Uterine nitric oxide levels and isofluopredone treatment effect in mares susceptible to persistent post-breeding endometritis

Transient endometritis is a normal consequence of breeding and results from uterine contamination with both semen and bacteria. The modulation of the inflammatory response with the use of isoflupredone has been proposed as efficient for the treatment of endometritis by increasing pregnancy rates. The aim of the current study was to determine the effects of isoflupredone on nitric oxide (NO) levels in uterine samples from mares susceptible to persistent postbreeding endometritis, presenting or not the infectious process. Seven consecutive estrous cycles were induced in 11 mares, being the first one used as control (no treatment). All mares were submitted to the following four treatments: treatment 1: control, treatment 2: glucocorticoid (GC) treatment (20 mg isoflupredone acetate) every 12 h, for three consecutive days, treatment 3: infected treatment (intrauterine infusion of 1x109 CFU/ml Streptococcus equi subsp. zooepidemicus), treatment 4: combination of GC + infected treatment (infusion of bacteria 24 h after the first GC treatment). At 12 h after the end of each treatment, uterine samples were collected by flushing and NO was determined. After nitrate reduction, total nitrite was determined by spectrophotometer. No significant differences on nitric oxide concentration were verified by analysis of variance in the different experimental groups. It is concluded that the use of isoflupredone did not alter the nitric oxide concentration in uterine flushing’s from susceptible mares 12 h after treatment.

Uterine nitric oxide levels and isofluopredone treatment effect in mares susceptible to persistent post-breeding endometritis

Transient endometritis is a normal consequence of breeding and results from uterine contamination with both semen and bacteria. The modulation of the inflammatory response with the use of isoflupredone has been proposed as efficient for the treatment of endometritis by increasing pregnancy rates. The aim of the current study was to determine the effects of isoflupredone on nitric oxide (NO) levels in uterine samples from mares susceptible to persistent postbreeding endometritis, presenting or not the infectious process. Seven consecutive estrous cycles were induced in 11 mares, being the first one used as control (no treatment). All mares were submitted to the following four treatments: treatment 1: control, treatment 2: glucocorticoid (GC) treatment (20 mg isoflupredone acetate) every 12 h, for three consecutive days, treatment 3: infected treatment (intrauterine infusion of 1x109 CFU/ml Streptococcus equi subsp. zooepidemicus), treatment 4: combination of GC + infected treatment (infusion of bacteria 24 h after the first GC treatment). At 12 h after the end of each treatment, uterine samples were collected by flushing and NO was determined. After nitrate reduction, total nitrite was determined by spectrophotometer. No significant differences on nitric oxide concentration were verified by analysis of variance in the different experimental groups. It is concluded that the use of isoflupredone did not alter the nitric oxide concentration in uterine flushings from susceptible mares 12 h after treatment.(AU)

Glutathione for the freezing of cooled equine semen, using different protocols

The aim of this study was to evaluate the effect of in vitro addition of glutathione in four different concentrations compared to the control group, using two different freezing systems: controlled-rate (AS) and manual (MS) freezer. The parameters evaluated were motility, strength, plasmatic and acrosomal membrane integrity of spermatozoa from twelve stallions. Ejaculates from stallions were collected three times per week, during four weeks. Gel-free semen was diluted in skim milk extender and cooled (16ºC) f or 24 h. After cooling, extended semen was centrifuged at 600 xg for 10 min. The supernatant was removed and sperm pellets were re-suspended using the freezing extender with different glutathione concentration s ( 0, 2.5 mM, 5 mM, 7.5 mM , and 10 mM). Samples were then packed into 0.5 m l straws, which were divided into two parts: one for cryopreservation in a controlled-rate freezer (AS) and the other for a manual system (MS). In this study, the control group showed higher motility and better membrane integrity within treatments for the AS (P < 0.05). Furthermore, when an MS group was evaluated, the 2.5 mM glutathione group demonstrated better preservation for total motility and plasmatic membrane integrity. However, concentrations higher than 2.5 mM were deleterious to the spermatozoa in both controlled and manual freezing systems.

Glutathione for the freezing of cooled equine semen, using different protocols

The aim of this study was to evaluate the effect of in vitro addition of glutathione in four different concentrations compared to the control group, using two different freezing systems: controlled-rate (AS) and manual (MS) freezer. The parameters evaluated were motility, strength, plasmatic and acrosomal membrane integrity of spermatozoa from twelve stallions. Ejaculates from stallions were collected three times per week, during four weeks. Gel-free semen was diluted in skim milk extender and cooled (16ºC) f or 24 h. After cooling, extended semen was centrifuged at 600 xg for 10 min. The supernatant was removed and sperm pellets were re-suspended using the freezing extender with different glutathione concentration s ( 0, 2.5 mM, 5 mM, 7.5 mM , and 10 mM). Samples were then packed into 0.5 m l straws, which were divided into two parts: one for cryopreservation in a controlled-rate freezer (AS) and the other for a manual system (MS). In this study, the control group showed higher motility and better membrane integrity within treatments for the AS (P < 0.05). Furthermore, when an MS group was evaluated, the 2.5 mM glutathione group demonstrated better preservation for total motility and plasmatic membrane integrity. However, concentrations higher than 2.5 mM were deleterious to the spermatozoa in both controlled and manual freezing systems.(AU)

The effect of sperm selection by Percoll or swim-up on the sex ratio of in vitro produced bovine embryos

The goal of this investigation was to determine the percentage of male and female bovine embryos produced after oocyte fertilization with Percoll density gradient centrifugation or with self-migration (swim-up) selected semen. In Experiment 1, sperm selection was performed by 90-45% discontinuous Percoll density gradient centrifugation (T1) and swim-up (T2). InExperiment 2, in addition to the discontinuous gradient, a 67.5% continuous gradient, and 5 or 10 min centrifugation were used. Four treatment groups were defined (TI = continuous, 5 min, TII = discontinuous, 5 min, TIII = continuous, 10 min and TIV = discontinuous,10 min). Embryos were sexed using PCR. In Experiment 1,Percoll density gradient centrifugation (n = 185) resulted in 48.6% (n = 90) male and 51.4% (n = 95) female embryos, and swim-up (n = 142) in 58.4% (n = 83) male and 41.6% (n = 59) female embryos. In Experiment 2, the percentages of male and female embryos obtained in TI (n = 93), TII (n = 70), TIII (n = 82) and TIV (n = 82) were 49.5% (n = 46) and 50.5% (n = 47), 57.1% (n = 40) and 42.9% (n = 30), 36.6% (n = 30) and 63.4% (n = 52) and 48.8% (n = 40) and 51.2% (n = 42), respectively. Swim-up (T2) and continuous Percoll gradient centrifugation for 10 min (TIII) showed a deviation toward males (P = 0.044) and females (P = 0.015), respectively, when compared to the 50% expected percentage of each gender. There was no difference in male and female percentages in the other treatment groups from Experiments 1 and 2, when compared to the 50% expected percentage of each gender neither between treatment.(AU)

The effect of sperm selection by Percoll or swim-up on the sex ratio of in vitro produced bovine embryos

The goal of this investigation was to determine the percentage of male and female bovine embryos produced after oocyte fertilization with Percoll density gradient centrifugation or with self-migration (swim-up) selected semen. In Experiment 1, sperm selection was performed by 90-45% discontinuous Percoll density gradient centrifugation (T1) and swim-up (T2). InExperiment 2, in addition to the discontinuous gradient, a 67.5% continuous gradient, and 5 or 10 min centrifugation were used. Four treatment groups were defined (TI = continuous, 5 min, TII = discontinuous, 5 min, TIII = continuous, 10 min and TIV = discontinuous,10 min). Embryos were sexed using PCR. In Experiment 1,Percoll density gradient centrifugation (n = 185) resulted in 48.6% (n = 90) male and 51.4% (n = 95) female embryos, and swim-up (n = 142) in 58.4% (n = 83) male and 41.6% (n = 59) female embryos. In Experiment 2, the percentages of male and female embryos obtained in TI (n = 93), TII (n = 70), TIII (n = 82) and TIV (n = 82) were 49.5% (n = 46) and 50.5% (n = 47), 57.1% (n = 40) and 42.9% (n = 30), 36.6% (n = 30) and 63.4% (n = 52) and 48.8% (n = 40) and 51.2% (n = 42), respectively. Swim-up (T2) and continuous Percoll gradient centrifugation for 10 min (TIII) showed a deviation toward males (P = 0.044) and females (P = 0.015), respectively, when compared to the 50% expected percentage of each gender. There was no difference in male and female percentages in the other treatment groups from Experiments 1 and 2, when compared to the 50% expected percentage of each gender neither between treatment.