褪黑素对绒山羊精液冷冻保存效果的影响
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摘要
为了研究褪黑素对绒山羊精液冷冻保存效果的影响,在冷冻稀释液中外源补充不同浓度(0.125,0.250,0.500,1.000 mg/mL)的褪黑素进行冷冻试验,冷冻解冻采用计算机辅助精液分析系统(CASAS)和流式细胞仪分别检测精子活率、质膜完整率、顶体完整率以及精子细胞中活性氧(ROS)的含量。结果表明:在冷冻稀释液中添加褪黑素能够显著降低精子细胞中ROS的含量,而且在冷冻稀释液中添加浓度为0.500 mg/mL的试验组绒山羊精子活率(53.3%)、质膜完整率(44.6%)、顶体完整率(77.9%)均显著高于空白对照组(47.8%、36.3%和53.2%)(P<0.05)。说明在绒山羊精液冷冻稀释液中添加0.500 mg/mL褪黑素能够显著提高绒山羊精子抗氧化能力和冷冻保存效果。
In this sdudy,different concentrations of melatonin(0.125, 0.250, 0.500, 1.000 mg/mL) were supplemented in frozen diluent to investigate the effect of melatonin on the cryopreservation of cashmere goat's semen. The analysis system(CASAS) and flow cytometry were used to detect sperm motility, plasma membrane integrity, acrosome integrity, and reactive oxygen species(ROS) in sperm cells after thawing. The results show that the addition of melatonin in the freeze-diluted solution significantly reduced the level of ROS in the sperm. The sperm viability(53.3%), plasm integrity rate(44.6%), intact acrosome rate(77.9%) were significantly higher than that of the control group(47.8%, 36.3% and 53.2%,P<0.05).It indicats that the addition of 0.500 mg/mL melatonin to the frozen dilution of cashmere goat semen can significantly improve the antioxidant capacity and cryopreservation effect of spermatozoa.
In this sdudy,different concentrations of melatonin(0.125, 0.250, 0.500, 1.000 mg/mL) were supplemented in frozen diluent to investigate the effect of melatonin on the cryopreservation of cashmere goat's semen. The analysis system(CASAS) and flow cytometry were used to detect sperm motility, plasma membrane integrity, acrosome integrity, and reactive oxygen species(ROS) in sperm cells after thawing. The results show that the addition of melatonin in the freeze-diluted solution significantly reduced the level of ROS in the sperm. The sperm viability(53.3%), plasm integrity rate(44.6%), intact acrosome rate(77.9%) were significantly higher than that of the control group(47.8%, 36.3% and 53.2%,P<0.05).It indicats that the addition of 0.500 mg/mL melatonin to the frozen dilution of cashmere goat semen can significantly improve the antioxidant capacity and cryopreservation effect of spermatozoa.
引文
[1] AITKEN R J.Reactive oxygen species as mediators of sperm capacitation and pathological damage[J]. Mol Reprod Dev,2017,84(10):1039-1052.
[2] AITKEN R J.Free radicals,lipid peroxidation and sperm function[J].Reprod Fertil Dev,1995,7(4):659-668.
[3] ROSSI T,MAZZILIL F,DELFINO M,et al.Improved human sperm recovery using superoxide dismutase and catalase supplementation in semen cryopreservation procedure[J].Cell Tissue Bank,2001,2(1):9-13.
[4] NADJARZADEH A,SHIDFAR F,AMIRJANNATI N,et al.Effect of coenzyme Q10 supplementation on antioxidant enzymes activity and oxidative stress of seminal plasma:a double-blind randomised clinical trial[J].Andrologia,2014,46(2):177-183.
[5] GADEA J,GARCíIA-VAZQUEZ F,MATáAS C,et al.Cooling and freezing of boar spermatozoa: supplementation of the freezing media with reduced glutathione preserves sperm function[J].J Androl,2005,26(3):396-404.
[6] RODRIGUEZ C,MAYO J C,SAINZ R M,et al.Regulation of antioxidant enzymes:a significant role for melatonin[J].J Pineal Res,2004,36(1):1-9.
[7] JANG H Y,KIM Y H,YANG B K,et al.Ameliorative effects of melatonin against hydrogen peroxide-induced oxidative stress on boar sperm characteristics and subsequent in vitro embryo development[J].Reprod Domest Anim,2010,45(6):943-950.
[8] 李博玲.在猪精液冷冻稀释液中添加褪黑素、谷胱甘肽、Vc对冷冻保存效果的影响[D].雅安:四川农业大学,2009.
[9] CHEN X J,ZHANG Y,JIA G X,et al.Effect of melatonin supplementation on cryopreserved sperm quality in mouse[J].Cryo Letters,2016,37(2):115-122.
[10] SUCCU S,BERLINGUER F,PASCIU V,et al.Melatonin protects ram spermatozoa from cryopreservation injuries in a dose-dependent manner[J].J Pineal Res,2011,50(3):310-318.
[11] 李晓霞,曹平华,禹学礼,等.添加褪黑素对牛精液品质的影响[J].中国畜牧兽医,2013,40(4):177-179.
[12] ASHRAFI I,KOHRAM H,ARDABILI F F.Antioxidative effects of melatonin on kinetics,microscopic and oxidative parameters of cryopreserved bull spermatozoa[J].Anim Reprod Sci,2013,139(1/2/3/4):25-30.
[13] 吴帅帅,李楠,付静涛,等.褪黑素对马精液冷冻保存效果影响的研究[C]//中国畜牧兽医学会马学分会成立大会学术论文集,北京:中国畜牧兽医学会马学分会,2014.
[14] 刘琦.不同精子质膜保护剂对猪精液常温保存效果的影响[D].杨凌:西北农林科技大学,2015.
[15] 杨威.三种抗氧化剂对猪精子冷冻保存效果的影响[D].杨凌:西北农林科技大学,2015.
[16] CARVAGAL G,CUELLO C,AUIZ M,et al.Effects of centrifugation before freezing on boar sperm cryosurvival[J].J Androl,2004,25(3):389-396.
[17] 商学军,黄宇烽,叶章群,等.褪黑素在活性氧致精子线粒体功能损伤中的保护作用[J].中华男科学杂志,2004,10(8):604-607.
[18] RODRIGUEZ C,MAYO J C,SAINZ R M,et al.Regulation of antioxidant enzymes:a significant role for melatonin[J].J Pineal Res,2004,36(1):1-9.
[19] 赵祥杰,李玉田,孙爽,等.猪精液颗粒冷冻技术的研究[J].东北农业大学学报,2008,39(3):70-75.
[20] LóPEZ A,GARCíAA J,ESCAMES G,et al.Melatonin protects the mitochondria from oxidative damage reducing oxygen consumption,membrane potential,and superoxide anion production[J].J Pineal Res,2009,46(2):188-198.
[21] 陈晓丽,朱化彬,郝海龙,等.猪精子冷冻损伤研究进展[J].中国生物工程杂志,2010,30(7):86-91.
[22] MAYO J C,SAINZ R M,ANTOLI,et al.Melatonin regulation of antioxidant enzyme gene expression[J].Cell Mol Life Sci,2002,59(10):1706-1713.
[23] 孙业清.褪黑素对牛精液品质、卵母细胞体外成熟及体外发育能力影响的研究[D].兰州:甘肃农业大学,2016.
[24] 杜立银,曹少先,郭炜,等.褪黑素在猪精液保存中的抗氧化作用[J].江苏农业学报,2009,25(2):315-319.
[2] AITKEN R J.Free radicals,lipid peroxidation and sperm function[J].Reprod Fertil Dev,1995,7(4):659-668.
[3] ROSSI T,MAZZILIL F,DELFINO M,et al.Improved human sperm recovery using superoxide dismutase and catalase supplementation in semen cryopreservation procedure[J].Cell Tissue Bank,2001,2(1):9-13.
[4] NADJARZADEH A,SHIDFAR F,AMIRJANNATI N,et al.Effect of coenzyme Q10 supplementation on antioxidant enzymes activity and oxidative stress of seminal plasma:a double-blind randomised clinical trial[J].Andrologia,2014,46(2):177-183.
[5] GADEA J,GARCíIA-VAZQUEZ F,MATáAS C,et al.Cooling and freezing of boar spermatozoa: supplementation of the freezing media with reduced glutathione preserves sperm function[J].J Androl,2005,26(3):396-404.
[6] RODRIGUEZ C,MAYO J C,SAINZ R M,et al.Regulation of antioxidant enzymes:a significant role for melatonin[J].J Pineal Res,2004,36(1):1-9.
[7] JANG H Y,KIM Y H,YANG B K,et al.Ameliorative effects of melatonin against hydrogen peroxide-induced oxidative stress on boar sperm characteristics and subsequent in vitro embryo development[J].Reprod Domest Anim,2010,45(6):943-950.
[8] 李博玲.在猪精液冷冻稀释液中添加褪黑素、谷胱甘肽、Vc对冷冻保存效果的影响[D].雅安:四川农业大学,2009.
[9] CHEN X J,ZHANG Y,JIA G X,et al.Effect of melatonin supplementation on cryopreserved sperm quality in mouse[J].Cryo Letters,2016,37(2):115-122.
[10] SUCCU S,BERLINGUER F,PASCIU V,et al.Melatonin protects ram spermatozoa from cryopreservation injuries in a dose-dependent manner[J].J Pineal Res,2011,50(3):310-318.
[11] 李晓霞,曹平华,禹学礼,等.添加褪黑素对牛精液品质的影响[J].中国畜牧兽医,2013,40(4):177-179.
[12] ASHRAFI I,KOHRAM H,ARDABILI F F.Antioxidative effects of melatonin on kinetics,microscopic and oxidative parameters of cryopreserved bull spermatozoa[J].Anim Reprod Sci,2013,139(1/2/3/4):25-30.
[13] 吴帅帅,李楠,付静涛,等.褪黑素对马精液冷冻保存效果影响的研究[C]//中国畜牧兽医学会马学分会成立大会学术论文集,北京:中国畜牧兽医学会马学分会,2014.
[14] 刘琦.不同精子质膜保护剂对猪精液常温保存效果的影响[D].杨凌:西北农林科技大学,2015.
[15] 杨威.三种抗氧化剂对猪精子冷冻保存效果的影响[D].杨凌:西北农林科技大学,2015.
[16] CARVAGAL G,CUELLO C,AUIZ M,et al.Effects of centrifugation before freezing on boar sperm cryosurvival[J].J Androl,2004,25(3):389-396.
[17] 商学军,黄宇烽,叶章群,等.褪黑素在活性氧致精子线粒体功能损伤中的保护作用[J].中华男科学杂志,2004,10(8):604-607.
[18] RODRIGUEZ C,MAYO J C,SAINZ R M,et al.Regulation of antioxidant enzymes:a significant role for melatonin[J].J Pineal Res,2004,36(1):1-9.
[19] 赵祥杰,李玉田,孙爽,等.猪精液颗粒冷冻技术的研究[J].东北农业大学学报,2008,39(3):70-75.
[20] LóPEZ A,GARCíAA J,ESCAMES G,et al.Melatonin protects the mitochondria from oxidative damage reducing oxygen consumption,membrane potential,and superoxide anion production[J].J Pineal Res,2009,46(2):188-198.
[21] 陈晓丽,朱化彬,郝海龙,等.猪精子冷冻损伤研究进展[J].中国生物工程杂志,2010,30(7):86-91.
[22] MAYO J C,SAINZ R M,ANTOLI,et al.Melatonin regulation of antioxidant enzyme gene expression[J].Cell Mol Life Sci,2002,59(10):1706-1713.
[23] 孙业清.褪黑素对牛精液品质、卵母细胞体外成熟及体外发育能力影响的研究[D].兰州:甘肃农业大学,2016.
[24] 杜立银,曹少先,郭炜,等.褪黑素在猪精液保存中的抗氧化作用[J].江苏农业学报,2009,25(2):315-319.