Catalase 첨가에 따른 돼지 정액 동결 및 융해 후 생존 정자에서 Hydrogen Peroxide의 감소 The Reduction of Hydrogen Peroxide in Viable Boar Sperm Cryopreserved in the Presence of Catalase원문보기
요 약: 정액 동결 과정은 활성산소종의 생성을 유발하며, 생성된 활성산소종은 정자의 손상을 일으키는 것으로 알려져 있다. 따라서 본 연구의 목적은 동결 과정 중 항산화 효소 중 하나인 catalase (CAT)를 첨가함으로써 융해 후 정자의 기능과 활성산소종의 수준에 미치는 효과를 알아보고자 하였다. 5마리 돼지에서 채취한 정액은 0 (대조군), 200, 400 U/mL CAT가 첨가되어 있는 동결 희석액으로 각각 동결하였다. 융해 후, 정자 운동성, 생존성, 정상 형태율, 형질막 온전성, 미토콘드리아 기능, 세포내 ROS를 평가하였다. CAT는 400 U/mL의 농도에서 전체 정자 운동성을 향상시켰지만 (P < 0.05), 전진 운동성, 생존성, 기형율, 형질막 온전성, 미토콘드리아 기능의 향상을 나타내지 않았다. 활성산소종의 평가에서, CAT는 융해된 생존 정자의 ${\cdot}O_2$의 감소에는 효과를 나타내지 않은 반면 $H_2O_2$를 감소시켰다(P < 0.05). 결론으로 CAT는 동결 및 융해된 정자의 질을 향상시키는 데 큰 효과를 나타내진 않았지만 생존 정자에서 $H_2O_2$을 제거함로써 생존정자의 산화적 손상을 감소시킬 수 있으리라 판단된다.
요 약: 정액 동결 과정은 활성산소종의 생성을 유발하며, 생성된 활성산소종은 정자의 손상을 일으키는 것으로 알려져 있다. 따라서 본 연구의 목적은 동결 과정 중 항산화 효소 중 하나인 catalase (CAT)를 첨가함으로써 융해 후 정자의 기능과 활성산소종의 수준에 미치는 효과를 알아보고자 하였다. 5마리 돼지에서 채취한 정액은 0 (대조군), 200, 400 U/mL CAT가 첨가되어 있는 동결 희석액으로 각각 동결하였다. 융해 후, 정자 운동성, 생존성, 정상 형태율, 형질막 온전성, 미토콘드리아 기능, 세포내 ROS를 평가하였다. CAT는 400 U/mL의 농도에서 전체 정자 운동성을 향상시켰지만 (P < 0.05), 전진 운동성, 생존성, 기형율, 형질막 온전성, 미토콘드리아 기능의 향상을 나타내지 않았다. 활성산소종의 평가에서, CAT는 융해된 생존 정자의 ${\cdot}O_2$의 감소에는 효과를 나타내지 않은 반면 $H_2O_2$를 감소시켰다(P < 0.05). 결론으로 CAT는 동결 및 융해된 정자의 질을 향상시키는 데 큰 효과를 나타내진 않았지만 생존 정자에서 $H_2O_2$을 제거함로써 생존정자의 산화적 손상을 감소시킬 수 있으리라 판단된다.
Semen cryopreservation induces the formation of reactive oxygen species (ROS), and the ROS cause sperm damage. We aimed to investigate the effects of the antioxidative enzyme catalase (CAT) on sperm quality and ROS during cryopreservation. Sperm rich fractions collected from five Duroc boars were cr...
Semen cryopreservation induces the formation of reactive oxygen species (ROS), and the ROS cause sperm damage. We aimed to investigate the effects of the antioxidative enzyme catalase (CAT) on sperm quality and ROS during cryopreservation. Sperm rich fractions collected from five Duroc boars were cryopreserved in freezing extender with (200 or 400 U/mL) or without CAT (control). After thawing, sperm motility, viability, normal morphology, plasma membrane integrity, mitochondrial function and intracellular ROS were evaluated. CAT significantly improved total sperm motility at a concentration of 400 U/mL (P < 0.05), but didn't improve progressive sperm motility, viability, morphological defects, plasma membrane integrity and mitochondrial function in frozen-thawed boar sperm. In evaluation of ROS, CAT had no effect on reduction in ${\cdot}O_2$, but scavenged $H_2O_2$ in viable frozen-thawed boar sperm at concentrations of 200 and 400 U/mL (P < 0.05). In conclusion, CAT was not enough to improve quality of frozen-thawed sperm, but can reduce $H_2O_2$ generation in viable boar sperm during cryopreservation.
Semen cryopreservation induces the formation of reactive oxygen species (ROS), and the ROS cause sperm damage. We aimed to investigate the effects of the antioxidative enzyme catalase (CAT) on sperm quality and ROS during cryopreservation. Sperm rich fractions collected from five Duroc boars were cryopreserved in freezing extender with (200 or 400 U/mL) or without CAT (control). After thawing, sperm motility, viability, normal morphology, plasma membrane integrity, mitochondrial function and intracellular ROS were evaluated. CAT significantly improved total sperm motility at a concentration of 400 U/mL (P < 0.05), but didn't improve progressive sperm motility, viability, morphological defects, plasma membrane integrity and mitochondrial function in frozen-thawed boar sperm. In evaluation of ROS, CAT had no effect on reduction in ${\cdot}O_2$, but scavenged $H_2O_2$ in viable frozen-thawed boar sperm at concentrations of 200 and 400 U/mL (P < 0.05). In conclusion, CAT was not enough to improve quality of frozen-thawed sperm, but can reduce $H_2O_2$ generation in viable boar sperm during cryopreservation.
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문제 정의
In conclusion, this study focused on the effect of CAT on ROS in viable boar sperm after cryopreservation. CAT led to a reduction in H2O2 in the viable frozen-thawed sperm.
Catalase (CAT) has been reported as a potential H2O2 detoxifier (3), but its ability to act as antioxidant in cryopreservation of boar sperm is not well known. Therefore, the aims of this study were to investigate the effects of CAT, as a supplement in sperm freezing extenders on sperm quality and ROS during cryopreservation of boar sperm.
제안 방법
All flow cytometry analyses were performed using a FACScalibur flow cytometer (Becton Dickinson, San José, CA, USA) equipped with a 15 mW air-cooled 488 nm argon-ion laser and Cell Quest Pro software (Becton Dickinson).
대상 데이터
Five fertile Duroc boars were used in this study. One ejaculation from each boar was collected using the gloved-hand method (n = 5).
데이터처리
The Shapiro-Wilk test was utilized to evaluate normality analysis. The non-parametric Friedman test was used for analysis of all data, and the Wilcoxon signed ranks test was used to calculate the difference between samples in cases showing significant differences with the Friedman test. Statistical significance was set at P < 0.
이론/모형
Five fertile Duroc boars were used in this study. One ejaculation from each boar was collected using the gloved-hand method (n = 5). Sperm rich fractions were extended (1:1 [v/v]) in BTS.
, Chicago, IL, USA). The Shapiro-Wilk test was utilized to evaluate normality analysis. The non-parametric Friedman test was used for analysis of all data, and the Wilcoxon signed ranks test was used to calculate the difference between samples in cases showing significant differences with the Friedman test.
The mean of six successive estimations was recorded as the final motility score. The viability of sperm was assessed by means of the eosin-nigrosin staining method (10). Viability was assessed by counting 200 sperm under a bright field microscope at × 400 magnification.
성능/효과
The percentage of motile and progressively motile sperm was determined by observing a minimum of 300 sperm, in at least six different fields under a bright field microscope at × 400 magnification.
후속연구
Further research is required to determine if the reduction in ethidium fluorescence by CAT is due to the H2O2-detecting ability of HE or ·O2 inactivation following removal of H2O2 by CAT.
However, it is still not clear that CAT provides viable boar sperm with sufficient protection from oxidative damage during cryopreservation. Further studies will be necessary to determine if the reduction of H2O2 in viable frozen-thawed sperm by CAT is sufficient to improve fertilizing ability after freeze-thawing.
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