that resveratrol has protective effects on sperm motility and viability.