精液冷冻贮藏

精液冷冻贮藏（Semen cryopreservation）是指以低温保存精子细胞活性的程序。精液可以透过深低温保存永久保存。若考虑以人类精液进行冷冻贮藏，之后又成功受孕的案例，据文献记载贮藏最长达24年^[1]。精液冷冻贮藏可以用在捐精（精子银行），可能捐精者和需要精子者相距距离较远，或是在捐精后一段时间才需要精子，若男性接受输精管切除术或是像化学疗法、放射线疗法等会影响生育能力的手术前，也可能会先将精液冷冻贮藏，保留日后再养育后代的可能性。在1954年时就已有用冷冻精液解冻后成功受孕的例子^[2]。

精液冷冻储藏技术广泛应用于男性不孕症患者，及畜牧业的人工繁殖上。对于即将进行化学治疗等细胞毒杀治疗的患者，该技术也能事先保存患者的生殖能力^[3]。

保存方法[编辑]

冷冻[编辑]

最常用的精液冷冻保护剂是甘油（占冷冻保护剂溶液的10%），而甘油溶液中也常加入蔗糖、其他的双糖或三糖。冷冻保护剂溶液会加入蛋黄或大豆卵磷脂，比较使用这两种物质，精液在解冻后，在精子活动力、变形程度、在体外和玻尿酸盐结合的能力以及其DNA的完整性上，两者没有显著的差异^[4]。

冷冻保护剂中的其他成分可以增加精子在冷冻后的存活率及受孕率。在冷冻贮藏前，用肝素结合蛋白来处理精子，可以减少精子的冷冻伤害，也减少自由基（Reactive oxygen species）的生成^[5]。在冷冻保护剂中加入神经成长因子（英语：nerve growth factor）可以减少精子的死亡速率，增加精子解冻后的活动力^[6]。若在冷冻前，用环糊精将胆固醇掺入精子细胞膜中，也可以增加精子活动力^[7]。

精液的冷冻会用控制速率的慢速冷却（深低温保存，简称SPF），另一种冷却方式是较新式的快速冷却，称为玻璃化（vitrification）。玻璃化的解冻后活动力以及冷冻存活比例都比慢速深低温保存要高^[8]。

解冻[编辑]

最理想的解冻温度是40 °C，不过解冻温度的细微调整对精子活动力、顶体状态、ATP含量以及DNA完整性的影响不大^[9]。精液冷冻后的解冻也有许多不同的技术，在Di Santo等人2012年的文献中都有提到^[10]。

再冷冻[编辑]

比较冷冻再解冻一次和三次的精液，在DNA片段化的风险差异不大。前提是精液和其原始的冷冻保护剂一起再冷冻，中间没有经过精子洗涤或其他类似程序，而用来进行人工受精（英语：assisted reproduction technology）前，会先用密度梯度离心（英语：density gradient centrifugation）或swim-up（英语：swim-up）进行分离^[11]。

对于精子品质的影响[编辑]

有些证据证实精子在冷冻后，单股断裂、DNA缩合（英语：DNA condensation）及片段化的比例增加，这也会增加后代DNA突变的风险。使用抗氧化剂以及良好的冷冻控温可能可以改善此情形^[12]。

在长期的追踪研究中，用冷冻后精液受孕的人和一般人比较，其先天性障碍或染色体畸变的比例相近，还没有证据支持用冷冻后精液受孕的人有较高的风险^[12]。

历史[编辑]

对于精液冷冻储藏的历史能够追溯到1776年，意大利人拉扎罗·斯帕兰札尼尝试使用冰雪保存精子^[3]。1949年克利斯朵夫·波尔吉利用甘油冷冻保护的特性^[13]，大幅改善了精液保存技术。自此，人们开始成功以冷冻储藏的精液，进行各种物种的人工受精^[3]。

参考资料[编辑]

^ Planer NEWS and Press Releases > Child born after 22 year semen storage using Planer controlled rate freezer 14/10/2004
^ Fatherhood After Death Has Now Been Proved Possible. Cedar Rapids Gazette. 1954-04-09.
^ ^3.0 ^3.1 ^3.2 Hezavehei, Maryam; Sharafi, Mohsen; Kouchesfahani, Homa Mohseni; Henkel, Ralf; Agarwal, Ashok; Esmaeili, Vahid; Shahverdi, Abdolhossein. Sperm cryopreservation: A review on current molecular cryobiology and advanced approaches. Reproductive BioMedicine Online. 2018-09, 37 (3): 327–339 [2020-02-19]. doi:10.1016/j.rbmo.2018.05.012. （原始内容存档于2022-06-18）（英语）.
^ Reed ML; et al. Soy lecithin replaces egg yolk for cryopreservation of human sperm without adversely affecting postthaw motility, morphology, sperm DNA integrity, or sperm binding to hyaluronate. Fertility and Sterility. 2009, 92 (5): 1787–1790. doi:10.1016/j.fertnstert.2009.05.026.
^ Patel, M., Gandotra, V. K., Cheema, R. S., Bansal, A. K., & Kumar, A. (2016). Seminal Plasma Heparin Binding Proteins Improve Semen Quality by Reducing Oxidative Stress during Cryopreservation of Cattle Bull Semen. Asian-Australasian Journal of Animal Sciences, 29(9), 1247–1255.
^ Saeednia S, Bahadoran H, Amidi F, Asadi MH, Naji M, Fallahi P, Nejad NA. Nerve growth factor in human semen: Effect of nerve growth factor on the normozoospermic men during cryopreservation process. Iranian Journal of Basic Medical Sciences. 2015, 18 (3): 292–299.
^ Purdy PH, Graham JK. Effect of cholesterol-loaded cyclodextrin on the cryosurvival of bull sperm. Cryobiology. 2004a, 48: 36–45. doi:10.1016/j.cryobiol.2003.12.001.
^ Vutyavanich T, Piromlertamorn W, Nunta S. Rapid freezing versus slow programmable freezing of human spermatozoa. Fertil. Steril. April 2010, 93 (6): 1921–8. PMID 19243759. doi:10.1016/j.fertnstert.2008.04.076.
^ Calamera JC, Buffone MG, Doncel GF, et al. Effect of thawing temperature on the motility recovery of cryopreserved human spermatozoa. Fertil. Steril. December 2008, 93 (3): 789–794. PMID 19059590. doi:10.1016/j.fertnstert.2008.10.021.
^ Di Santo M, Tarozzi N, Nadalini M, and Borini A. Human Sperm Cryopreservation: Update on Techniques, Effect on DNA Integrity, and Implications for ART (Review). Adv. Urology. 2012, 2012: 854837 [2019-07-12]. PMC 3238352 . PMID 22194740. doi:10.1155/2012/854837. （原始内容存档于2018-06-02）.
^ Thomson LK, Fleming SD, Barone K, Zieschang JA, Clark AM. The effect of repeated freezing and thawing on human sperm DNA fragmentation. Fertil Steril. March 2010, 93 (4): 1147–1156. PMID 19135665. doi:10.1016/j.fertnstert.2008.11.023.
^ ^12.0 ^12.1 Kopeika, J.; Thornhill, A.; Khalaf, Y. The effect of cryopreservation on the genome of gametes and embryos: principles of cryobiology and critical appraisal of the evidence. Human Reproduction Update. 2014, 21 (2): 209–227. ISSN 1355-4786. PMID 25519143. doi:10.1093/humupd/dmu063.
^ Polge, C.; Smith, A. U.; Parkes, A. S. Revival of Spermatozoa after Vitrification and Dehydration at Low Temperatures. Nature. 1949-10, 164 (4172): 666–666 [2020-02-19]. ISSN 1476-4687. doi:10.1038/164666a0. （原始内容存档于2022-06-18）（英语）.

[1] Planer NEWS and Press Releases > Child born after 22 year semen storage using Planer controlled rate freezer 14/10/2004

[2] Fatherhood After Death Has Now Been Proved Possible. Cedar Rapids Gazette. 1954-04-09.

[:0-3] 3.0 ^3.1 ^3.2 Hezavehei, Maryam; Sharafi, Mohsen; Kouchesfahani, Homa Mohseni; Henkel, Ralf; Agarwal, Ashok; Esmaeili, Vahid; Shahverdi, Abdolhossein. Sperm cryopreservation: A review on current molecular cryobiology and advanced approaches. Reproductive BioMedicine Online. 2018-09, 37 (3): 327–339 [2020-02-19]. doi:10.1016/j.rbmo.2018.05.012. （原始内容存档于2022-06-18）（英语）.

[4] Reed ML; et al. Soy lecithin replaces egg yolk for cryopreservation of human sperm without adversely affecting postthaw motility, morphology, sperm DNA integrity, or sperm binding to hyaluronate. Fertility and Sterility. 2009, 92 (5): 1787–1790. doi:10.1016/j.fertnstert.2009.05.026.

[5] Patel, M., Gandotra, V. K., Cheema, R. S., Bansal, A. K., & Kumar, A. (2016). Seminal Plasma Heparin Binding Proteins Improve Semen Quality by Reducing Oxidative Stress during Cryopreservation of Cattle Bull Semen. Asian-Australasian Journal of Animal Sciences, 29(9), 1247–1255.

[6] Saeednia S, Bahadoran H, Amidi F, Asadi MH, Naji M, Fallahi P, Nejad NA. Nerve growth factor in human semen: Effect of nerve growth factor on the normozoospermic men during cryopreservation process. Iranian Journal of Basic Medical Sciences. 2015, 18 (3): 292–299.

[7] Purdy PH, Graham JK. Effect of cholesterol-loaded cyclodextrin on the cryosurvival of bull sperm. Cryobiology. 2004a, 48: 36–45. doi:10.1016/j.cryobiol.2003.12.001.

[8] Vutyavanich T, Piromlertamorn W, Nunta S. Rapid freezing versus slow programmable freezing of human spermatozoa. Fertil. Steril. April 2010, 93 (6): 1921–8. PMID 19243759. doi:10.1016/j.fertnstert.2008.04.076.

[9] Calamera JC, Buffone MG, Doncel GF, et al. Effect of thawing temperature on the motility recovery of cryopreserved human spermatozoa. Fertil. Steril. December 2008, 93 (3): 789–794. PMID 19059590. doi:10.1016/j.fertnstert.2008.10.021.

[10] Di Santo M, Tarozzi N, Nadalini M, and Borini A. Human Sperm Cryopreservation: Update on Techniques, Effect on DNA Integrity, and Implications for ART (Review). Adv. Urology. 2012, 2012: 854837 [2019-07-12]. PMC 3238352 . PMID 22194740. doi:10.1155/2012/854837. （原始内容存档于2018-06-02）.

[11] Thomson LK, Fleming SD, Barone K, Zieschang JA, Clark AM. The effect of repeated freezing and thawing on human sperm DNA fragmentation. Fertil Steril. March 2010, 93 (4): 1147–1156. PMID 19135665. doi:10.1016/j.fertnstert.2008.11.023.

[KopeikaThornhill2014-12] 12.0 ^12.1 Kopeika, J.; Thornhill, A.; Khalaf, Y. The effect of cryopreservation on the genome of gametes and embryos: principles of cryobiology and critical appraisal of the evidence. Human Reproduction Update. 2014, 21 (2): 209–227. ISSN 1355-4786. PMID 25519143. doi:10.1093/humupd/dmu063.

[13] Polge, C.; Smith, A. U.; Parkes, A. S. Revival of Spermatozoa after Vitrification and Dehydration at Low Temperatures. Nature. 1949-10, 164 (4172): 666–666 [2020-02-19]. ISSN 1476-4687. doi:10.1038/164666a0. （原始内容存档于2022-06-18）（英语）.

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