甘氨酸对H_2O_2诱导氧化应激的小鼠胚胎抗氧化能力的影响
摘要
本实验旨在探究甘氨酸(Gly)对H_2O_2诱导氧化应激的小鼠胚胎抗氧化能力的影响。实验分为对照组、H_2O_2处理组和H_2O_2+Gly处理组,对照组胚胎不经H_2O_2处理培养于M16培养液中;其他2组胚胎均置于25μmol/L的H_2O_2中孵育30 min,然后,H_2O_2处理组和H_2O_2+Gly处理组分别在M16培养液和最佳浓度Gly的培养液中进行培养。通过2',7'-二氯荧光素二乙酸盐(DCHFDA)法测定胚胎内部的活性氧水平和巢式PCR法检测内源抗氧化酶基因的表达。结果表明:胚胎经不同浓度Gly处理后,4 mmol/L Gly处理组的4-cell发育率(P>0.05)和囊胚发育率(P<0.05)均高于对照组;氧化应激胚胎经4 mmol/L的Gly处理后,胚胎的4-cell发育率(P<0.01)和囊胚发育率(P<0.05)显著高于H_2O_2处理组;H_2O_2+Gly处理组的活性氧水平显著低于H_2O_2处理组(P<0.05);Gly能上调CAT(P<0.05)和SOD1(P<0.01)基因的表达,而对SOD2基因的表达无影响(P>0.05)。综上可知,添加Gly可以有效提高胚胎的抗氧化能力,在胚胎体外培养过程中添加Gly的最适宜浓度为4 mmol/L。
引文
[1]Dalvitg C,Cetica P D,Pintos L N,et al.Reactive oxygen species in bovine embryo in vitro production[J].Biocell,2005,29(2):209-212.
[2]Scherz-Shouval R,Elazar Z.Regulation of autophagy by ROS:physiology and pathology[J].Trends Biochem Sci,2011,36(1):30-38.
[3]Gupta S,Malhotra N,Sharma D,et al.Oxidative stress and its role in female infertility and assisted reproduction:Clinical implications[J].Int J Fertil Steril,2009,2(4):147-164.
[4]Sabatini S E,Juárez A B,Eppis M R,et al.Oxidative stress and antioxidant defenses in two green microalgae exposed to copper[J].Ecotoxicol Environ Saf,2009,72(4):1200-1206.
[5]潘晓菲,谭初兵,时丽丽,等.Nrf2/ARE信号通路及其相关药物研究进展[J].药物评价研究,2013,36(1):54-58.
[6]Liang M,Wang Z,Li H,et al.L-Arginine induces antioxidant response to prevent oxidative st ress via stimulation of glutathione synthesis and activation of Nrf2 pathway[J].Food Chem Toxicol,2018(18):30178-30179.
[7]Rubio V,Zhang J,Valverde M,et al.Essential role of Nrf2 in protection against hydroquinone-and benzoquinone-induced cytotoxicity[J].Toxicol In Vitro,2011,25(2):521-529.
[8]Xu F L,You H B,Li X H,et al.Glycine attenuates endotoxin induced liver injury by down regulation TLR4 signaling in Kupffer cells[J].Am J Surg,2008,196(1):139-148.
[9]Yamashina S,Ikejima K,Enomoto N,et al.Glycine as a therapeutic immuno-nutrient for alcoholic liver disease[J].Alcohol Clin Exp Res,2005,29(11 Suppl):162-165.
[10]袁晓娟,吴希茜.甘氨酸的生理作用与应用[J].饮料工业,2011,7(2):1007-7871.
[11]Nonogaki T,Noda Y,Narimoto K,et al.Effects of superoxide dismutase on mouse in vitro fertilization and embryo culture system[J].Assist Reprod Gen,1992,9(3):274-280.
[12]胡德宝,张也,张日欣,等.WY14643对小鼠早期胚胎抗氧化损伤的影响[J].畜牧兽医学报,2014,45(4):547-552.
[13]Mc Car ty M F,O'Keefe J H,Di Nicolantonio J J.Dietar y glycine is rate-limiting for glutathione synthesis and may have broad potential for health protection[J].Ochsner J,2018,18(1):81-87.
[14]Lane M.Mechanisms for managing cellular and homeostatic stress in vitro[J].Theriogenology,2001,55(1):225-236.
[15]Cao X Y,Rose J,Wang S Y,et al.Glycine increases preimpla nt at ion development of mou se oocy tes follow i ng vitrifi cation at the germinal vesicle stage[J].Sci Rep,2016,6:37262.
[16]Mito T,Yoshioka K,Yamashita S,et al.Glucose and glycine synergistically enhance the in vitro development of porcine blastocysts in a chemically defined medium[J].Reprod Fertil Dev,2012,24(3):443-500.
[17]Guerin P,Mouatassim S EI,Menezo Y.Oxidative stress and protection against reactive oxygen species in the preimplantation embryo and its surroundings[J].Hum Reprod Update,2001,7(2):175-189.
[18]Takahashi M.Oxidative stress and redox regulation on in vitro development of mammalian embryos[J].J Reprod Develop,2012,58(1):1-9.
[19]Osbur n W O,Kensler T W.Nrf 2 sig naling:an adaptive response pathway for protection against environmental toxic insults[J].Mutat Res,2008,659(1-2):31-39.
[20]Shi X,Zhou B.The role of Nrf 2 and MAPK pathways in PFOS-Induced oxidative stress in zebraf ish embr yos[J].Toxucol Sci,2010,115(2):391-400.
[2]Scherz-Shouval R,Elazar Z.Regulation of autophagy by ROS:physiology and pathology[J].Trends Biochem Sci,2011,36(1):30-38.
[3]Gupta S,Malhotra N,Sharma D,et al.Oxidative stress and its role in female infertility and assisted reproduction:Clinical implications[J].Int J Fertil Steril,2009,2(4):147-164.
[4]Sabatini S E,Juárez A B,Eppis M R,et al.Oxidative stress and antioxidant defenses in two green microalgae exposed to copper[J].Ecotoxicol Environ Saf,2009,72(4):1200-1206.
[5]潘晓菲,谭初兵,时丽丽,等.Nrf2/ARE信号通路及其相关药物研究进展[J].药物评价研究,2013,36(1):54-58.
[6]Liang M,Wang Z,Li H,et al.L-Arginine induces antioxidant response to prevent oxidative st ress via stimulation of glutathione synthesis and activation of Nrf2 pathway[J].Food Chem Toxicol,2018(18):30178-30179.
[7]Rubio V,Zhang J,Valverde M,et al.Essential role of Nrf2 in protection against hydroquinone-and benzoquinone-induced cytotoxicity[J].Toxicol In Vitro,2011,25(2):521-529.
[8]Xu F L,You H B,Li X H,et al.Glycine attenuates endotoxin induced liver injury by down regulation TLR4 signaling in Kupffer cells[J].Am J Surg,2008,196(1):139-148.
[9]Yamashina S,Ikejima K,Enomoto N,et al.Glycine as a therapeutic immuno-nutrient for alcoholic liver disease[J].Alcohol Clin Exp Res,2005,29(11 Suppl):162-165.
[10]袁晓娟,吴希茜.甘氨酸的生理作用与应用[J].饮料工业,2011,7(2):1007-7871.
[11]Nonogaki T,Noda Y,Narimoto K,et al.Effects of superoxide dismutase on mouse in vitro fertilization and embryo culture system[J].Assist Reprod Gen,1992,9(3):274-280.
[12]胡德宝,张也,张日欣,等.WY14643对小鼠早期胚胎抗氧化损伤的影响[J].畜牧兽医学报,2014,45(4):547-552.
[13]Mc Car ty M F,O'Keefe J H,Di Nicolantonio J J.Dietar y glycine is rate-limiting for glutathione synthesis and may have broad potential for health protection[J].Ochsner J,2018,18(1):81-87.
[14]Lane M.Mechanisms for managing cellular and homeostatic stress in vitro[J].Theriogenology,2001,55(1):225-236.
[15]Cao X Y,Rose J,Wang S Y,et al.Glycine increases preimpla nt at ion development of mou se oocy tes follow i ng vitrifi cation at the germinal vesicle stage[J].Sci Rep,2016,6:37262.
[16]Mito T,Yoshioka K,Yamashita S,et al.Glucose and glycine synergistically enhance the in vitro development of porcine blastocysts in a chemically defined medium[J].Reprod Fertil Dev,2012,24(3):443-500.
[17]Guerin P,Mouatassim S EI,Menezo Y.Oxidative stress and protection against reactive oxygen species in the preimplantation embryo and its surroundings[J].Hum Reprod Update,2001,7(2):175-189.
[18]Takahashi M.Oxidative stress and redox regulation on in vitro development of mammalian embryos[J].J Reprod Develop,2012,58(1):1-9.
[19]Osbur n W O,Kensler T W.Nrf 2 sig naling:an adaptive response pathway for protection against environmental toxic insults[J].Mutat Res,2008,659(1-2):31-39.
[20]Shi X,Zhou B.The role of Nrf 2 and MAPK pathways in PFOS-Induced oxidative stress in zebraf ish embr yos[J].Toxucol Sci,2010,115(2):391-400.