Protective effects of nobiletin on cisplatin induced neurotoxicity in rats.

OBJECTIVES: This study was designed to investigate the possible antioxidant, antiapoptotic and neuroprotective effects of nobiletin on cisplatin-induced neurotoxicity rat model by evaluating neurotrophins, antioxidants and histopathology. METHODS: Forty male Wistar Albino rats were divided into four groups: control, cisplatin (CIS), cisplatin + nobiletin (CIS + NOB) and nobiletin + cisplatin (NOB + CIS). CIS + NOB was applied nobiletin (10 mg/kg, i.p.) during the last four days whereas NOB + CIS was applied nobiletin during the first four days of the study. Cisplatin (4 mg/kg, i.p. twice a day) was administered to the experimental groups on the 5th day of the study. All rats were sacrificed on the 10th day of the study. BDNF, NGF, G6PD, GPx, tGSH and MDA levels were determined in brain. In addition, routin histolopathological analysis and caspase-3 immunoreactivity assay were conducted. RESULTS: BDNF concentrations increased in nobiletin-administered groups, compared to Control and CIS and that the increase was statistically significant in NOB + CIS (p < 0.05). It was also found that G6PD activity increased (p < 0.05) in the nobiletin-administered groups, compared to control and CIS. Histopathologically, neuronal degeneration, oedema and gliosis increased in CIS compared to Control, and nobiletin administration decreased neuronal degeneration and oedema compared to CIS (p < 0.05). Cisplatin increased (p < 0.05) caspase-3 immunoreactivity in cerebrovascular endothelium and neurons compared to Control, while nobiletin administration decreased caspase-3 immunoreactivity in cerebrovascular endothelium. Caspase-3 immunoreactivity in neurons decreased only in NOB + CIS (p < 0.05). CONCLUSION: Nobiletin increased BDNF concentration and G6PD activity in brain and when evaluated together with histopathological and immunohistochemical findings, it may have antioxidant, antiapoptotic and neuroprotective effects against cisplatin.

Combination of fetuin and trehalose in presence of low glycerol has beneficial effects on freeze-thawed ram spermatozoa.

BACKGROUND: Freeze-thawing process negatively affects ram spermatozoa in terms of sperm quality, DNA integrity and antioxidant defence system. Thus, antioxidant supplementation of spermatozoa during freeze-thawing is suggested to improve sperm parameters. OBJECTIVES: The aim of this study was to determine the effects of fetuin and trehalose added into ram semen extender on sperm parameters, antioxidant parameters, antioxidant-related gene expressions and DNA integrity during the freeze-thawing process, in low glycerol concentration. METHODS: Semen samples collected from six mature rams were pooled and splitted into equal aliquots and diluted with a tris-based extender containing different concentrations of glycerol (G5; %5 and G3; %3), fetuin (F; 2.5, 5 and 15 mg/mL) and trehalose (60 mm) as eight groups (G5F0, G5F2.5, G5F5, G5F15, G3F0, G3F2.5, G3F5 and G3F15). RESULTS: G3F5 group resulted in the highest motility, mitochondrial activity and viability and the lowest DNA fragmentation and DNA damage (p < 0.05). Also, G3F0 displayed considerably more cryoprotective effect compared with G5F0 group (p < 0.05) in terms of motility, mitochondrial activity and viability rates. Lipid peroxidation levels decreased in G5F5 group compared with G5F0 group (p < 0.05). The levels of total glutathione increased in G3F2.5 group (p < 0.05) in comparison with the G5F0 group. NQO1 gene levels were upregulated approximately twofold in G5F5, G5F15, G3F2.5, G3F5 and G3F15 groups compared with G5F0 group (p < 0.05). The levels of GCLC gene were approximately twofold higher in G3F0, G3F2.5, G3F5 and G3F15 groups compared with G5F0 group (p < 0.05). GSTP1 gene levels were significantly higher with different levels in all treatment groups except for G5F2.5 and G3F0 groups in comparison with G5F0 group (p < 0.05). CONCLUSIONS: Co-supplementation of tris-based extender having low glycerol (3%) with trehalose and fetuin to enhance the quality of ram spermatozoa after freeze-thawing process is recommended.

Decreasing glycerol content by co-supplementation of trehalose and taxifolin hydrate in ram semen extender: Microscopic, oxidative stress, and gene expression analyses.

This study aimed to evaluate the comparative effects of taxifolin hydrate and trehalose on the quality of frozen-thawed ram spermatozoa for the first time. Ejaculates collected from six mature rams were pooled, and divided to eight equal aliquots to extend them with different concentrations of glycerol (%5 and %3), taxifolin hydrate (10, 100, and 500 µM), and trehalose (60 mM) as eight groups (G5T0, G5T10, G5T100, G5T500, G3T0, G3T10, G3T100, and G3T500). After freeze-thawing process of cryopreservation, microscopic and oxidative stress parameters, and gene expression levels were investigated for understanding of possible impacts of taxifolin hydrate and trehalose. The study showed that G3T10 resulted in the highest post-thawed viability and mitochondrial activity. Moreover, all extenders with taxifolin hydrate reduced DNA fragmentation in comparison to G5T0, but DNA damage was prevented at the highest rate in presence of G5T10. The level of LPO significantly decreased in the groups G5T500 and G3T100, and the expression levels of NQO1, GCLC, and GSTP1 genes significantly increased in the groups G5T100, G5T500, G3T10, and G3T100 compared to the group G5T0. Finally, co-supplementation of tris-based extender having 3% glycerol with 60 mM trehalose and 10 µM taxifolin hydrate in cryopreservation extender may be recommended to improve the quality of post-thawed ram spermatozoa. However, further in vivo and in vitro studies are suggested to evaluate fertility rates of frozen-thawed ram spermatozoa co-supplemented with trehalose and taxifolin hydrate.

Erythrocyte and spermatozoa glucose-6-phosphate dehydrogenase activity in merino rams: An experimental study.

BACKGROUND: Glucose-6-phosphate dehydrogenase (G6PD) is the first enzyme of the pentose phosphate metabolic pathway that supplies reducing agents by maintaining the level of reduced nicotinamide adenine dinucleotide phosphate. OBJECTIVE: It was aimed to determine the activity of erythrocyte and spermatozoa G6PD in the breeding and non-breeding seasons in Merino rams. And also, to find out the relation of these parameters with sperm quality parameters for better understanding the role of this enzyme in male fertility. MATERIALS AND METHODS: 1.5-2 yr-old healthy, 14 Merino rams were involved. Ejaculate samples were collected using an artificial vagina, in October (the breeding season) and April (the non-breeding season). Blood samples were collected prior to sperm collection. Sperm volume (ml), motility (%), mass activity (1-5), concentration (×106), viability (%), abnormal acrosome morphology (%) and abnormal sperm morphology (%) was evaluated. The activities of spermatozoa and erythrocyte G6PD were determined and the relation of sperm parameters with G6PD activity was evaluated. RESULTS: Erythrocyte G6PD activity was higher (p≤0.001), whereas spermatozoa G6PD activity was lower (p≤0.001) in the breeding season (1.928±0.231 U/g hemoglobin, 129.65±28.41 U/g protein, respectively) from that in the non-breeding (0.530±0.066 U/g hemoglobin, 562.36±94.92 U/g protein, respectively). There were also significant differences among sperm quality parameters within the seasons. Positive correlation was determined between spermatozoa G6PD activity (r=0.053, p=0.03 and sperm concentration in the breeding season. CONCLUSION: Higher spermatozoa G6PD activity in October, where the level of polyunsaturated fatty acids is suggested to be increased, may reflect the increased need of nicotinamide adenine dinucleotide phosphate and thus higher G6PD activity for the oxidative balance.

The effects of various levels of boron supplementation on live weight, plasma lipid peroxidation, several biochemical and tissue antioxidant parameters of male mice**: Effects of boron on performance, antioxidant and some metabolits of mice.

In this study, live weight, live weight gain, plasma GPx, GR, LDH, ALT activities, triglyceride, total protein, albumin and LPO levels, also liver and brain SOD and GPx activities were investigated after administration of boron (0.10, 0.20 and 0.30 mg/day) into male mice with drinking water for 60 days. Blood albumin and triglyceride levels were not affected with boron (p > 0.05) where triglyceride levels, with increasing amounts of boron, displayed a slight decrease within the normal ranges. From the antioxidant-oxidant balance parameters, LPO and GR levels were not affected from boron, where GPx activity was increased significantly (p < 0.001) comparing the groups of boron and control. LDH and ALT activities were affected significantly (p < 0.001) with decreased ALT and increased LDH levels with increasing amounts of boron. In regards of liver and brain GPx and SOD activities, significant increases were determined. Liver GPx and SOD activities were increased within the groups with the increasing amount of boron, where in brain, SOD (p < 0.05) was affected significantly but GPx (p > 0.05) displayed a gradual insignificant increase. As regards live weight gain, a gradual increase was determined during experimental period, but only the 45th day, the increase was statistically significant (p < 0.05). It is suggested that, new studies on the effects of different doses and compounds of boron in laboratory animals in regards of antioxidant and metabolic effects may be helpful for the understanding of the subject.

Effects of cysteine and ergothioneine on post-thawed Merino ram sperm and biochemical parameters.

The aim of the current study was to evaluate the effects of cysteine and ergothioneine on the post-thawed sperm parameters, lipid peroxidation and antioxidant activities. Semen samples from 5 mature Merino rams were used in the study. Semen samples, which were diluted with a Tris-based extender containing l-Cysteine and l-(+)-Ergothioneine and no antioxidant (control), were cooled to 5°C and frozen in 0.25 ml French straws. Frozen straws were then thawed individually at 37°C for 20s in a water bath for evaluation. Ergothioneine at doses of 2 and 4mM increased percentages of subjective motility, VSL and VCL, compared to controls following the freeze-thawing (P<0.001). Ergothioneine at three different doses led to higher rates of progressive motility and VAP, compared to control groups (P<0.001). Cysteine and ergothioneine at three doses provided the higher rates of ALH, in comparison to no antioxidant group (P<0.001). As regards CASA motility, supplementation with antioxidants did not provide any significant difference on the percentage of post-thaw sperm CASA motilities, in comparison to the control. In regards of sperm membrane integrity, only cysteine 1mM provided a greater protective effect, compared to control (P<0.001). Percentages of sperm with high mitochondrial activity were dramatically increased with cysteine at doses of 1 and 2mM, compared to control (P<0.05). No significant differences were observed in sperm acrosome integrities among groups. CAT activity was increased significantly only in cysteine1mM compared to control group (P<0.001). Cysteine at doses of 2 and 4mM showed a tendency of increased activities of CAT when compared to control. But these increases were not statistically significant. Supplementation with antioxidants did not significantly affect activities of SOD and GPx. Findings of this study showed that ergothioneine supplementation in semen extenders, was of greater benefit to motility and motion characteristics of frozen-thawed ram sperm.

Effect of glutamine and sugars after bull spermatozoa cryopreservation.

The objective of this study was to evaluate the effects of the addition of different sugars (raffinose, sucrose, and trehalose) on bull spermatozoa cryopreserved in a commercial extender (Optidyl) supplemented with glutamine on semen parameters, fertilizing ability and superoxide dismutase (SOD) activity. Nine ejaculates for each bull were used in the study. Semen was frozen in five different extenders: raffinose 25 mM plus glutamine 3 mM (RGO), sucrose 25 mM plus glutamine 3 mM (SGO), trehalose 25 mM plus glutamine 3 mM (TGO), glutamine 3 mM (GO) and control (O). Insemination doses were processed so that each 0.25 mL straw contained 15 x 10(6)sperm. Groups of GO and RGO resulted in the higher rates of subjective (54.0 ± 1.7% and 64.0 ± 1.1%; P < 0.01) and CASA motilities (53.0 ± 2.7% and 61.0 ± 4.4%; P < 0.001), respectively compared to the other groups. The supplementation of additives did not provide an effect on the level of post-thaw sperm CASA progressive motilities, the sperm motion characteristics and pregnancy rates. GO and RGO provided the better protective effect for sperm acrosome (4.0 ± 0.5% and 12.0 ± 0.6%) and total abnormalities (5.0 ± 0.3% and 13.0 ± 0.7%; P < 0.001), respectively. At the HOST values, the additives did not give to result the protective effect in comparison to Optydil extender without additives (P > 0.05). For pregnancy rates, there were no significant differences among the groups. The supplementation of additives did not provide any significant difference on the level of SOD activity (P > 0.05). It can be also thought that these sugars might have worked with glutamine in a synergy. Thereby, sugars such as raffinose and sucrose with glutamine in freezing extender may be recommended to facilitate bull semen freezability.

The effect of cysteine and glutathione on sperm and oxidative stress parameters of post-thawed bull semen.

The aim of this study was to determine the effects of antioxidants such as reduced glutathione (GSH) and cysteine in Laiciphose® extender on semen parameters, fertilizing ability, lipid peroxidation (LPO) level and glutathione peroxidise (GPx) activity of post-thawed bull semen. Totally 54 ejaculates of three bulls were used in the study. Five groups, namely; GSH (0.5 and 2 mM), cysteine (5 and 10 mM) and control group, were conducted to test the antioxidants in Laiciphose®. Insemination doses were processed that each 0.25-mL straw contained 15 x 106 sperm. The addition of antioxidants did not present any significant effect on the percentages of post-thaw sperm morphology (acrosome and total abnormalities), subjective, CASA and progressive motilities, as well as sperm motility characteristics (VAP, VSL, VCL, LIN and ALH), compared to the control groups (P > 0.05). GSH 0.5mM (55.5±7.38%) and cysteine 10 mM (48±5.65%) led to lower rates of DNA damage, compared to control (P < 0.05). As regards to MDA level, cysteine at 10 mM dose gave the highest level (4.99±0.44 nmol/L) (P < 0.001). GPx activity was demonstrated to be higher level upon the addition of 5 mM cysteine when compared to the other groups (P < 0.05). With respect to fertility results based on 60-day non-returns, the supplementation of antioxidants did not present significant differences (P > 0.05). The results of this study may provide an useful information for the future studies in this area. So, further studies could be suggested to achieve better information in terms of the DNA damage and fertilizing capacity of bull sperm frozen with effective antioxidants.

Effects of antioxidants on post-thawed bovine sperm and oxidative stress parameters: antioxidants protect DNA integrity against cryodamage.

This study was conducted to determine the effects of methionine, inositol and carnitine on sperm (motility, abnormality, DNA integrity and in vivo fertility) and oxidative stress parameters (lipid peroxidation, total glutathione and antioxidant potential levels) of bovine semen after the freeze-thawing process. Nine ejaculates, collected with the aid of an artificial vagina twice a week from each Simmental bovine, were included in the study. Each ejaculate, splitted into seven equal groups and diluted in Tris-based extender containing methionine (2.5 and 7.5 mM), carnitine (2.5 and 7.5 mM), inositol (2.5 and 7.5 mM) and no additive (control), was cooled to 5 °C and then frozen in 0.25 ml straws. Frozen straws were then thawed individually at 37 °C for 20s in a water bath for the evaluation. The extender supplemented with 7.5 mM doses of carnitine and inositol led to higher subjective motility percentages (61.9±1.3% and 51.3±1.6%) compared to the other groups. The addition of methionine and carnitine at doses of 2.5 and 7.5 mM and inositol at doses of 7.5mM provided a greater protective effect in the percentages of total abnormality in comparison to the control and inositol 2.5 mM (P < 0.001). As regards CASA motility, 7.5 mM carnitine (41.6±2.9% and 54.2±4.9%) and inositol (34.9±2.0% and 47.3±2.2%) caused insignificant increases in CASA and total motility in comparison to the other groups. All of the antioxidants at 2.5 and 7.5 mM resulted in lower sperm with damaged DNA than that of control, thus reducing the DNA damage (P < 0.05). No significant differences were observed in CASA progressive motility and sperm motion characteristics among the groups. In fertility results based on 59-day non-returns, no significant differences were observed in non-return rates among groups. As regards biochemical parameters, supplementation with antioxidants did not significantly affect LPO and total GSH levels in comparison to the control group (P > 0.05). The maintenance of AOP level in methionine 2.5 mM was demonstrated to be higher (5.06±0.38 mM) than that of control (0.96±0.29 mM) following the freeze-thawing (P < 0.001). Supplementation with these antioxidants prior to the cryopreservation process protected the DNA integrity against the cryodamage. Furthermore, future research should focus on the molecular mechanisms of the antioxidative effects of the antioxidants methionine, carnitine and inositol during cryopreservation.

Influence of methionine and dithioerythritol on sperm motility, lipid peroxidation and antioxidant capacities during liquid storage of ram semen.

The aim of this study was to investigate the effects of methionine and dithioerythritol, added to the Tris extender, on ram sperm motility and LPO (lipid peroxidation) and antioxidant capacities during liquid storage up to 72 h at 5°C. Ejaculates collected from five Merino rams, were evaluated and pooled at 37°C. This study included two experiments. In experiment 1, each pooled ejaculate was divided into four equal aliquots and diluted (37°C) with the base extender, containing 0 (control), 1, 2 and 4 mM methionine, at a final concentration of approximately 4×10(8)sperms/ml (single step dilution), in a 15-ml plastic centrifuge tube. In experiment 2, dithioerythritol, at concentrations of 0 (control), 0.5, 1 and 2 mM, was used as an additive in the extender, and the procedure explained above was applied for the division of aliquots and the dilution of semen. Diluted semen samples were kept in glass tubes and cooled from 37 to 5°C in a cold cabinet, and maintained at 5°C. Sperm motility and LPO and total glutathione (GSH) and glutathione peroxidase (GPx) capacities were determined at 5°C for periods of 0, 24, 48 and 72 h of liquid storage. The extender supplemented with 1 mM methionine led to higher motility percentages (77.0±1.2%), in comparison to the control group (66.0±4.9%), during 72 h of liquid storage (P<0.05). As regards dithioerythritol, it did not statistically improve the motility rates for any of the storage times at 5°C. In biochemical assays, differences in LPO levels between the groups with antioxidants and the control groups were not statistically significant. Compared to the control group, no significant difference was observed in GSH and GPx activities following the addition of methionine, during 72 h of storage. Total GSH and GPx activities did not increase significantly upon supplementation with 0.5 and 1 mM of dithioerythritol, compared to the control group, at any of the time points (P>0.05). Dithioerythritol at 2 mM led (P<0.01) to elevating GSH activity, compared to the control group, during 72 h of liquid storage. GPx activity was approximately 10 times higher for 2 mM of dithioerythritol (P<0.001), compared to that of the control group at all time points. The question regarding the sustainability of sperm survival, LPO and antioxidant capacities following liquid storage of semen remains unanswered. Further studies are required for a better understanding of the biochemical changes and to obtain more information on the determination of lipid peroxidation and antioxidant capacities during cooled storage of ram semen.

Comparison of oxidative stress and antioxidant capacity before and after running exercises in both sexes.

BACKGROUND: It has been difficult to determine, from the published literature, whether men or women have higher levels of exercise-induced oxidative stress. OBJECTIVE: The aim of this study was to compare variations between the sexes in lipid hydroperoxide (LPO), superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), and lactate dehydrogenase (LDH) after 3 different running exercises performed at the same speed. METHODS: Eligible participants were healthy university students of both sexes. The participants performed running exercise tests at distances of 800, 1500, and 3000 m at a speed of 10 km/h. Blood samples were taken from the participants just before and immediately after the running activities to determine LPO, SOD, CAT, GR, and LDH, and these measures were compared both before and after exercise and between the sexes. RESULTS: A total of 17 young and healthy, but not physically trained, students (n = 8 men; mean age, 22.00 years; n = 9 women; mean age, 21.78 years) participated in this study. Height, weight, and maximum oxygen consumption values were significantly higher in men than in women (P = 0.01). Significant gender effects were found in LPO levels at 3000 m (F = 5.51; P = 0.03) and in SOD activity at 800 m (F = 7.92; P = 0.01) and 3000 m (F = 6.05; P = 0.03). CAT activity also differed between the sexes at 800 m (F = 15.67; P = 0.01) and 1500 m (F = 6.55; P = 0.02). However, no significant gender-time interaction effect was observed for any measurement at the 800-, 1500-, and 3000-m distances. CONCLUSIONS: Changes in LPO, SOD, and CAT activities at different running distances were not different between men and women over time because of a nonsignificant gender-time interaction. With regard to changes in oxidative stress, men and women had similar responses to exercise at the same absolute workload, despite significant differences in physical characteristics.